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diff --git a/js/npm.js b/js/npm.js
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+++ b/js/npm.js
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+// This file is autogenerated via the `commonjs` Grunt task. You can require() this file in a CommonJS environment.
+require('../../js/transition.js')
+require('../../js/alert.js')
+require('../../js/button.js')
+require('../../js/carousel.js')
+require('../../js/collapse.js')
+require('../../js/dropdown.js')
+require('../../js/modal.js')
+require('../../js/tooltip.js')
+require('../../js/popover.js')
+require('../../js/scrollspy.js')
+require('../../js/tab.js')
+require('../../js/affix.js')
\ No newline at end of file
diff --git a/search-result.html b/search-result.html
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+ Welcome to Virtual Labs - A MHRD Govt of india Initiative
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+Welcome to Virtual Labs - A MHRD Govt of india Initiative
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+Welcome to Virtual Labs - A MHRD Govt of india Initiative
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+Welcome to Virtual Labs - A MHRD Govt of india Initiative
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+Welcome to Virtual Labs - A MHRD Govt of india Initiative
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+Welcome to Virtual Labs - A MHRD Govt of india Initiative
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+Welcome to Virtual Labs - A MHRD Govt of india Initiative
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+Welcome to Virtual Labs - A MHRD Govt of india Initiative
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+Welcome to Virtual Labs - A MHRD Govt of india Initiative
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+Welcome to Virtual Labs - A MHRD Govt of india Initiative
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+Welcome to Virtual Labs - A MHRD Govt of india Initiative
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+Welcome to Virtual Labs - A MHRD Govt of india Initiative
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+Welcome to Virtual Labs - A MHRD Govt of india Initiative
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+Welcome to Virtual Labs - A MHRD Govt of india Initiative
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+Welcome to Virtual Labs - A MHRD Govt of india Initiative
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diff --git a/src/lab/Acknowledgement.html b/src/lab/Acknowledgement.html
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diff --git a/src/lab/BackgroundPrinciple.html b/src/lab/BackgroundPrinciple.html
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+ Circular Dichroism
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+We acknowledge the efforts of the members of the development team:
+Antarip Halder, Bipin Singh, Swarnabha Sen, Jayaguru Panda, Ankit Baheti, Ishan Srivastava,Mohak Gupta, Sourav Chatterjee, Anubhooti and Parshvika Sharma.
Special thanks are due to Dr. Syed Maqbool Ahmed, PSO at the Central Instruments Laboratory, University of Hyderabad.
+Lab Developers:
+Dr. Abhijit Mitra (Professor, IIIT Hyderabad)
+Dr. Naveena Yanamala (ARS, University of Pittsburgh &Adjunct; Faculty, IIIT Hyderabad)
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diff --git a/src/lab/Feedback.html b/src/lab/Feedback.html
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Optical spectroscopic methods:
+Understanding structures of unknown molecules, or determining concentrations of known molecules, present in solutions, are extremely important for chemical and biochemical research. Optical spectroscopic methods constitute a nondestructive approach towards carrying out these tasks. In several of these methods, UV or visible light is allowed to pass through the solution and the required qualitative and quantitative information regarding the molecules in the solution is extracted by analyzing the changes in the properties of the transmitted light.
+
Depending on its composition, a medium can have two types of effect on the transmitted light. The medium can reduce the velocity and/or reduce the intensity of the transmitted light. The first effect relates to refraction (http://en.wikipedia.org/wiki/Refraction) due to the medium while the second relates to the absorbance (http://en.wikipedia.org/wiki/Absorbance) of the medium. Both these effects will depend on the nature and the number of the molecules present in the medium as well as the wavelength of the light. Naturally other experimental conditions such as temperature also influence the behavior of light. What is important to note is that, in some sense, optical spectroscopic methods can help us to 'see' and 'count' molecules.
++
The chiroptical phenomenon:
+When the molecules in the medium are chiral (http://en.wikipedia.org/wiki/Chirality_(chemistry)), then depending on whether the molecules are right handed or left handed, they affect individual interacting photons in different ways. These effects come under what are called chiroptical phenomena. Essentially, photons are associated with oscillating electric and magnetic fields, perpendicular to each other as well as to their direction of propagation. Since chiral molecules provide a chiral electrical environment they can affect the oscillating electric vectors, associated with interacting photons, in different ways. An ordinary beam of light consists of photons having their oscillating electric vectors uniformly distributed in all possible planes and, though the plane of the vectors associated with individual photons may change because of chiral interactions, the transmitted beam will still have a uniform distribution of the planes. So we can not track the chiroptical effects using ordinary light beams.
++
The phenomenon of polarization of light:
+
However, if we use an optically anisotropic medium, such as a nicol prism, we can filter a beam of ordinary light so that the transmitted beam consists only of photons with associated electric vectors aligned to a particular plane. You are advised to go through the pages provided at the link http://www.enzim.hu/~szia/cddemo/edemo1.htm, to understand electromagnetic waves and types of polarization. The following phenomena are explained in greater detail:
+-
+
- #Types of polarization (linear, circular) +
- #Superposition of waves +
- #Interference of waves +
+
Interaction of polarized light with matter:
++
We can detect chiroptical phenomena if we study the interaction of polarized light with matter. We know that interaction of light and matter leads to the phenomena of absorption and refraction. We have also seen above that any plane polarized light can be represented as a combination of two component linear polarized lights oriented horizontally and vertically, respectively, with respect to a reference plane. They can also be represented as a combination of two circularly polarized light components, namely right circularly polarized light (R-CPL) and left circularly polarized light (L-CPL). When plane polarized light travels through anisotropic materials, we can observe birefringence (differential refractive indices for the components respectively) and/or (usually both) dichroism (differential absorbance of the two components respectively). If the anisotropic material is chiral (optically active) then we observe differential refractive indices for R-CPL and L-CPL respectively, and results in a net rotation in the plane of polarization of the incident polarized light. In addition, we will also observe differential absorbanceâs for R-CPL and L-CPL respectively, leading to an elliptical polarization of the emergent light, referred to as circular dichroism. These phenomena are nicely explained in the link http://www.enzim.hu/~szia/cddemo/edemo9.htm. You are advised to go through the pages.
+Based on the principles discussed above, we have three spectroscopic methods for 'seeing' and 'counting' chiral molecules using polarized light
+Optical rotation:
+This involves the measurement of the rotation of linearly polarized light by the sample.
+
Optical rotary dispersion:
+ This involves the recording of the spectrum of optical rotation, i. e. understand the variation
+ of optical rotation as a function of wavelength.
+
Circular Dichroism:
This involves recording the difference in absorption of left and right circularly polarized light. Measurements carried out in the visible and ultra-violet region of the electro-magnetic spectrum monitor electronic transitions, and, if the molecule under study contains chiral chromophores then one CPL state will be absorbed to a greater extent than the other and the CD signal over the corresponding wavelengths will be non-zero. A circular dichroism signal can be positive or negative, depending on whether L-CPL is absorbed to a greater extent than R-CPL (CD signal positive) or to a lesser extent (CD signal negative). An example circular dichroism spectrum of a sample with multiple CD peaks is shown below, demonstrating how CD varies as a function of wavelength, and that a CD spectrum may exhibit both positive and negative peaks.
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diff --git a/src/lab/Introduction.html b/src/lab/Introduction.html
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Feedback
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diff --git a/src/lab/List ofExperiments.html b/src/lab/List ofExperiments.html
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Circular dichroism (CD) spectroscopy is a spectroscopic technique where the CD of molecules is measured over a range of wavelengths. CD spectroscopy is used extensively to study chiral molecules of all types and sizes, but it is in the study of large biological molecules where it finds its most important applications. A primary use is in analyzing the secondary structure or conformation of macromolecules, particularly proteins, and because secondary structure is sensitive to its environment, e.g. temperature or pH, circular dichroism can be used to observe how secondary structure changes with environmental conditions or on interaction with other molecules. Structural, kinetic and thermodynamic information about macromolecules can be derived from circular dichroism spectroscopy. +
+
Circular dichroism spectroscopy is particularly useful for:
+
-
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- Determining whether a protein is folded, and if so characterizing its secondary structure, tertiary structure, and the structural family to which it belongs +
- Comparing the structures of a protein obtained from different sources (e.g. species or expression systems) or comparing structures for different mutants of the same protein +
- Demonstrating comparability of solution conformation and/or thermal stability after changes in manufacturing processes or formulation +
- Studying the conformational stability of a protein under stress -- thermal stability, pH stability, and stability to denaturants and how this stability is altered by buffer composition or addition of stabilizers and excipients +
- CD is excellent for finding solvent conditions that increase the melting temperature and/or the reversibility of thermal unfolding, conditions which generally enhance shelf life +
- Determining whether protein-protein or protein-ligand interactions alter the conformation of protein +
- If there are any conformational changes, this will result in a spectrum which will differ from the sum of the individual components. Small conformational changes have been seen, for example, upon formation of several different receptor/ligand complexes +
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diff --git a/src/lab/Pre-Requisite.html b/src/lab/Pre-Requisite.html
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+ Circular Dichroism
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In this lab we will first carry out a few simple experiments to understand the principle working behind CD spectroscopy. We will then proceed with experiments involving the CD spectrophotometer, which is by far the most effective of the three methods.
+-
+
- Experiment 1: To determine the specific rotation of a sugar using a polarimeter. +
- Experiment 2: To understand the effect of chiral substances on plane polarized light as a function of wavelength. +
- Experiment 3:To study the Optical Rotatory Dispersion (ORD) of some chiral substances. +
- Experiment 4: To study the Circular Dichroism of some substances with chiral chromophores. +
- Experiment 5: To Study the operational details of a CD spectrometer. +
- Experiment 6: To Study the effect of Secondary Structure elements on CD spectrum of proteins. +
- Experiment 7: To deconvolute the CD spectrum of a given protein solution and to classify it in terms of its secondary structure elements. +
- Experiment 8: To study the thermal stability of proteins. +
- Experiment 9:To study the effect of chemical denaturants on protein stability. +
- Experiment 10: To study the comparison between protein unfolding and protein aggregation in the presence of chemical denaturing agents. +
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diff --git a/src/lab/TargetAudience.html b/src/lab/TargetAudience.html
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+ Chemical Sciences
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+ Circular Dichroism
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+Install Flash Plugin to run the Simulation + +
+Basic understanding of high-school level science is sufficient to perform the experiments designed in this lab. But it is expected that a student will understand the following topics in details. +
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- Basic principles of Optics: light as an electromagnetic wave. +
- Basic principles of Spectroscopy: a method to study atomic/molecular structure. +
- Basic principles of Physical Chemistry: structure of an atom, inter atomic bonds, inter molecular bonds, ions, pH etc. +
- Basic concepts of Biology: concept of cellular mechanisms, enzymes etc. +
+
+
System Requirements
+-
+
- The labs are best viewed in Google Chrome. +
- Flash plugin must be installed in your browser. +
- Java plugin must be installed in your browser. +
+Install Flash Plugin to run the Simulation + +
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diff --git a/src/lab/exp1/Experiment.html b/src/lab/exp1/Experiment.html
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+ Chemical Sciences
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+ Circular Dichroism
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+ Circular Dichroism Virtual Laboratory is desigen for mainly Under Graduate (3rd/4th year) and Post Graduate students. This lab will serve the purpose of interest of a large and diverse group of students. +
+Following students may be benefited from this lab. +
+
-
+
- Undergraduate (B.Tech, B.Sc.), Postgraduates (M.Sc, M.Pharm) as part of courses Biotechnology, Biochemistry, Chemistry. +
- Undergradute and post graduate level (M.S., diploma, and Ph.D level) as part of courses Biotechnology, Biochemical engineering, Biophysics, Structural Biophysics, Structural Molecular Biology, Molecular Biophysics. +
+
This lab may also be helpful for research scientists interested in photochemistry and photobiology.
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diff --git a/src/lab/exp1/Feedback.html b/src/lab/exp1/Feedback.html
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+ Chemical Sciences →Circular Dichroism Virtual Lab →List Of Experiments
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+ Chemical Sciences →Circular Dichroism Virtual Lab →List Of Experiments
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+ To determine the specific rotation of a sugar using a polarimeter.
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diff --git a/src/lab/exp1/Introduction.html b/src/lab/exp1/Introduction.html
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+ Chemical Sciences →Circular Dichroism Virtual Lab →List Of Experiments
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+ Chemical Sciences →Circular Dichroism Virtual Lab →List Of Experiments
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+ To determine the specific rotation of a sugar using a polarimeter.
Feedback
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diff --git a/src/lab/exp1/Manual.html b/src/lab/exp1/Manual.html
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+ Chemical Sciences →Circular Dichroism Virtual Lab →List Of Experiments
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+ Chemical Sciences →Circular Dichroism Virtual Lab →List Of Experiments
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+ To determine the specific rotation of a sugar using a polarimeter.
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+The study of optical activity of liquids began in the early 19th century with Biot and other scientists. They found that solutions of sugar and certain other naturally occurring chemicals would rotate a beam of polarized light passing through the solution. They called such substances optically active, a term which is still used. The instrument used to demonstrate or to measure this rotation was given the name polarimeter. +
++Clockwise rotation is given a positive (+) sign; counterclockwise rotation is given a negative (-) sign. Certain substances rotate light to a much greater extent than others. Both the direction of rotation and the amount of rotation per gram of solute in a given volume of solution are characteristic properties and can be used to identify an unknown substance. When the identity of the solute is known, the polarimeter can be used to determine the concentration of the solution. +
++It may be noted that approximately 25% of all drugs are marketed as either racemates (mixtures of two enantiomers) or mixtures of diasteromers. The orientation around a chiral center can have a dramatic impact on the pharmacological response of that drug in the human body. Such recent observations brought about severe tightening in the laws surrounding the introduction of new drugs into the market. Thus, chiral synthesis and purification became a crucial aspect of all successful drug manufacturing procedures. This is just one of the several areas highlighting the importance of polarimetric studies. +
+
+Since polarized light is the basis for all studies of optical activity, you are advised to review the "Background principles" described in the home page of this laboratory. In particular, please go through the tutorials in the pages: +
+
+http://www.enzim.hu/~szia/cddemo/edemo1.htm and
+http://www.enzim.hu/~szia/cddemo/edemo9.htm
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diff --git a/src/lab/exp1/Objective.html b/src/lab/exp1/Objective.html
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+ Chemical Sciences →Circular Dichroism Virtual Lab →List Of Experiments
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+ Chemical Sciences →Circular Dichroism Virtual Lab →List Of Experiments
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+ To determine the specific rotation of a sugar using a polarimeter.
+ To know details regarding the instrumentation of the polarimeter.
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+ To directly perform an experiment using the polarimeter.
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+ Estimate the specific rotation [α] using the equation for liquids.
+ Compare the obtained specific rotation [α] value against the table provided below to identify the identity of the sugar used in the experiment.
+ Click on save form and perform calculations and choose the appropriate identity of the unknown sugar you have used in the experiment.
+ Either click on “Click to restart the experiment” and follow the instructions from step 3 or “Click to try with same sample sugar” and follow the instructions from step 3, substep (ii).
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+ In this experiment, the user will prepare a sugar solution of known concentration (c), but unknown identity. The user will obtain the observed rotation (αobs) from the experiment using the polarimeter and use that information to calculate the specific rotation [α] of the given sample using the above formula. The identity of the sample can thus be found out from the given list of specific rotations for different chemicals. +
++ The procedure to perform this experiment is self-explanatory and leads the user in a step-by step manner to accomplish the task. The protocol briefly involves the following steps:
++
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+
- User will click on the virtual lab link (simulator).
+ i.Click on “Know Your Polarimeter” option.
+ ii.Click on the Polarimeter to Zoom-in as indicated by an arrow.
+ iii.Click on the red button provided to switch on the polarimeter and to see the light source in the polarimeter instrument.
+ iv.Click on the panel indicated by arrow to open the sample chamber.
+ v.After you see 100% Intensity of the light coming through the eye piece at α20Dthen click Continue.
+ vi.Click on the button “click to record experiment without login” to perform an experiment and further follow instructions as detailed in step 3.
+
+ i.Click on any of the 6 sugar sample in the bottles on the shelves.
+ ii.Type in the amount of sugar (in gm/100mL) that you want to use for the experiment in the text box provided and then presses enter on the keyboard.
+ iii.Click on one of the sample cells to transfer the prepared sugar solution into a cell of particular path length.
+ iv.After transferring the contents into the sample cell, click on the sample cell to place it into the polarimeter’s sample chamber.
+ v.Click on the power button to switch on the polarimeter to record the optical rotation of the sample.
+ vi.To rotate the dial of the eye piece in clockwise direction click on ‘+’ button and for anticlockwise direction click on ‘-’ button.
+ vii.To change the Increment Factor click on the IF* buttons 1, 5 or 10 accordingly.
+ viii.Rotate the dial both in clockwise and anti-clockwise directions for the whole 360o until you see a maximum light intensity in the right semicircle, matching the left semicircle exactly.
+ ix.Click on open record form and note down all the 4 different αobs angles where you observed maximum Intensity.
+
+
+
| Chemical name | +Specific rotation[α]20D | +
| D-glucose | ++52.7 | +
| Lactose | ++55.4 | +
| D-fructose | +-92.4 | +
| L-arabinose | ++104.5 | +
| D-mannose | ++14.2 | +
| D-arabinose | ++105.0 | +
| D-xylose | ++18.8 | +
| D-galactose | ++80.2 | +Sucrose | ++66.5 | + +Maltose | ++130.5 | + +Dextrin | ++195 | + +
*NOTE: While doing the experiments you will notice that if the maximum intensity of light is observed at x° degree, it is also observed at (180+ x)° degree. From a single experiment, it is not possible to infer which the value for total rotation αtotal actually is. Then again, the observation only tells us about the orientation of the plane of polarization of the emergent light relative to that of the incident light. However, one can not off hand say anything about what is the actual total rotation the plane has undergone. Thus, for example, the observation of maxima at x° and at (180 + x)° may mean that the value of αtotal could also be (360n + x) °; where n = 0, 1, 2, … Similarly, it could also be that αtotal is -(360 - x)° or –(360 - (180 + x))° depending on whether the rotation of the light has taken place in the anticlockwise or clockwise direction. For example if the maximum intensity is observed at α value of 30° and 210° , then could be any one of the values: 30°, 210°, -330°, -150° or even 390° or 570°. To confirm the actual value of αtotal, one needs to repeat the experiment using different concentration and variable path lengths. Though there would be an ambiguity regarding the αtotal for each of these experiments, the correct choices would provide the same specific rotation [α]Tλ or [α]20D +
++
Sample data for the experiment
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diff --git a/src/lab/exp1/Quizzes.html b/src/lab/exp1/Quizzes.html
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+ Chemical Sciences →Circular Dichroism Virtual Lab →List Of Experiments
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+ Chemical Sciences →Circular Dichroism Virtual Lab →List Of Experiments
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+ To determine the specific rotation of a sugar using a polarimeter.
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+The purpose of this lab is to use optical rotation as a method for determining the identity of unknown sugars. By doing this experiment the user will be able to: +
++
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+
- Understand the interaction of plane polarized light with solutions of chiral substances + +
- Understand various processes and techniques involved in measuring the optical activity of sugar solutions using polarimeter. + +
- Understand the step-by-step procedure including most details from preparing the sample to identification of sugar solution used as part of the experiment. + +
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diff --git a/src/lab/exp1/Theory.html b/src/lab/exp1/Theory.html
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+ To determine the specific rotation of a sugar using a polarimeter.
+Click here to have a self evaluation.
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Click here to have a self evaluation.
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diff --git a/src/lab/exp1/scraplab.py b/src/lab/exp1/scraplab.py
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+from bs4 import BeautifulSoup
+heading=""
+breadcrumb=""
+def writefile(fname,s,s1,heading):
+ f=open(fname, 'w+')
+ f.write(template)
+ f.seek(0)
+
+ content = f.read()
+ content=content.replace('Disciplines and Domains',breadcrumb)
+ f.seek(0)
+ f.write(content)
+ print content
+
+ f.seek(0)
+ content=f.read()
+ k=content.index('
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+ Chemical Sciences →Circular Dichroism Virtual Lab →List Of Experiments
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+ Chemical Sciences →Circular Dichroism Virtual Lab →List Of Experiments
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+Chirality and optical activity:
+Chiral molecules have an asymmetrical center which respond to light as a lens and rotate the light. The ability to rotate light is termed optical activity and substances that exhibit this property are called optically active substances. Optically active organic molecules have a spiral structure like a right-handed or left-handed screw. It is this spiral nature of the molecule, which rotates the plane of polarization of light passing through it. Right-handed molecules will rotate the plane of polarization clockwise as viewed in the direction of the beam, while left-handed molecules rotate the plane in a counter clockwise direction. If right-handed and left-handed species of a given molecule occurred with equal abundance, then there would be no net effect on the polarization of light passing through. However, naturally occurring biological molecules of a given species are always either purely right-handed or purely left-handed. However, these enantiomer compounds rotate light by exactly the same amount but in the opposite direction. The degree to which a substance rotates light may be used to determine a) the identity of the substance, b) the enantiomer purity of the substance or c) the concentration of a known substance in a solution. In order to observe rotation, the light which is passed through the solution must be plane polarized. Ordinary light has waves which are oriented in all directions. Plane polarized light is made up of waves which are oriented parallel to a defined plane.
+
+
+ |
+
+When a beam of plane polarized light passes through a solution of optically active material the light will rotate. +
+
+
+ |
+
+This is because, when light interacts with matter, two basic phenomena occur, namely, absorption and the decrease in the velocity of light. Absorption is the decrease in the intensity of light because a part of the incident light is absorbed by the material. The decrease in velocity of light is due to refractive index of the material, because the velocity of light is smaller in the material than in the vacuum. +
+
+If the refraction index of the material differs between left and right circularly polarized light, then such materials are shown to exhibit a phenomenon called circular birefringence. Circular birefringence rotates the plane of polarization of the resultant plane-polarized light. This is termed as optical rotation. The angle by which the polarization plane of the light exiting the medium rotates with respect to the original polarization plane is determined by the difference between the refraction indices for the two circularly polarized components (and on the length of path traversed in the medium). Any molecule which exhibits optical rotation or make plane-polarized light elliptically polar is called an optically active material. +
+How do chiral molecules exhibit Optical Rotation?
++Interestingly, a monochromatic linearly polarized light beam can be considered as a superposition of two circularly polarized electromagnetic waves that are propagating in the same direction with the same frequency but the opposite sense of rotation. The plane of polarization of the resulting linearly polarized wave thus prepared can be changed (rotated) by applying a phase shift between its two circularly polarized components. With the help of this concept we can explain the phenomenon of optical rotation: We have seen that chiral molecules interact slightly differently with the two circularly polarized components of a linearly polarized light beam. This is true both for absorption and refraction. Left- and right hand circularly polarized light beams also have slightly different refractive indices in a chiral medium. This means that even if they are not absorbed they travel at different speeds through the medium. Therefore, this causes a phase shift between the two circularly polarized components which increases proportional to the path length that the light travels through the chiral medium. This phase shift manifests itself as a rotation of the plane of polarization of the resultant linearly polarized light beam - optical rotation. +
+
+The degree of rotation of the plane polarized light depends on the wavelength of the light (usually, the yellow sodium D line near 589 nm wavelength is used), the optical path length, the concentration of the solution, and the chemistry of the molecule. Under identical conditions, some molecules rotate polarized light more than the others do. In order measure how good chiral molecules rotate plane- polarized light, a term called as the "specific rotation" was coined. The specific rotation of a substance is an intrinsic characteristic similar to other properties such melting point, or solubility. By convention, the specific rotation of a chemical is defined as the observed rotation when light of a specified wavelength passes through sample path length of one decimeter (1 dm = 10 cm) and a sample concentration of 1 g/mL. +
+Circular birefringence and optical rotation
++Chiral molecules exhibit circular birefringence, which means that a solution of a chiral substance presents an anisotropic medium through which left circularly polarised (L-CPL) and right circularly polarised (R-CPL) propagate at different speeds. A linearly polarised wave can be thought of as the resultant of the superposition of two circularly polarised waves, one left-circularly polarised, the other right-circularly polarised. On traversing the circularly birefringent medium, the phase relationship between the circularly polarised waves changes and the resultant linearly polarised wave rotates. This is the origin of the phenomenon known as optical rotation, which is measured using a polarimeter. Measuring optical rotation as a function of wavelength is termed optical rotatory dispersion (ORD) spectroscopy. +
+
+
+ |
+
Circular birefringence:the orange cuboid represents the sample
+
+For pure liquids:
+
[α]Tλ = α/(l * d)
+ +
+In this equation, +
++l is the path length in decimeters +
++d is the density of the liquid in g/mL for a sample at a +
++temperature T (given in degrees Celsius) and wavelength λ (in nanometers). +
+
+If the wavelength of the light used is 589 nanometer (the sodium D line), the symbol “D” is used. The sign of the rotation (+ or -) is always given. +
+
+For solutions, a different equation is used: +
+[α]Tλ = 100 * α/(l * c)
+
+In this equation, +
++l is the path length in decimeters +
++c is the concentration in g/100mL for a sample at a +
++temperature T (given in degrees Celsius) and wavelength λ (in nanometers). +
++When using this equation, the concentration and the solvent are always provided in parentheses after the rotation. The rotation is reported using degrees, and no units of concentration are given (it is assumed to be g/100mL). +
+
Functioning of the polarimeter
+a
+
+
+ |
+
b
+
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+ |
+
+Fig. 1: Schematic representation of the functioning of the polarimeter. +
++a. When the sample tube is empty, the planes of polarization of the polarizing and the analyzing prisms are same and αobs is 0° +
++b. When the sample tube has a solution of a chiral (optically active) substance, the plane of polarization of the emergent polarized light changes. One now needs to rotate the analyzer prism for its plane of polarization to coincide with the plane of the emergent light. This corresponds to the maximum intensity of the transmitted light. The αobs is shown with a green arrow. +
+ +')
+
+ t1=content.index('')
+ print t1
+ f.seek(t1+13)
+ s1=s1+f.read()
+ #print s1
+ f.seek(t1+13)
+ f.write(s1)
+ f.seek(0)
+ content=f.read()
+ t= content.index('')
+ f.seek(t+13)
+ s='
' +while sectionNumber<=len(sectionno): + tag="" + att = ''+'experiment-article-section-'+str(sectionNumber)+'-heading' + tagger = soup.findAll('div', attrs={'id':att,'class':'heading'}) + if not tagger: + sectionNumber=sectionNumber+1 + continue + tag1=str(tagger[0].text) + tag1=tag1.strip() + print tag1 + if tag1=='Prerequisite S/W': + tag1='Prerequisites' + st+='
'+heading+'
'+s + s=s+f.read() + f.seek(t+13) + f.write(s) +f=open("template1.html",'r') +template=f.read() +f=open("content.html",'r') +srchtml=f.read() +f.close() +soup = BeautifulSoup(srchtml, 'html.parser') +sectionno=soup.find_all('section') +d=['introduction','theory','objective','experiment','manual','quizzes','further_readings','procedure','hh','ll'] +print len(sectionno) +sectionNumber=1 +st="" + +att = ''+'experiment-article-heading' +tagger = soup.findAll('header', attrs={'id':att,'class':'heading'}) +heading=str(tagger[0].text) +heading=heading.strip() +print heading +att = ''+'experiment-header-heading' +tagger = soup.findAll('div', attrs={'id':att,'class':'heading'}) +heading1=str(tagger[0].text) +print heading +heading1=heading1.strip() +breadcrumb='Chemical Sciences →'+heading1+' →List Of Experiments' +while sectionNumber<=len(sectionno): + tag="" + att = ''+'experiment-article-section-'+str(sectionNumber)+'-heading' + tagger = soup.findAll('div', attrs={'id':att,'class':'heading'}) + if not tagger: + sectionNumber=sectionNumber+1 + continue + tag1=str(tagger[0].text) + tag1=tag1.strip() + print tag1 + if tag1=='Prerequisite S/W': + tag1='Prerequisites' + st+='
'+tag1+'
' + sectionNumber=sectionNumber+1 +st+='Feedback
' +sectionNumber=1 +while sectionNumber<=len(sectionno): + tag="" + tag="" + att = ''+'experiment-article-section-'+str(sectionNumber)+'-heading' + tagger = soup.findAll('div', attrs={'id':att,'class':'heading'}) + if not tagger: + sectionNumber=sectionNumber+1 + continue + tag1=str(tagger[0].text) + tag1=tag1.strip() + if tag1=='Prerequisite S/W': + tag1='Prerequisites' + att = ''+'experiment-article-section-'+str(sectionNumber)+'-content' + tagger = soup.findAll('div', attrs={'id':att,'class':'content'}) + + tag+=str(tagger[0]) + #print tag + #print st + + + writefile(tag1+'.html',tag,st,heading) + #print sectionNumber + sectionNumber=sectionNumber+1 + +f=open("Feedback.html",'w+') +f.write(template) +f.seek(0) + +content = f.read() +content=content.replace('Disciplines and Domains',breadcrumb) +f.seek(0) +f.write(content) +#print content +f.seek(0) +content=f.read() +k=content.index('')
+
+t1=content.index('')
+print t1
+f.seek(t1+13)
+st=st+f.read()
+#print s1
+f.seek(t1+13)
+f.write(st)
+f.seek(0)
+content=f.read()
+t= content.index('')
+f.seek(t+13)
+s='Welcome to Virtual Labs - A MHRD Govt of india Initiative
+
+
+
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+Welcome to Virtual Labs - A MHRD Govt of india Initiative
+
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+Welcome to Virtual Labs - A MHRD Govt of india Initiative
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+Welcome to Virtual Labs - A MHRD Govt of india Initiative
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+Welcome to Virtual Labs - A MHRD Govt of india Initiative
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+Welcome to Virtual Labs - A MHRD Govt of india Initiative
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+Welcome to Virtual Labs - A MHRD Govt of india Initiative
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+Welcome to Virtual Labs - A MHRD Govt of india Initiative
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'+heading+'
'+'Feedback' +s=s+f.read() +f.seek(t+13) +f.write(s) diff --git a/src/lab/exp1/template1.html b/src/lab/exp1/template1.html new file mode 100644 index 0000000..382fb88 --- /dev/null +++ b/src/lab/exp1/template1.html @@ -0,0 +1,205 @@ + + + + + + + + + + +
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diff --git a/src/lab/exp10/Experiment.html b/src/lab/exp10/Experiment.html
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+ Disciplines and Domains
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diff --git a/src/lab/exp10/Feedback.html b/src/lab/exp10/Feedback.html
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+ Chemical Sciences →Circular Dichroism Virtual Lab →List Of Experiments
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+ Chemical Sciences →Circular Dichroism Virtual Lab →List Of Experiments
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+ To study the comparison between protein unfolding and protein aggregation in the presence of chemical denaturing agents.
+
+Click here to start the experiment. + +
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diff --git a/src/lab/exp10/Introduction.html b/src/lab/exp10/Introduction.html
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+ Chemical Sciences →Circular Dichroism Virtual Lab →List Of Experiments
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+ Chemical Sciences →Circular Dichroism Virtual Lab →List Of Experiments
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+ To study the comparison between protein unfolding and protein aggregation in the presence of chemical denaturing agents.
Feedback
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diff --git a/src/lab/exp10/Manual.html b/src/lab/exp10/Manual.html
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+ +
+ Chemical Sciences →Circular Dichroism Virtual Lab →List Of Experiments
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+
+ Chemical Sciences →Circular Dichroism Virtual Lab →List Of Experiments
+
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+ To study the comparison between protein unfolding and protein aggregation in the presence of chemical denaturing agents.
+
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+
+In experiment 8 and 9 we have seen that how a protein can go from a folded to an unfolded state at extreme environmental conditions (high temperature or presence of other chemical ). At extreme conditions intramolecular bonds are broken and the protein molecule gains a deformed shape. But in a protein solution, in the unfolded state, proteins may form intermolecular bonds and can aggregate among themselves. Whether a protein will aggregate or not that depends on the environment.
+
+So to study the characteristics of the protein in the unfolded state it is important to know whether the protein has undergone aggregation or not. CD spectroscopy provides a nice way to find out whether aggregation is taking place at the unfolded state. +
+
+In this experiment we will study the method to determine aggregation from CD spectroscopy. And will try to determine whether aggregation is happening in two different samples of Rhodopsin.
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diff --git a/src/lab/exp10/Objective.html b/src/lab/exp10/Objective.html
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+ Chemical Sciences →Circular Dichroism Virtual Lab →List Of Experiments
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+ Chemical Sciences →Circular Dichroism Virtual Lab →List Of Experiments
+
+
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+ To study the comparison between protein unfolding and protein aggregation in the presence of chemical denaturing agents.
+
+
+
1. Video Tutorial
++ +
++
click here to ENLARGE.
++
2. Step-by-step Procedure
+
+
Step1: Select any one of the two samples.
+
Step2: Follow the instructions given in the Experiemntal-setup page.
+
Step3: Select the temperature range and step size of your interest.
+
Step4: Go through the instructions and fill up the table by clicking at the 222nm.
+
Step5: Observe the plot and find out the transition temperature.
+
Step6: Click on the “Measure Dynode Voltage”.
+
Step7: Follow the instruction and fill up the table by clicking at the 222nm.
+
Step8: Observe the change in Dynode voltage.
+
Step9: Click on “Go to Analysis”
+
Step 10: Submit your answers. You can verify them from redoing the experiment or going through the theory.
+
Step 11: Once you have studied both the samples you may compare them by clicking “COMPARE”.
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diff --git a/src/lab/exp10/Quizzes.html b/src/lab/exp10/Quizzes.html
new file mode 100644
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+ Chemical Sciences →Circular Dichroism Virtual Lab →List Of Experiments
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+ Chemical Sciences →Circular Dichroism Virtual Lab →List Of Experiments
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+ To study the comparison between protein unfolding and protein aggregation in the presence of chemical denaturing agents.
+
+
The purpose of this lab is to make students familiar with the concept of protein aggregation. In the unfolded state a protein undergo aggregation. And CD spectroscopic methods provides us a nice way to determine that. By doing this experiment user will be able to,
++
-
+
- Understand how at unfolded state a protein undergoes aggregation. +
- Understand how CD spectroscopic methods are useful to determine aggregation effect. +
- Experience how environmental conditions control the aggregation effect. +
- Understand how comparative study of Dynode voltage helps to determine whether aggregation has taken place or not. +
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diff --git a/src/lab/exp10/Theory.html b/src/lab/exp10/Theory.html
new file mode 100644
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+ +
+ Chemical Sciences →Circular Dichroism Virtual Lab →List Of Experiments
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+
+ Chemical Sciences →Circular Dichroism Virtual Lab →List Of Experiments
+
+
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+ To study the comparison between protein unfolding and protein aggregation in the presence of chemical denaturing agents.
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diff --git a/src/lab/exp10/scraplab.py b/src/lab/exp10/scraplab.py
new file mode 100644
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--- /dev/null
+++ b/src/lab/exp10/scraplab.py
@@ -0,0 +1,124 @@
+from bs4 import BeautifulSoup
+heading=""
+breadcrumb=""
+def writefile(fname,s,s1,heading):
+ f=open(fname, 'w+')
+ f.write(template)
+ f.seek(0)
+
+ content = f.read()
+ content=content.replace('Disciplines and Domains',breadcrumb)
+ f.seek(0)
+ f.write(content)
+ print content
+
+ f.seek(0)
+ content=f.read()
+ k=content.index('
+
+
+
+
+
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+
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+
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+ +
+ Chemical Sciences →Circular Dichroism Virtual Lab →List Of Experiments
+
+
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+
+ Chemical Sciences →Circular Dichroism Virtual Lab →List Of Experiments
+
+
+
+
+
+
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+
+
+ To study the comparison between protein unfolding and protein aggregation in the presence of chemical denaturing agents.
+
+
+
+
+What is Protein Aggregation?
++
Protein aggregation is a common issue encountered during manufacture of biotherapeutics. Aggregation is a general term that encompasses several types of interactions or characteristics. Aggregates of proteins may arise from several mechanisms and may be classified in numerous ways, including soluble/insoluble, covalent/noncovalent, reversible/irreversible, and native/denatured. There is no consistent definition of what is meant by a “soluble” aggregate, so working definitions are often employed. Soluble aggregates refer to those that are not visible as discrete particles and that may not be removed by a filter with a pore size of 0.22 μm. Conversely, insoluble aggregates may be removed by filtration and are often visible to the unaided eye. However, the levels of soluble aggregates such as dimers and trimers that are acceptable are not well defined. Covalent aggregates arise from the formation of a chemical bond between 2 or more monomers. Disulfide bond formation resulting from previously unpaired free thiols is a common mechanism for covalent aggregation. Oxidation of tyrosines may also result in covalent aggregation through the formation of bi-tyrosine. For some proteins, a covalent interaction between monomers is required to form a stable protein structure. Reversible protein aggregation typically results from relatively weak noncovalent protein interactions. The reversibility is sometimes indicative of the presence of equilibrium between the monomer and higher order forms. This equilibrium may shift as a result of a change in solution conditions such as a decrease in protein concentration or a change in pH. A weak, reversible self-association of this type has been observed in a monoclonal antibody to VEGF. On occasion, reversible protein self-association manifests itself as an increase in viscosity.
+The effect of the presence of self-associated species is not always known. Both the thermodynamics and the kinetics of the system may assist in understanding how to control the association of the protein. In addition, this knowledge aids in determining how serious the presence of associated species may be during the development of a protein therapeutic. Both the potential for increased exposure to the associated species and the route of administration present potential safety concerns.
+Historically, investigators believed that denaturation was a prerequisite for protein aggregation. Exposure of hydrophobic surfaces upon denaturation results in favorable protein: protein interactions in aqueous solutions. It is true that this type of interaction leads to the formation of aggregates in many proteins and may cause extreme precipitation. However, the role of native protein interactions in the formation of self-associated species has recently become more appreciated. Small perturbations in protein structure may expose hydrophobic surfaces that lead to aggregation. Electrostatic interactions have been implicated in the formation of self-associated species of a monoclonal antibody, while dipole-dipole interactions are believed to be the cause of fibrillogenic association of β-sheets.
+Just as there are many types of interactions that can lead to protein aggregation, there are many environmental factors that can lead to aggregation. Solution conditions such as temperature, protein concentration, pH, and the ionic strength may affect the amount of aggregate observed. The presence of certain ligands, including specific ions, may enhance aggregation. Stresses to the protein such as freezing, exposure to air, or interactions with metal surfaces may result in surface denaturation, which then leads to the formation of aggregates. Finally, mechanical stresses may cause protein aggregation. Each of these environmental factors is typically encountered during bioprocessing.
++
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++
How CD technique can help in measuring Aggregation:
+
Circular dichroism spectroscopy is widely used to follow the stability of a protein by increasing denaturant conditions. The most common denaturants are temperature, chemicals (usually urea and guanidinium chloride) and extremes of pH (usually acidic). As the denaturant condition increases the stability of the protein decreases and then unfolds (Ramos and Ferreira, 2005). CD is a convenient technique to follow protein unfolding because the spectra of folded and random coil are quite different.
+
+ + |
+
Characteristic far-UV CD spectra for an all-α-helix, an all-β-sheet and a random coil +protein. The spectrum for an all-α-helix protein (Alpha) has two negative bands of similar +magnitude at 222 and 208 nm, and a positive band at ~ 190 nm. The spectrum for an all β- +sheet protein (Beta) has in general a negative band between 210-220 nm and a positive band +between 195 - 200 nm. The spectrum for a disorderly (random) protein has a negative +band of great magnitude at around 200 nm. |
+
The indication of absorption of light is given by the dynode voltage module (in volts, V), which measures the response of the photomultiplier during the measurement and is an indicator of the quantity of photons that are not absorbed or are scattered by the sample. The higher the amount recorded, the greater the absorption or the scattering. A high absorption causes an increase in noise and consequently increases the errors of elipticity measurement. In general, the acceptable limit for reliable reading is 600 V, but acquisitions of up to 700 V are acceptable provided that the number of spectra collection and accumulation is highly enough to reduce the noise generated by absorption. It is advisable not to perform readings with voltage above 700 V as this may cause damage to the unit. The use of cells with thinner pathlengths decreases the number of photons that the sample is able to absorb. By using the Beer-Lambert law one can vary the concentration of sample and the length of the optical path in order to optimize the measurement.
++
+
+
+
+
+From the data of Dynode Voltage how we can infer whether Aggregation has take place or not:-
+
The unfolding transition can be easily determined by choosing a wavelength where the difference in signal for folded and unfolded protein is large. For instance, α-helical proteins have a large CD signal at 222 nm in which unfolded proteins has none or little signal. As discussed above the dynode voltage V (high-voltage applied to the photomultiplier of the UV detector to compensate for the reduction in the intensity light) is a result from light absorption or scattering and should be recorded for all CD measurements. For being related to scattering the recorded dynode voltage V can be used to probe the turbidity of the solution and hence aggregation (Benjwal et al., 2006). Accordingly to the Beer-Lambert law a heat induced change in the dynode voltage can only be originated from variations in the light scattering which depends on the particle size. Thus the signal of CD at 222 nm and the dynode voltage V as a function of temperature can be used to determine the amount of aggregation from denaturation. The curve profile shows that the CD signal starts to decrease at about 85ºC followed by a two-state like transition. This transition indicates that the protein unfolds as verified by the decrease in CD signal and aggregates as verified by the increase in the voltage which is related to light scattering. To sum up, as the protein aggregated its size increased and more light was scattered.
+
+ + |
+
Circular dichroism spectra of HoloMyoglobin (Mb) as a function of temperature. Circular dichroism measurements were recorded using a Jasco J-810 spectropolarimeter with 10 mm pathlength cell following the signal at 222 nm and the dynode voltage V as a function of temperature. A heat-induced change in the dynode voltage is originated from variations in the light scattering which depends on the particle size and thus from aggregation. The transitions indicate that the protein unfolds as verified by the decrease in CD signal and aggregates as verified by the increase in the voltage which is related to light scattering. A non-aggregated protein would have shown a constant dynode voltage throughout the whole scan. |
+
+
')
+
+ t1=content.index('')
+ print t1
+ f.seek(t1+13)
+ s1=s1+f.read()
+ #print s1
+ f.seek(t1+13)
+ f.write(s1)
+ f.seek(0)
+ content=f.read()
+ t= content.index('')
+ f.seek(t+13)
+ s='
' +while sectionNumber<=len(sectionno): + tag="" + att = ''+'experiment-article-section-'+str(sectionNumber)+'-heading' + tagger = soup.findAll('div', attrs={'id':att,'class':'heading'}) + if not tagger: + sectionNumber=sectionNumber+1 + continue + tag1=str(tagger[0].text) + tag1=tag1.strip() + print tag1 + if tag1=='Prerequisite S/W': + tag1='Prerequisites' + st+='
'+heading+'
'+s + s=s+f.read() + f.seek(t+13) + f.write(s) +f=open("template1.html",'r') +template=f.read() +f=open("content.html",'r') +srchtml=f.read() +f.close() +soup = BeautifulSoup(srchtml, 'html.parser') +sectionno=soup.find_all('section') +d=['introduction','theory','objective','experiment','manual','quizzes','further_readings','procedure','hh','ll'] +print len(sectionno) +sectionNumber=1 +st="" + +att = ''+'experiment-article-heading' +tagger = soup.findAll('header', attrs={'id':att,'class':'heading'}) +heading=str(tagger[0].text) +heading=heading.strip() +print heading +att = ''+'experiment-header-heading' +tagger = soup.findAll('div', attrs={'id':att,'class':'heading'}) +heading1=str(tagger[0].text) +print heading +heading1=heading1.strip() +breadcrumb='Chemical Sciences →'+heading1+' →List Of Experiments' +while sectionNumber<=len(sectionno): + tag="" + att = ''+'experiment-article-section-'+str(sectionNumber)+'-heading' + tagger = soup.findAll('div', attrs={'id':att,'class':'heading'}) + if not tagger: + sectionNumber=sectionNumber+1 + continue + tag1=str(tagger[0].text) + tag1=tag1.strip() + print tag1 + if tag1=='Prerequisite S/W': + tag1='Prerequisites' + st+='
'+tag1+'
' + sectionNumber=sectionNumber+1 +st+='Feedback
' +sectionNumber=1 +while sectionNumber<=len(sectionno): + tag="" + tag="" + att = ''+'experiment-article-section-'+str(sectionNumber)+'-heading' + tagger = soup.findAll('div', attrs={'id':att,'class':'heading'}) + if not tagger: + sectionNumber=sectionNumber+1 + continue + tag1=str(tagger[0].text) + tag1=tag1.strip() + if tag1=='Prerequisite S/W': + tag1='Prerequisites' + att = ''+'experiment-article-section-'+str(sectionNumber)+'-content' + tagger = soup.findAll('div', attrs={'id':att,'class':'content'}) + + tag+=str(tagger[0]) + #print tag + #print st + + + writefile(tag1+'.html',tag,st,heading) + #print sectionNumber + sectionNumber=sectionNumber+1 + +f=open("Feedback.html",'w+') +f.write(template) +f.seek(0) + +content = f.read() +content=content.replace('Disciplines and Domains',breadcrumb) +f.seek(0) +f.write(content) +#print content +f.seek(0) +content=f.read() +k=content.index('')
+
+t1=content.index('')
+print t1
+f.seek(t1+13)
+st=st+f.read()
+#print s1
+f.seek(t1+13)
+f.write(st)
+f.seek(0)
+content=f.read()
+t= content.index('')
+f.seek(t+13)
+s='Welcome to Virtual Labs - A MHRD Govt of india Initiative
+
+
+
+
+
+
+
+
+
+
+
+
+
+
+
+
+
+
+Welcome to Virtual Labs - A MHRD Govt of india Initiative
+
+
+
+
+
+
+
+
+
+
+
+
+
+
+
+
+
+
+Welcome to Virtual Labs - A MHRD Govt of india Initiative
+
+
+
+
+
+
+
+
+
+
+
+
+
+
+
+
+
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+Welcome to Virtual Labs - A MHRD Govt of india Initiative
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+Welcome to Virtual Labs - A MHRD Govt of india Initiative
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+Welcome to Virtual Labs - A MHRD Govt of india Initiative
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+Welcome to Virtual Labs - A MHRD Govt of india Initiative
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+Welcome to Virtual Labs - A MHRD Govt of india Initiative
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'+heading+'
'+'Feedback' +s=s+f.read() +f.seek(t+13) +f.write(s) diff --git a/src/lab/exp10/template1.html b/src/lab/exp10/template1.html new file mode 100644 index 0000000..382fb88 --- /dev/null +++ b/src/lab/exp10/template1.html @@ -0,0 +1,205 @@ + + + + + + + + + + +
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diff --git a/src/lab/exp2/Experiment.html b/src/lab/exp2/Experiment.html
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+ Disciplines and Domains
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diff --git a/src/lab/exp2/Feedback.html b/src/lab/exp2/Feedback.html
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+ Chemical Sciences →Circular Dichroism Virtual Lab →List Of Experiments
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+ Chemical Sciences →Circular Dichroism Virtual Lab →List Of Experiments
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+ To understand the effect of chiral substances on plane polarized light as a function of wavelength.
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+
Click here to observe dispersive effect.
+
+Click here to observe absorptive effect.
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diff --git a/src/lab/exp2/Introduction.html b/src/lab/exp2/Introduction.html
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+ Chemical Sciences →Circular Dichroism Virtual Lab →List Of Experiments
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+ Chemical Sciences →Circular Dichroism Virtual Lab →List Of Experiments
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+ To understand the effect of chiral substances on plane polarized light as a function of wavelength.
Feedback
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diff --git a/src/lab/exp2/Manual.html b/src/lab/exp2/Manual.html
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+ Chemical Sciences →Circular Dichroism Virtual Lab →List Of Experiments
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+ Chemical Sciences →Circular Dichroism Virtual Lab →List Of Experiments
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+ To understand the effect of chiral substances on plane polarized light as a function of wavelength.
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+We have already learned from Experiment 1, how chiral molecules are optically active and how chiral substances +can be characterized on the basis of their specific rotation [α]. You will have noticed that in that experiment +we have used a sodium lamp with a wavelength of 589 nm as the light source, and may be wondering why the wave length +of light source needed to be specified. +
++Well, as has been explained in the following video using sucrose solution as an example, the specific rotation +[α] for a given substance, and hence the molar rotation [φ], varies as a function of the wavelength (λ) + of the light source.This variation in the optical rotation ([α] or [φ] ) of a substance, with the variation in + the wavelength (λ) of light, is known as optical +rotatory dispersion (ORD). +
+
+
+
+When a plane polarised wave passes through an optically active medium some times its state of polarisation +also gets affected along with its plane of polarisation and it comes out from the medium as an elliptically +polarised light. This phenomena is known as Circular Dichroism. +
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diff --git a/src/lab/exp2/Objective.html b/src/lab/exp2/Objective.html
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+ Chemical Sciences →Circular Dichroism Virtual Lab →List Of Experiments
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+ Chemical Sciences →Circular Dichroism Virtual Lab →List Of Experiments
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+ To understand the effect of chiral substances on plane polarized light as a function of wavelength.
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+
The experiment is devided inte two parts:
+
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+
- Study of dispersive effect +
- Study of absorptive effect +
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+Study of dispersive effect:
+
- + In the first step you can observe "Change in Optical Rotation Due to Change in Difference of RI(Refractive Index)". Here n1 means RI of LCPL and n2 means RI of RCPL. + +
- + 5 sets of n1 and n2 are given. You can choose any one and see its effect on a plane polarised light. + +
- + The value of specific rotation in radian/ cm is also provided. One can study the changes in that with different choices of n1 and n2. + +
- + On clicking “Next” you will be lead to step 2. + +
- + In Step2 one can study "Change in Optical Rotation Due to Change in Wavelength". + +
- +User can select any wave length from the choices given and see how the wave is bevaving in the anisotropic medium. + +
- +The value of Specific rotation in rad/cm is also given. + +
+
+
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+Study of absorptive effect:
+
- +In the first step you can observe "Change in Ellipticity Due to Change in Difference of Extinction Coefficients". Here e1 means extinction coefficients of LCPL and e2 means extinction coefficients of RCPL. + +
- +5 sets of e1 and e2 are given. You can choose any one and see its effect on a plane polarised light. + +
- +The value of ellipticity is also provided. One can study the changes in that with different choices of e1 and e2. + +
- +On clicking “Next” you will be lead to step 2. + +
- +In Step2 one can study “Change in Optical Rotation and Ellipticity Due to Change in Wavelength”. + +
- +User can select any wave length from the choices given and see how the wave is behaving in the anisotropic medium. + +
- +The value of Specific rotation and ellipticity is also given. + +
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diff --git a/src/lab/exp2/Quizzes.html b/src/lab/exp2/Quizzes.html
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+ Chemical Sciences →Circular Dichroism Virtual Lab →List Of Experiments
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+ Chemical Sciences →Circular Dichroism Virtual Lab →List Of Experiments
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+ To understand the effect of chiral substances on plane polarized light as a function of wavelength.
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The purpose of the lab is to understand the effect of chiral substance on a plane polarised + light as a function of wave length. +
++By doing this experiment the user will be able to understand that:
+(1) A plane polarised light is made up of one Right Circularly and one Left Circularly polarised light. +
++(2) Different Refractive Indexes for different circularly polarised components of an anisotropic medium give +rise to Optical Rotation. +
+(3) Different rate of absorption for different circularly polarised components of an anisotropic medium + gives rise to Circular Dichroism. +
++(4) Optical Rotation depends on the wave length of incident light and results Optical Rotatory Dispersion. +
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diff --git a/src/lab/exp2/Theory.html b/src/lab/exp2/Theory.html
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+ Chemical Sciences →Circular Dichroism Virtual Lab →List Of Experiments
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+ Chemical Sciences →Circular Dichroism Virtual Lab →List Of Experiments
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+ To understand the effect of chiral substances on plane polarized light as a function of wavelength.
+Click here to have a self evaluation.
Click here to have a self evaluation.
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diff --git a/src/lab/exp2/scraplab.py b/src/lab/exp2/scraplab.py
new file mode 100644
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+from bs4 import BeautifulSoup
+heading=""
+breadcrumb=""
+def writefile(fname,s,s1,heading):
+ f=open(fname, 'w+')
+ f.write(template)
+ f.seek(0)
+
+ content = f.read()
+ content=content.replace('Disciplines and Domains',breadcrumb)
+ f.seek(0)
+ f.write(content)
+ print content
+
+ f.seek(0)
+ content=f.read()
+ k=content.index('
+
+
+
+
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+ +
+ Chemical Sciences →Circular Dichroism Virtual Lab →List Of Experiments
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+
+
+ Chemical Sciences →Circular Dichroism Virtual Lab →List Of Experiments
+
+
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+
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+ To understand the effect of chiral substances on plane polarized light as a function of wavelength.
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+Figure demonstrating the concept of elliptical polarization of light and CD
+
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+
Recap of Experiment 1
+In experiment:1 we have talked + about the phenomenon of polarisation of electromagnetic waves. We have tried to emphasize on two aspects of a plane polarised + or linearly polarised wave.
+-
+
+A plane polarised wave is a composition of a left and a right +circularly polarised waves propagating in phase. +It is visually described by the following animation. Here the green and the red curve shows +propagation of right and left circularly polarised wave respectively. And superposition of them +is the blue curve which is a plane polarised wave.
+Whenever a electromagnetic wave enters a medium its velocity gets reduced. The ratio of its velocity in vacuum to its velocity in the medium is known as + refractive index and is represented by, n = c0 / v where c0 is the velocity of the wave in vacuum + and v is the velocity of the wave in the medium
+
+
+
+source:http://www.enzim.hu/~szia/cddemo/edemo8.htm+ |
+
+
Anisotropic Medium And Speed of Light
+
+In an isotropic medium the properties of the medium are the same in all directions. + But in an anisotropic medium the properties become directionally dependent. + Anisotropic materials may have different indices of refraction associated with + different crystallographic directions. A common situation is that, there are + two distinct indices of refraction, and they are called birefringent + materials. In case of optically active chilral molecules, + they have different refractive indices for different circularly polarised components of a plane polarised wave(nL ≠ nR). + That means, once the plane polarised wave enters the medium speed of its right circular part and left circular part will + be different(cL ≠ cR). This property is known as circular birefringence. +
++These two aspects together give rise to the phenomena called Optical + Rotation, rotation of plane of polarisation +of a plane polarised wave after it passes through an optically active medium. +
++In experiment 1 we have seen and the Optical +Rotation of a light wave (which is an example of EM wave) when it passes through an optically +active medium like sugar solution. There we have measured the specific rotation [α]Tλ of the +light wave for different concentrations of medium at a fixed wave length of incident light λ~589nm(Na D-line)and fixed temperature T. In the this experiment we are going to observe how the specific rotation changes with wave length of incident light. +
++
Dependence of specific rotation on the wavelength: Optical Rotatory Dispersion
+
+A wave can be characterized by its amplitude and phase. Amplitude of a wave is the maximum +displacement of the particles of the medium from its equilibrium position. Whereas phase +of a wave is the definition of a point in a waveform. Interaction of the wave with a material +effects its velocity and magnitude and this may in turn cause a change in these two characters. +If the material is transparent then no change takes place in phase. But in the case of an +anisotropic medium there may be difference in velocity of different components of the wave +(which causes change in phase). If the medium has absorptive property, then change in amplitude may take place. +
+
+The electric vector of a EM wave
+is always perpendicular to
+the direction of its propagation. For a plane polarised wave the electric vector of its Left circular
+part (EL) and the electric vector of its Right circular part (ER) make equal angles with the plane of
+polarization (αL and αR) ie they are in phase. As we have discussed, after entering an optically active
+medium velocities of the two components of a plane polarised wave (vL and vR) differs from each other.
+As a result the angles that EL and ER make with the initial plane of polarisation, no longer remains the
+same i.e. αL ≠ αR.
+This will give rise to a phase shift between the two components of a plane polarised wave
+,δ = αL – αR .
+
+
+The following figure describes this. +
+
+ |
+
+But the superposition of these two oppositely circularly polarised wave will still +generate a plane polarised wave with its plane of polarisation rotated to an angle +α',as described by the following figure.From the concepts of geometry it may be shown that the angle of rotation α' is half of the phase shift δ.[*] +
+
+ |
+
+Now we will see how this angle of rotation depends on the wave length λ of the incident light. +To understand the dependence let us first write down the electric vectors of a right circularly +polarised and a left circularly polarised wave propagating in the same direction (z) as follows +
++
+ER(z)= Eo/2 cosω(t - z/VR)
+EL(z)= Eo/2 cosω(t - z/VL)
+
+
Here, Eo = the magnitude
+ ω = angular frequency
+ z = distance traveled
+ v = velocity in the medium of the wave
+
+ +
+At any instant, the resultant of ER and EL (i.e. ER + EL) will produce a + plane polarised wave. We will see how much phase shift is generated when +a wave travels a distance, say d, through an optically active medium. + For that we can look at the equations of waves at z = 0 and at z = d. +
++
+
+ER(o)= Eo/2 cosω t
+EL(o)= Eo/2 cosω t
+
+
+
+
+ER(d)= Eo/2 cosω(t - d/VR)
+EL(d)= Eo/2 cosω(t - d/VL)
+
+
+
+Applying trigonometric laws we +can show that the phase difference +between ER(0)+EL(0) and ER(d)+EL(d) is given by +
++
+ +δ=ω .d (1/VR - 1/VL)=ω .d/c (nL - nR) + +
++
+Now as we know, velocity of any EM wave in any medium = (wave length of wave in the mediun) +X (frequency in that medium) and angular frequency = 2πX frequency of wave. So, +
++
+ +δ=2π/λ(nL - nR)d + +
++
+But α'= δ /2. So we can have angle of rotation directly as a function of wave length as, +
++
+ +α '= δ/2= π/λ(nL - nR)d + +
++
+So with change in the wavelength of a light, its optical rotation will also change for the same medium. This phenomena + is known as Optical Rotatory Dispersion. +
++In practical purpose, to study ORD one usually looks at two +parameters – specific rotation [α]Tλ and molecular rotation [M]Tλ. They are defined as follows. +
+
+ |
+
+
+A plot of [α]Tλ or [M]Tλwith wavelength λ is known as ORD spectra. From an ORD +spectra of an unknown material we can have an understanding of the optical properties of the material.
++
+
Absorption and Extinction Coefficient- A Recap
+
When an EM wave passes through a medium, the energy of its photon may be taken up by the electrons of the atoms +of the medium. This is called absorption. +As a result the wave loses its energy and its amplitude gets reduced. This is called attenuation of a wave. +Absorption may occur for any type of polarised wave. The following figures exhibit how a linearly polarised +wave and a circularly polarised wave are attenuated by an absorptive medium. +
++
 |
+ |
+
| Absorption of plane polarised light source:http://www.enzim.hu/~szia/cddemo/edemo10.htm |
+Absorption of circularly polarised light source:http://www.enzim.hu/~szia/cddemo/edemo11.htm |
+
+
+The ratio of radiant power transmitted (P) by a sample to the radiant power incident (P0) on the sample is called the transmittance, T: +T = P/P0 +
+
+Absorbance (A), then, is defined as the logarithm (base 10) of the reciprocal of the transmittance:
+A = -log T = log (1/T)
+
+
+In a spectrophotometer, monochromatic plane-parallel light enters a sample at right angles to the plane-surface
+of the sample. In these conditions, the transmittance and absorbance of a sample depends on the molar concentration
+(c), light path length in centimetres (L), and molar absorptivity (ε) for the dissolved substance at the specified
+wavelength (λ).
+
+Tλ = 10εcL or Aλ = εcL
+
+
+Beer’s Law states that molar absorptivity is constant (and the absorbance is proportional to concentration) +for a given substance dissolved in a given solute and measured at a given wavelength. +
++
+For this reason, molar absorptivities are called molar absorption coefficients or molar extinction coefficients. +Because transmittance and absorbance are unitless, the units for molar absorptivity must cancel with units of measure +in concentration and light path. Therefore, molar absorptivities have units of M-1cm-1. +
++
Circular Dichroism – The transformation of plane polarized light into elliptically polarized light
+
+Some asymmetric (chiral) materials possess a special property that they absorb left circularly polarized light (LCPL) to +a different extent than right circularly polarized light(RCPL). Such materials are said to be exhibiting the phenomenon +of Circular Dichroism or CD (The word 'dichroism' means that the material absorbs two different types of light differently). +
++
+Following Beer's law if absorbance of LCPL AL is different from absorbance of RCPL AR then,
+AR ≠AL
+i.e. εRcL ≠εLcL
+i.e. εR ≠ εL
+
+
+
+We can study the effects of circular dichroism as a difference between molar extinction coefficients +(∆ε = εR - εL) of LCPL and RCPL. +
++
+Now let us discuss the results of circular dichroism on a plane polarised light. So if a plane polarised +light enters a a circular dichroic substance its two circular components undergo through separate absorptive +effect of the medium. So the resultant wave will no longer be a linearly polarized wave i.e. the resulting field +vector does not oscillate along a straight line but it rotates along an ellipsoid path. Such a light wave is called +an elliptically polarized light. The direction of rotation of the elliptically polarized light (or, more exactly, its +field vector) is determined by the circular component that remains stronger after traversing the material +
++
+
+
+ source:http://www.enzim.hu/~szia/cddemo/edemo14.htm |
+
+
+
+As is evident from the figure above, one of the axis of the ellipse is parallel to the polarization plane +of the original light wave and another is orthogonal to it. This is the case when the material doesn't show +any refraction. However, there may be a difference between the refractive indices of the material for the RCPL +and LCPL as well. Then the axis of of the elliptically polarised resultant wave undergoes a rotion and no longer +remains parallel to the plane of polarisation of the original wave any more. The material is said to posses both +Circular Birefringence and Circular Dichroism. +
++As we have measured ORD by specific rotation, in this case we also use parameters like delta absorbance, +molar circular dichroism, molar ellipticity, molar residue ellipticity etc to measure the CD. Here are definition, +description and the inter relation of these parameters. +
+Goal of The Experiment
+
This experiment has two parts. The first part elaborates the effect of an anisotropic medium on the velocity of light passing through it. It shows how the plane of polarisation of a plane polarised light changes when RI of its two circularly polarised components are different from each other. It also describes the dispersive phenomena of optical rotation caused by the medium. +
+The second part of the experiment demonestrates the effect of an anisotropic medium on the amplitude of the wave passing through it. It exhibits how state of polarisation of a plane polarised wave changes when rate of absorbance of its two circularly polarised components are different. +
++
+
Notes:
+[*]Geometric explanation of α'= δ/2
+
+
+Disclaimer:
+
+The figures used for explaining phase shift due to Optical Rotation is adopted from
+http://www.pci.uzh.ch/nonella/Anleitungen/ORD.pdf
+The download site of Prof Marco Nonella
+Institute of Physical Chemistry,
+University of Zurich
+
+
')
+
+ t1=content.index('')
+ print t1
+ f.seek(t1+13)
+ s1=s1+f.read()
+ #print s1
+ f.seek(t1+13)
+ f.write(s1)
+ f.seek(0)
+ content=f.read()
+ t= content.index('')
+ f.seek(t+13)
+ s='
' +while sectionNumber<=len(sectionno): + tag="" + att = ''+'experiment-article-section-'+str(sectionNumber)+'-heading' + tagger = soup.findAll('div', attrs={'id':att,'class':'heading'}) + if not tagger: + sectionNumber=sectionNumber+1 + continue + tag1=str(tagger[0].text) + tag1=tag1.strip() + print tag1 + if tag1=='Prerequisite S/W': + tag1='Prerequisites' + st+='
'+heading+'
'+s + s=s+f.read() + f.seek(t+13) + f.write(s) +f=open("template1.html",'r') +template=f.read() +f=open("content.html",'r') +srchtml=f.read() +f.close() +soup = BeautifulSoup(srchtml, 'html.parser') +sectionno=soup.find_all('section') +d=['introduction','theory','objective','experiment','manual','quizzes','further_readings','procedure','hh','ll'] +print len(sectionno) +sectionNumber=1 +st="" + +att = ''+'experiment-article-heading' +tagger = soup.findAll('header', attrs={'id':att,'class':'heading'}) +heading=str(tagger[0].text) +heading=heading.strip() +print heading +att = ''+'experiment-header-heading' +tagger = soup.findAll('div', attrs={'id':att,'class':'heading'}) +heading1=str(tagger[0].text) +print heading +heading1=heading1.strip() +breadcrumb='Chemical Sciences →'+heading1+' →List Of Experiments' +while sectionNumber<=len(sectionno): + tag="" + att = ''+'experiment-article-section-'+str(sectionNumber)+'-heading' + tagger = soup.findAll('div', attrs={'id':att,'class':'heading'}) + if not tagger: + sectionNumber=sectionNumber+1 + continue + tag1=str(tagger[0].text) + tag1=tag1.strip() + print tag1 + if tag1=='Prerequisite S/W': + tag1='Prerequisites' + st+='
'+tag1+'
' + sectionNumber=sectionNumber+1 +st+='Feedback
' +sectionNumber=1 +while sectionNumber<=len(sectionno): + tag="" + tag="" + att = ''+'experiment-article-section-'+str(sectionNumber)+'-heading' + tagger = soup.findAll('div', attrs={'id':att,'class':'heading'}) + if not tagger: + sectionNumber=sectionNumber+1 + continue + tag1=str(tagger[0].text) + tag1=tag1.strip() + if tag1=='Prerequisite S/W': + tag1='Prerequisites' + att = ''+'experiment-article-section-'+str(sectionNumber)+'-content' + tagger = soup.findAll('div', attrs={'id':att,'class':'content'}) + + tag+=str(tagger[0]) + #print tag + #print st + + + writefile(tag1+'.html',tag,st,heading) + #print sectionNumber + sectionNumber=sectionNumber+1 + +f=open("Feedback.html",'w+') +f.write(template) +f.seek(0) + +content = f.read() +content=content.replace('Disciplines and Domains',breadcrumb) +f.seek(0) +f.write(content) +#print content +f.seek(0) +content=f.read() +k=content.index('')
+
+t1=content.index('')
+print t1
+f.seek(t1+13)
+st=st+f.read()
+#print s1
+f.seek(t1+13)
+f.write(st)
+f.seek(0)
+content=f.read()
+t= content.index('')
+f.seek(t+13)
+s='Welcome to Virtual Labs - A MHRD Govt of india Initiative
+
+
+
+
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+Welcome to Virtual Labs - A MHRD Govt of india Initiative
+
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+Welcome to Virtual Labs - A MHRD Govt of india Initiative
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+Welcome to Virtual Labs - A MHRD Govt of india Initiative
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+Welcome to Virtual Labs - A MHRD Govt of india Initiative
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+Welcome to Virtual Labs - A MHRD Govt of india Initiative
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+Welcome to Virtual Labs - A MHRD Govt of india Initiative
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+Welcome to Virtual Labs - A MHRD Govt of india Initiative
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'+heading+'
'+'Feedback' +s=s+f.read() +f.seek(t+13) +f.write(s) diff --git a/src/lab/exp2/template1.html b/src/lab/exp2/template1.html new file mode 100644 index 0000000..382fb88 --- /dev/null +++ b/src/lab/exp2/template1.html @@ -0,0 +1,205 @@ + + + + + + + + + + +
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diff --git a/src/lab/exp3/Experiment.html b/src/lab/exp3/Experiment.html
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+ Disciplines and Domains
+ +
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diff --git a/src/lab/exp3/Feedback.html b/src/lab/exp3/Feedback.html
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+ Chemical Sciences →Circular Dichroism Virtual Lab →List Of Experiments
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+ Chemical Sciences →Circular Dichroism Virtual Lab →List Of Experiments
+
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+ To study the Optical Rotatory Dispersion (ORD) of some chiral substances.
+
+
Click here to start the experiment.
++
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diff --git a/src/lab/exp3/Introduction.html b/src/lab/exp3/Introduction.html
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+ +
+ Chemical Sciences →Circular Dichroism Virtual Lab →List Of Experiments
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+ Chemical Sciences →Circular Dichroism Virtual Lab →List Of Experiments
+
+
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+ To study the Optical Rotatory Dispersion (ORD) of some chiral substances.
Feedback
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diff --git a/src/lab/exp3/Manual.html b/src/lab/exp3/Manual.html
new file mode 100644
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+ +
+ Chemical Sciences →Circular Dichroism Virtual Lab →List Of Experiments
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+
+ Chemical Sciences →Circular Dichroism Virtual Lab →List Of Experiments
+
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+ To study the Optical Rotatory Dispersion (ORD) of some chiral substances.
+
+
+
+In Experiment 2 we have seen how an electro magnetic wave interacts with an anisotropic medium. +We have observed the phenomena called Optical Rotatory Dispersion where the optical rotation of +a plane polarised light is studied as a function of wave length of the incident light. Now we will +try to understand how the ORD spectra can be used to study the structure of an optically active substance. +We will study different types of normal and abnormal ORD spectra and try to correlate them with structural +properties of the chiral molecule. +
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diff --git a/src/lab/exp3/Objective.html b/src/lab/exp3/Objective.html
new file mode 100644
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+ +
+ Chemical Sciences →Circular Dichroism Virtual Lab →List Of Experiments
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+ Chemical Sciences →Circular Dichroism Virtual Lab →List Of Experiments
+
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+ To study the Optical Rotatory Dispersion (ORD) of some chiral substances.
+
+Follow the steps given below to successfully complete the experiment.
+
-
+
- +In the “Understanding Structure of Chiral Molecules” step you will be given four samples. You can click on any sample and study its structure and conformation. + +
- +Once you are satisfied you can click "Next" to go to the next Step. + +
- +Next Step is "Assigning R & S Configuration". Here you are given structures of four samples as individul Jmol applets. The Table "Jmol Instructions" provides guide lines to study those structures efficiently. You have to decide whether the structure is a R or a S enantiomer and select the corresponding Radio Button for each structure. Once you decide and choose your option for all the four structures you should submit your answer. You will be informed whether your decission is correct or not. You can skip this step at any point of time by clicking "Skip". To decide whether the structure is R or S you may follow the following instructions + +
- + First look at the atoms that are directly attached to the stereocenter of the compound. A substituent that has a higher atomic number takes precedence over a substituent that has a lower atomic number. Hydrogen is the lowest priority substituent. When you are dealing with isotopes, the atom with the higher atomic mass received higher priority. + +
-
+Note: The lowest priority substituents should always go away from the viewer When you have two substituents that have equal rank, we proceed along the two respective substituent chains until we find a point of difference.
+
+ Note: Keep in mind that priority is determined by the first point of difference along the two similar + substituent chains. After you have reached the first point of difference, the rest of the chain is irrelevant. +
+ - + When you are looking for the first point of difference on similar substituent chains, you may encounter branching. If there is branching, we choose the branch that is higher in priority. When the two substituents have similar branches, you rank the elements within the branches until you reach a point of difference. + +
- + When you come across double and triple bonds, treat them as if they were the same number of single bonds connected to that atom instead. Think of the atoms as duplicated or triplicated: + +
- + After all your substituents have been prioritized in the correct manner, you can name it either R or S. By putting the lowest priority substituent in the back (dashed line), the rotation of the other three substituents will help determine whether the molecule is R or S. In the order of priority (1, 2, 3), if the molecule rotates to the right, it is an R. Conversely, if it rotates to the left, it is considered a S. + +
- +In next step you will be measuring the ORD spectrum of different chiral samples. First click the "Switch the ORD mode" button. The samples will be activated for selecting. Select any sample and "Measure ORD". The ORD spectrum will be shown on screen. You can “Measure Again” or skip the step. + +
- +Next Step is the "Analysis" step. At the left side of your screen you are given a structure and you are provided with a prior knowledge that "R-spiroacytal has -ve ORD curves". Now you have to decide among the four given ORD spectra the given structure matches with which which one. Once you select one graph you should submit your result and you will informed whether you are right or wrong. If you do correctly you can load the next sample. You can skip this step also at any point of time by clicking "Skip". Otherwise you will be asked to perform the analysis for all four samples. +
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diff --git a/src/lab/exp3/Quizzes.html b/src/lab/exp3/Quizzes.html
new file mode 100644
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+++ b/src/lab/exp3/Quizzes.html
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+ +
+ Chemical Sciences →Circular Dichroism Virtual Lab →List Of Experiments
+
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+
+ Chemical Sciences →Circular Dichroism Virtual Lab →List Of Experiments
+
+
+
+
+
+
+
+
+
+ To study the Optical Rotatory Dispersion (ORD) of some chiral substances.
+
+
+ The purpose of this lab is to make students familiar with different types of ORD + spectra obtained from different chiral substances. By doing this experiment user will be able to, +
++
-
+
- + +Develop an understanding about assigning R and S configuration to chiral structures. + +
- + +Relate plain ORD curves and their sign with absolute configuration of sample. + +
- + +Understand why anomalous ORD or Cotton effect is observed. + +
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diff --git a/src/lab/exp3/Theory.html b/src/lab/exp3/Theory.html
new file mode 100644
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+++ b/src/lab/exp3/Theory.html
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+ +
+ Chemical Sciences →Circular Dichroism Virtual Lab →List Of Experiments
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+
+ Chemical Sciences →Circular Dichroism Virtual Lab →List Of Experiments
+
+
+
+
+
+
+
+
+
+ To study the Optical Rotatory Dispersion (ORD) of some chiral substances.
+Click here to have a self evaluation.
+
+
Click here to have a self evaluation.
+
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diff --git a/src/lab/exp3/scraplab.py b/src/lab/exp3/scraplab.py
new file mode 100644
index 0000000..79d06d8
--- /dev/null
+++ b/src/lab/exp3/scraplab.py
@@ -0,0 +1,124 @@
+from bs4 import BeautifulSoup
+heading=""
+breadcrumb=""
+def writefile(fname,s,s1,heading):
+ f=open(fname, 'w+')
+ f.write(template)
+ f.seek(0)
+
+ content = f.read()
+ content=content.replace('Disciplines and Domains',breadcrumb)
+ f.seek(0)
+ f.write(content)
+ print content
+
+ f.seek(0)
+ content=f.read()
+ k=content.index('
+
+
+
+
+
+
+
+
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+ +
+ Chemical Sciences →Circular Dichroism Virtual Lab →List Of Experiments
+
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+
+ Chemical Sciences →Circular Dichroism Virtual Lab →List Of Experiments
+
+
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+
+ To study the Optical Rotatory Dispersion (ORD) of some chiral substances.
+
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+In the previous experiment we have seen that optical rotation of a plane polarised light changes with the wave length of the light when it passes through an anisptropic medium. We measure this change in optical rotation per unit length of the medium as specific rotation. In ORD spectroscopy we change the wavelength of the incident light and measure the change in specific rotation of the light. So an ORD spectrum will give us a plot of specific rotation (some times molar specific rotation) against wavelength. Depending on the nature of the chiral center different anisotropic medium produces different ORD spectrum. Hence ORD spectra may be used as a very efficient tool to determine the nature of chirality in an anisotropic medium.
+
ORD spectra
++Now let us see how different types of ORD spectra represents different physical properties of a medium. +
+
+Dispersion and Plain ORD Curve
+These curves are observed for transparent compounds in the UV range.A plain curve results showing a steady increase (or decrease) of optical rotation with decrease in wavelength. These curves does not cross the zero-rotation axis and are devoid of maxima or minima within measurable range. So-called plain curve is the ORD for a chiral compound that lacks a chromophore. For Example, alcohols and hydrocarbons exhibit such kind of ORD curves.
+
+ |
+
+
+Another Property, Absorbance : Anomalous ORD Curve +
++We have mentioned the term chromophore. It is the part of a molecule where absorbance of light takes place. This occurs because at this part +energy of electronic transition between two molecular orbitals +with energy of the photons of the incident light. Incident light that hits the chromophore can thus be absorbed by exciting an electron from its +ground state into an excited state. So there is a range of wavelength,called the absorption band, where the light is absorbed. This absorption band +has a maxima at a particular wavelength λmax. As a result there comes an anomaly in the ORD curve at the neighborhood of the chromophore. The absolute magnitude of the optical rotation at first varies rapidly with wavelength, crosses zero at absorption maxima and then again varies rapidly with wavelength but in opposite direction. Such anomaly is known as Cotton effect. As opposed to plain curves, these curves exhibit a number of extreme peaks and troughs depending on the number of absorbing groups. The Cotton effect is called positive if the optical rotation first increases as the wavelength decreases (as first observed by Cotton), and negative if the rotation first decreases. The following figure exhibits both types of Cotton effects. +
++
+ + |
+
Figure shows absorption band and -ve & +ve cotton effect near the absorption maxima
+ ++
+Octant Rule: an empirical method to determine type of Cotton effect in Ketones
+In case of saturated alkyl ketones there is a way, known as octant rule, to predict the sign of cotton effect (+ve or -ve) directly from the understanding of the structure. We can divide the space around the ketone into eight octants by looking down the carbonyl as shown below.
+ + |
+
+
Four octants in the front of the oxygen (toward the observer) and four behind. The front four can be ignored but the rear four usually contain all the atoms in the molecule. Each of these four rear octants are assigned a sign(+ ve or -ve) and all the atoms of the molecule are distributed in these four octants. Now depending on the occupancy of these four octants the sign of the cotton effect may be determined. The sign of the octant having highest occupancy determines the sign of the cotton effect. +
++
+
Goal of The Experiment
++
+In this experiment user will observe CD and ORD spectrum of same samples. These samples are chiral molecules with chromophore. It is expected that by doing this experiment the user will develop an understanding of interrelations between ORD and CD spectra. +
++
+
')
+
+ t1=content.index('')
+ print t1
+ f.seek(t1+13)
+ s1=s1+f.read()
+ #print s1
+ f.seek(t1+13)
+ f.write(s1)
+ f.seek(0)
+ content=f.read()
+ t= content.index('')
+ f.seek(t+13)
+ s='
' +while sectionNumber<=len(sectionno): + tag="" + att = ''+'experiment-article-section-'+str(sectionNumber)+'-heading' + tagger = soup.findAll('div', attrs={'id':att,'class':'heading'}) + if not tagger: + sectionNumber=sectionNumber+1 + continue + tag1=str(tagger[0].text) + tag1=tag1.strip() + print tag1 + if tag1=='Prerequisite S/W': + tag1='Prerequisites' + st+='
'+heading+'
'+s + s=s+f.read() + f.seek(t+13) + f.write(s) +f=open("template1.html",'r') +template=f.read() +f=open("content.html",'r') +srchtml=f.read() +f.close() +soup = BeautifulSoup(srchtml, 'html.parser') +sectionno=soup.find_all('section') +d=['introduction','theory','objective','experiment','manual','quizzes','further_readings','procedure','hh','ll'] +print len(sectionno) +sectionNumber=1 +st="" + +att = ''+'experiment-article-heading' +tagger = soup.findAll('header', attrs={'id':att,'class':'heading'}) +heading=str(tagger[0].text) +heading=heading.strip() +print heading +att = ''+'experiment-header-heading' +tagger = soup.findAll('div', attrs={'id':att,'class':'heading'}) +heading1=str(tagger[0].text) +print heading +heading1=heading1.strip() +breadcrumb='Chemical Sciences →'+heading1+' →List Of Experiments' +while sectionNumber<=len(sectionno): + tag="" + att = ''+'experiment-article-section-'+str(sectionNumber)+'-heading' + tagger = soup.findAll('div', attrs={'id':att,'class':'heading'}) + if not tagger: + sectionNumber=sectionNumber+1 + continue + tag1=str(tagger[0].text) + tag1=tag1.strip() + print tag1 + if tag1=='Prerequisite S/W': + tag1='Prerequisites' + st+='
'+tag1+'
' + sectionNumber=sectionNumber+1 +st+='Feedback
' +sectionNumber=1 +while sectionNumber<=len(sectionno): + tag="" + tag="" + att = ''+'experiment-article-section-'+str(sectionNumber)+'-heading' + tagger = soup.findAll('div', attrs={'id':att,'class':'heading'}) + if not tagger: + sectionNumber=sectionNumber+1 + continue + tag1=str(tagger[0].text) + tag1=tag1.strip() + if tag1=='Prerequisite S/W': + tag1='Prerequisites' + att = ''+'experiment-article-section-'+str(sectionNumber)+'-content' + tagger = soup.findAll('div', attrs={'id':att,'class':'content'}) + + tag+=str(tagger[0]) + #print tag + #print st + + + writefile(tag1+'.html',tag,st,heading) + #print sectionNumber + sectionNumber=sectionNumber+1 + +f=open("Feedback.html",'w+') +f.write(template) +f.seek(0) + +content = f.read() +content=content.replace('Disciplines and Domains',breadcrumb) +f.seek(0) +f.write(content) +#print content +f.seek(0) +content=f.read() +k=content.index('')
+
+t1=content.index('')
+print t1
+f.seek(t1+13)
+st=st+f.read()
+#print s1
+f.seek(t1+13)
+f.write(st)
+f.seek(0)
+content=f.read()
+t= content.index('')
+f.seek(t+13)
+s='Welcome to Virtual Labs - A MHRD Govt of india Initiative
+
+
+
+
+
+
+
+
+
+
+
+
+
+
+
+
+
+
+Welcome to Virtual Labs - A MHRD Govt of india Initiative
+
+
+
+
+
+
+
+
+
+
+
+
+
+
+
+
+
+
+Welcome to Virtual Labs - A MHRD Govt of india Initiative
+
+
+
+
+
+
+
+
+
+
+
+
+
+
+
+
+
+
+Welcome to Virtual Labs - A MHRD Govt of india Initiative
+
+
+
+
+
+
+
+
+
+
+
+
+
+
+
+
+
+
+Welcome to Virtual Labs - A MHRD Govt of india Initiative
+
+
+
+
+
+
+
+
+
+
+
+
+
+
+
+
+
+
+Welcome to Virtual Labs - A MHRD Govt of india Initiative
+
+
+
+
+
+
+
+
+
+
+
+
+
+
+
+
+
+
+Welcome to Virtual Labs - A MHRD Govt of india Initiative
+
+
+
+
+
+
+
+
+
+
+
+
+
+
+
+
+
+
+Welcome to Virtual Labs - A MHRD Govt of india Initiative
+
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'+heading+'
'+'Feedback' +s=s+f.read() +f.seek(t+13) +f.write(s) diff --git a/src/lab/exp3/template1.html b/src/lab/exp3/template1.html new file mode 100644 index 0000000..382fb88 --- /dev/null +++ b/src/lab/exp3/template1.html @@ -0,0 +1,205 @@ + + + + + + + + + + +
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diff --git a/src/lab/exp4/Experiment.html b/src/lab/exp4/Experiment.html
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+ Disciplines and Domains
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diff --git a/src/lab/exp4/Feedback.html b/src/lab/exp4/Feedback.html
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+ Chemical Sciences →Circular Dichroism Virtual Lab →List Of Experiments
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+ Chemical Sciences →Circular Dichroism Virtual Lab →List Of Experiments
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+ To study the Circular Dichroism of some substances with chiral chromophores.
+
Simulation Under Construction.
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diff --git a/src/lab/exp4/Introduction.html b/src/lab/exp4/Introduction.html
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+ Chemical Sciences →Circular Dichroism Virtual Lab →List Of Experiments
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+ Chemical Sciences →Circular Dichroism Virtual Lab →List Of Experiments
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+ To study the Circular Dichroism of some substances with chiral chromophores.
Feedback
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diff --git a/src/lab/exp4/Manual.html b/src/lab/exp4/Manual.html
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+ Chemical Sciences →Circular Dichroism Virtual Lab →List Of Experiments
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+ Chemical Sciences →Circular Dichroism Virtual Lab →List Of Experiments
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+ To study the Circular Dichroism of some substances with chiral chromophores.
+
+In Experiment 3 we have seen variation in ORD spectra for different chiral molecules. In this experiment we will observe variation of CD spectra for chiral molecules with chromophores. We will aslo try to understand the relation between anomalous ORD spectra and CD spectra. +
+A CD spectra can be used as an identifying signature of chiral molecules where chromophore is present. For large chiral molecules like protein CD spectroscopy is used as a primary tool for structural characterization. +
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diff --git a/src/lab/exp4/Objective.html b/src/lab/exp4/Objective.html
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+ Chemical Sciences →Circular Dichroism Virtual Lab →List Of Experiments
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+ Chemical Sciences →Circular Dichroism Virtual Lab →List Of Experiments
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+ To study the Circular Dichroism of some substances with chiral chromophores.
+
+The experiment is composed of a number of steps. One may follow the steps given below to successfully run the experiment.
+
-
+
- +In the “Understanding Structure of Chiral Molecules” step you will be given two samples. +You can click on any sample and study its structure and conformation. + +
- +Once you are satisfied you can click “Next” to go to the next Step. + +
- +Next Step is “Assigning R & S Configuration”. Here you are given structures of four samples as individual Jmol applets. +The Table “Jmol Instructions” provides guide lines to study those structures efficiently. You have to decide whether +the structure is a R or a S enantiomer and select the corresponding Radio Button for each structure. Once you decide and +choose your option for both the structures you should submit your answer. You will be informed whether your decission is correct or +not. You can skip this step at any point of time by clicking “Skip”. To decide whether the structure is R or S you may follow the + following instructions + +
-
+First look at the atoms that are directly attached to the stereocenter of the compound.
+A substituent that has a higher atomic number takes precedence over a substituent that has a lower atomic number.
+Hydrogen is the lowest priority substituent. When you are dealing with isotopes, the atom with the higher atomic mass
+received higher priority.
+Note: The lowest priority substituents should always go away from the viewer
+Note: Keep in mind that priority is determined by the first point of difference along the two similar substituent chains. After you have reached the first point of difference, the rest of the chain is irrelevant. +
+
+ - +When you have two substituents that have equal rank, we proceed along the two respective substituent chains until we +find a point of difference. + +
- +When you are looking for the first point of difference on similar substituent chains, +you may encounter branching. If there is branching, we choose the branch that is higher in priority. +When the two substituents have similar branches, you rank the elements within the branches until you reach a point of difference. + +
- +When you come across double and triple bonds, treat them as if they were the same number of single bonds connected to that atom instead. +Think of the atoms as duplicated or triplicated: + +
- +After all your substituents have been prioritized in the correct manner, you can name it either R or S. +By putting the lowest priority substituent in the back (dashed line), the rotation of the other three substituents + will help determine whether the molecule is R or S. In the order of priority (1, 2, 3), if the molecule rotates to the right, + it is an R. Conversely, if it rotates to the left, it is considered a S. + +
- +In next step you will be measuring the CD and ORD spectrum of different chiral samples. First click the "Switch the CD mode" button. + The samples will be activated for selecting. Select any sample and "Measure ORD". The ORD spectrum will be shown on screen. You can + "Measure Again" or skip the step. + +
- +Next Step is the "Analysis" step. At the left side of your screen you are given a picture of two ORD spectrum. And there is one CD +spectrum example on the screen. Now you have to decide among the ftwo given ORD spectra, with which the given CD spectra corresponds + with. Once you select one graph you should submit your result and you will informed whether you are right or wrong. If you do correctly + you can load the next sample. You can skip this step also at any point of time by clicking “Skip”. Otherwise you will be asked to perform the analysis for both the samples. + +
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diff --git a/src/lab/exp4/Quizzes.html b/src/lab/exp4/Quizzes.html
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+ Chemical Sciences →Circular Dichroism Virtual Lab →List Of Experiments
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+ Chemical Sciences →Circular Dichroism Virtual Lab →List Of Experiments
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+ To study the Circular Dichroism of some substances with chiral chromophores.
+
+
+ The purpose of this lab is to make students familiar with different types of CD + spectra obtained from different chiral substances with chromophore. By doing this experiment user will be able to, +
++
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+Develop an understanding about assigning R and S configuration to chiral structures.
+
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+ -
+Be familiar with the nature of CD curves of samples of different chirality.
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+ -
+Understand relation between CD and ORD spectra obtained from the same sample.
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diff --git a/src/lab/exp4/Theory.html b/src/lab/exp4/Theory.html
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+ Chemical Sciences →Circular Dichroism Virtual Lab →List Of Experiments
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+ Chemical Sciences →Circular Dichroism Virtual Lab →List Of Experiments
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+ To study the Circular Dichroism of some substances with chiral chromophores.
+Click here to have a self evaluation.
+
+
Click here to have a self evaluation.
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diff --git a/src/lab/exp4/scraplab.py b/src/lab/exp4/scraplab.py
new file mode 100644
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+from bs4 import BeautifulSoup
+heading=""
+breadcrumb=""
+def writefile(fname,s,s1,heading):
+ f=open(fname, 'w+')
+ f.write(template)
+ f.seek(0)
+
+ content = f.read()
+ content=content.replace('Disciplines and Domains',breadcrumb)
+ f.seek(0)
+ f.write(content)
+ print content
+
+ f.seek(0)
+ content=f.read()
+ k=content.index('
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+ Chemical Sciences →Circular Dichroism Virtual Lab →List Of Experiments
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+ Chemical Sciences →Circular Dichroism Virtual Lab →List Of Experiments
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+ To study the Circular Dichroism of some substances with chiral chromophores.
+
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+Circular dichroism leads to the transformation of plane polarized light into elliptically polarized light, +and is directly related to differential absorbance of LEft Circularly Polarised (LCP) and Right Circularly Polarised (RCP) components of plane polarized light, by +chiral substances which also absorb light. The plot of the variation in the ellipticity, of the elliptically +polarized light, as a function of wavelength, gives us the CD spectrum. In order to understand circular dichroism, +and its role in CD spectroscopy, we need to visit some definitions and concepts. +
++
CD spectroscopy – Concepts, measurable parameters and quantitative relationships
+
As we have discussed CD is the direct consequence of differential absorbance. So studying variation of
+differential absorbance with variation in wavelength is a good approach. We define delta absorbance,
+a parameter to study CD, as
+ΔA=AL-AR
+where AL and AR represents the absorbance of LCP light and RCP light respectively.
+
+Though the the difference in absorption to be measured is very small (usually a few 1/100ths to a few 1/10th +of a percent), but it can be determined quite accurately. +
++A circular dichroism signal can be positive or negative, depending on whether left circularly polarised light +is absorbed to a greater extent than right circularly polarised light (CD signal positive) or to a lesser extent +(CD signal negative). An example CD spectrum of a sample with multiple CD peaks is shown below, demonstrating how +CD varies as a function of wavelength, and that a CD spectrum may exhibit both positive and negative peaks. +
++
+As discussed above, from Beer-Lambert law we can express delta absorbance as
+
+
+ΔA= (εL - εR)Cl
+where, C is the molar concentration, and l is the path length in centimetres(cm).
+
+Therefore we can plot the difference in molar extinction coefficient of LCP and RCP light
+Δε= εL - εR as a function of wavelength. Thus we have an intrinsic
+property called molar circular dichroism, Δε to represent CD spectroscopy.
+
+
+Upto now we have seen CD as an intrinsic property of the molecule but in many practical applications of +circular dichroism (CD), the measured CD may also be a function of temperature, concentration, and the +chemical environment, including solvents. So in those cases we should follow another way to represent CD +spectra. To do that instead of looking at the cause of circular dichroism (differential absorbance) we may +look at the effect of CD (ellipticity). +
++
+Hence we define ellipticity of polarization as the inverse tangent of the quotient of the minor and major axes +of the elliptically polarised wave. And it is derived as, +
+ source:http://en.wikipedia.org/wiki/File:Electric_Vectors_1.png |
+ |
+
+where
+ER and EL are the magnitudes of the electric field vectors of the right-circularly and left-circularly polarized light, respectively.
+
+
+This ellipticity will depend on the path length and concentration of medium. So we can think of specific ellipticity,
+ellipticity per unit length per unit concentration.We can think of an intrinsic property in terms of this ellipticity,
+molar ellipticity ([θ]), which is simply the product of molecular weight of the molecule and specific ellipticity.
+
+[θ] = M x specific ellipticity x 10-2
+
+
+Now the Molar circular dichroism, Δε, and molar ellipticity, [θ], are readily interconvertable by the equation:
+[θ] = 3298.2 ε
+
+After having discussed different measurable parameters related to CD spectra lets have a look at how it is measured in +practical situations. +
++
+Circular dichroism spectra are measured using a CD spectrometer which is a highly specialised derivative of an ordinary +absorption spectrometer. CD spectrometers measure alternately the absorption of left and right circularly polarised lights, +usually at a frequency of 50 kHz, and then calculate the CD signal. CD is reported either in units of ΔA, the delta +absorbance (is discussed below), or in degrees ellipticity. [θ], the molar ellipticity in deg cm2 dmol–1 = 3298.2 δA +which is also discussed in detail below. +
++
+
+Note: One should be very careful with the concentration of the sample taken. There is a misconception +that more the concentration, better the CD signal which is totally incorrect. However, in reality, the reverse happens. +If the concentration is more, less light will pass through it and it will give the CD signal with error. Therefore, an +optimal concentration of the sample should be chosen to obtain an error-free CD signal. +
++
+
CD vs ORD : their interrelation and preference of CD over ORD
+
We have seen Circular dichroism as an absorptive effect and ORD as a dispersive effect of interaction between an EM wave and an anisotropic medium. We also have discussed the measuring parameters of ORD and CD. In case of ORD the molecular rotation [M]Tλ and in case of CD the molar ellipticity [θ]Tλ are the most important parameters. They are also interrelated by Kroning-Kramer transformation as shown below. +
+
+ |
+
+
+
+So we can easily switch between CD and ORD data. One must keep in mind that ORD is observed in any molecule which is +chiral but CD is observed only when there is a chromophore. So when CD is observed a cotton effect must be present there. +
++
+But in practical purpose we prefer CD data over ORD data. There are several reasons behind doing that
+
-
+
- +In ORD if several cotton effects are present then it is difficult to estimate their magnitude, sign and wavelength. +In CD spectra it is much easier to interpret. There exists an one to one correspondence between absorption peaks and +CD signals. + +
- +CD spectra has a better sensitivity. + +
- +Since CD spectra is not spread over a wide range like ORD spectra, it has a better resolution. + +
+
Goal of The Experiment
++In this experiment we have two samples. They are enantiomers of each other. We will learn to determine which one is R and which one is S enantiomer. After that we will measure the ORD and CD spectrum of both the samples. Once we get all the spectrum we will try to match the ORD curve of a smaple to its own CD graph. +
++
')
+
+ t1=content.index('')
+ print t1
+ f.seek(t1+13)
+ s1=s1+f.read()
+ #print s1
+ f.seek(t1+13)
+ f.write(s1)
+ f.seek(0)
+ content=f.read()
+ t= content.index('')
+ f.seek(t+13)
+ s='
' +while sectionNumber<=len(sectionno): + tag="" + att = ''+'experiment-article-section-'+str(sectionNumber)+'-heading' + tagger = soup.findAll('div', attrs={'id':att,'class':'heading'}) + if not tagger: + sectionNumber=sectionNumber+1 + continue + tag1=str(tagger[0].text) + tag1=tag1.strip() + print tag1 + if tag1=='Prerequisite S/W': + tag1='Prerequisites' + st+='
'+heading+'
'+s + s=s+f.read() + f.seek(t+13) + f.write(s) +f=open("template1.html",'r') +template=f.read() +f=open("content.html",'r') +srchtml=f.read() +f.close() +soup = BeautifulSoup(srchtml, 'html.parser') +sectionno=soup.find_all('section') +d=['introduction','theory','objective','experiment','manual','quizzes','further_readings','procedure','hh','ll'] +print len(sectionno) +sectionNumber=1 +st="" + +att = ''+'experiment-article-heading' +tagger = soup.findAll('header', attrs={'id':att,'class':'heading'}) +heading=str(tagger[0].text) +heading=heading.strip() +print heading +att = ''+'experiment-header-heading' +tagger = soup.findAll('div', attrs={'id':att,'class':'heading'}) +heading1=str(tagger[0].text) +print heading +heading1=heading1.strip() +breadcrumb='Chemical Sciences →'+heading1+' →List Of Experiments' +while sectionNumber<=len(sectionno): + tag="" + att = ''+'experiment-article-section-'+str(sectionNumber)+'-heading' + tagger = soup.findAll('div', attrs={'id':att,'class':'heading'}) + if not tagger: + sectionNumber=sectionNumber+1 + continue + tag1=str(tagger[0].text) + tag1=tag1.strip() + print tag1 + if tag1=='Prerequisite S/W': + tag1='Prerequisites' + st+='
'+tag1+'
' + sectionNumber=sectionNumber+1 +st+='Feedback
' +sectionNumber=1 +while sectionNumber<=len(sectionno): + tag="" + tag="" + att = ''+'experiment-article-section-'+str(sectionNumber)+'-heading' + tagger = soup.findAll('div', attrs={'id':att,'class':'heading'}) + if not tagger: + sectionNumber=sectionNumber+1 + continue + tag1=str(tagger[0].text) + tag1=tag1.strip() + if tag1=='Prerequisite S/W': + tag1='Prerequisites' + att = ''+'experiment-article-section-'+str(sectionNumber)+'-content' + tagger = soup.findAll('div', attrs={'id':att,'class':'content'}) + + tag+=str(tagger[0]) + #print tag + #print st + + + writefile(tag1+'.html',tag,st,heading) + #print sectionNumber + sectionNumber=sectionNumber+1 + +f=open("Feedback.html",'w+') +f.write(template) +f.seek(0) + +content = f.read() +content=content.replace('Disciplines and Domains',breadcrumb) +f.seek(0) +f.write(content) +#print content +f.seek(0) +content=f.read() +k=content.index('')
+
+t1=content.index('')
+print t1
+f.seek(t1+13)
+st=st+f.read()
+#print s1
+f.seek(t1+13)
+f.write(st)
+f.seek(0)
+content=f.read()
+t= content.index('')
+f.seek(t+13)
+s='Welcome to Virtual Labs - A MHRD Govt of india Initiative
+
+
+
+
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+Welcome to Virtual Labs - A MHRD Govt of india Initiative
+
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+Welcome to Virtual Labs - A MHRD Govt of india Initiative
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+Welcome to Virtual Labs - A MHRD Govt of india Initiative
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+Welcome to Virtual Labs - A MHRD Govt of india Initiative
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+Welcome to Virtual Labs - A MHRD Govt of india Initiative
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+Welcome to Virtual Labs - A MHRD Govt of india Initiative
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+Welcome to Virtual Labs - A MHRD Govt of india Initiative
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'+heading+'
'+'Feedback' +s=s+f.read() +f.seek(t+13) +f.write(s) diff --git a/src/lab/exp4/template1.html b/src/lab/exp4/template1.html new file mode 100644 index 0000000..382fb88 --- /dev/null +++ b/src/lab/exp4/template1.html @@ -0,0 +1,205 @@ + + + + + + + + + + +
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diff --git a/src/lab/exp5/Experiment.html b/src/lab/exp5/Experiment.html
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+ Disciplines and Domains
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diff --git a/src/lab/exp5/Feedback.html b/src/lab/exp5/Feedback.html
new file mode 100644
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+ Chemical Sciences →Circular Dichroism Virtual Lab →List Of Experiments
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+ Chemical Sciences →Circular Dichroism Virtual Lab →List Of Experiments
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+ To Study the operational details of a CD spectrometer.
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diff --git a/src/lab/exp5/Introduction.html b/src/lab/exp5/Introduction.html
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+ Chemical Sciences →Circular Dichroism Virtual Lab →List Of Experiments
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+ Chemical Sciences →Circular Dichroism Virtual Lab →List Of Experiments
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+ To Study the operational details of a CD spectrometer.
Feedback
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diff --git a/src/lab/exp5/Manual.html b/src/lab/exp5/Manual.html
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+ Chemical Sciences →Circular Dichroism Virtual Lab →List Of Experiments
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+ Chemical Sciences →Circular Dichroism Virtual Lab →List Of Experiments
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+ To Study the operational details of a CD spectrometer.
+
+ As we are already familiar with the concepts of circular dichroism, ellipticity etc. from the previous experiments, here we are going to provide only a brief review of the CD.
+CD instruments (known as spectropolarimeters) measure the difference in absorbance between the L and R circularly polarized components (ΔA = A L - A R), and generally report this in terms of the ellipticity (θ) in degrees. It should be noted that θ = tan-1 (b/a) where b and a are the minor and major axes of the resulting ellipse . There is a simple numerical relationship between ΔA and ellipticity (in degrees), namely θ = 32.98 ΔA. The CD spectrum is obtained when the dichroism is measured as a function of wavelength.
+In a CD instrument, plane polarised light is split into the L and R components by passage through a modulator subjected to an alternating electric field (50 kHz is the frequency most commonly employed). The modulator normally used consists of a piezoelectric quartz crystal and a thin plate of isotropic material (e.g., fused silica) tightly coupled to the crystal. The alternating electric field induces structural changes in the quartz crystal; these make the plate transmit circularly polarised light at the extremes of the field. As the transmitted radiation is switched between L and R components, these are detected in turn by the photomultiplier tube.
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diff --git a/src/lab/exp5/Objective.html b/src/lab/exp5/Objective.html
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+ Chemical Sciences →Circular Dichroism Virtual Lab →List Of Experiments
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+ Chemical Sciences →Circular Dichroism Virtual Lab →List Of Experiments
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+ To Study the operational details of a CD spectrometer.
+
+Module:1
+
Step:1
+Click on the regulator of the N2 gas cylinder to open the gas for purging.
+Step:2
+Click on the knob present on the cylinder to control the flow rate of N2 gas according
+to short wave length limit of measurement.
+Step:3
+After setting the flow rate now turn on the CD spectrometer by clicking the green button present on the left side of the machine.
+Step:4
+Now turn on the computer by clicking the CPU power button.
+Step:5
+Now turn on the water bath by clicking on the power button and set the temperature to 20°C for normal measurement.
+Step:6
+Now turn on the peltier temperature control(PTC) to regulate the temperature and stirrer revolution for the sample by clicking the power button.After turning on PTC, click the start button to activate the PTC.
+The PTC has two mode:
+(i)Stirrer mode:Regulate the stirrer revolution rate by scrolling the slider present on the PTC.
+(ii)Temperature mode:Regulate the temperature of the sample by clicking the Ctrl and +/- arrow keys.
+After setting the stirrer revolution rate and temperature click on the Enter button.
+Step:7
+Now click on the lid of the CD spectrometer,the instrument will now
+ undergo its initialization process. Once the initialization is completed, the lid will open and the lamp will be
+ turned on.
+
+Step:8
+After the initialization of the CD spectrometer now it is ready for the spectrum measurement.
+
+Module:2
+
+Step:1
+First click on beaker ,then on the sample solution to bring it to the table.
+Step:2
+Then pour the sample solution into the beaker.
+Step:3
+Click on the pipette to bring it to the experiment table, then click the pipette to fill it with the sample solution.
+Step:4
+Now click on cuvette to bring it to the experiment table,and then click the pipette to fill the solution into the cuvette.
+Step:5
+Now click the cuvette to place into the cuvette holder of the CD spectrometer.
+Step:6
+Now the scan will take place and the spectrum for the sample will be recorded.The detail
+process of the modulation and the detection of the CD signal is shown in the simulated diagram.
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diff --git a/src/lab/exp5/Quizzes.html b/src/lab/exp5/Quizzes.html
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+ Chemical Sciences →Circular Dichroism Virtual Lab →List Of Experiments
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+ Chemical Sciences →Circular Dichroism Virtual Lab →List Of Experiments
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+
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+ To Study the operational details of a CD spectrometer.
+
+
+The objective of this experiment is to make the user/student understand the working of CD spectrometer, its different parts and how to set the important parameters related to CD spectrometer initialization and spectrum measurement.Unfortunately, it is not always clear that such instruments are being used to their best advantage so the aim of this experiment is to provide a brief summary of the CD spectrometer functioning and some important practical aspects of performing CD experiments. +
+
This experiment have three modules:
+(1) CD spectrometer initialization:How to set up the CD spectrometer for initialization
+(2) Sample preparation:How to prepare a sample for the CD spectrum measurement
+(3) Graph section:This section will demonstrate real CD spectra of a protein(Lysozyme) along with a buffer at different concentration and cell length.
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diff --git a/src/lab/exp5/Theory.html b/src/lab/exp5/Theory.html
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+ Chemical Sciences →Circular Dichroism Virtual Lab →List Of Experiments
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+ Chemical Sciences →Circular Dichroism Virtual Lab →List Of Experiments
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+ To Study the operational details of a CD spectrometer.
+
+
+
+
+Here are two sets of interactive questions to help you to make your understanding more clear.
+After you go through the theory section you should try out the Pre-experiment quiz before you start the actual experiment.
+Once you perform the original experiment you should try out the Post-experiment quiz to have a better insight of the concepts.
+
+
Pre-experiment Quiz
+
Click here to have a self evaluation.
+
Post-experiment Quiz
+
Click here to have a self evaluation.
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diff --git a/src/lab/exp5/scraplab.py b/src/lab/exp5/scraplab.py
new file mode 100644
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+++ b/src/lab/exp5/scraplab.py
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+from bs4 import BeautifulSoup
+heading=""
+breadcrumb=""
+def writefile(fname,s,s1,heading):
+ f=open(fname, 'w+')
+ f.write(template)
+ f.seek(0)
+
+ content = f.read()
+ content=content.replace('Disciplines and Domains',breadcrumb)
+ f.seek(0)
+ f.write(content)
+ print content
+
+ f.seek(0)
+ content=f.read()
+ k=content.index('
+
+
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+ +
+ Chemical Sciences →Circular Dichroism Virtual Lab →List Of Experiments
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+
+ Chemical Sciences →Circular Dichroism Virtual Lab →List Of Experiments
+
+
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+ To Study the operational details of a CD spectrometer.
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+Introduction to a general layout of a spectrometer +
+ ++The basic CD spectrometer design most widely adopted by the leading manufacturers comprises a high intensity broadband light source (usually a high pressure Xenon arc lamp), a double prism monochromator which not only serves to disperse the wavelength but is also used to linearly polarise the light beam, an electro-optic modulator which transforms the linearly polarised light into circularly polarised states, a sample chamber and photomultiplier detector. Sophisticated control and signal processing electronics, all under PC control, completes a typical instrument (Figure below) +
+
+
+
+
+Block diagram of a spectropolarimeter. Plane polarised +radiation is produced by passage of light from the source (LS) through 2 +prisms (P1 and P2) and a series of mirrors (M0 to M5) and slits (S1 to S3). +The ordinary ray (O) is focussed by a lens (L), and passed through a filter +(F) to the modulator (CDM). The circularly polarised components are then +passed through the shutter (SH) to the sample compartment, before +detection by the photomultiplier (PM). (E represents the extra-ordinary ray). +
++
+
LS light source, is a 150W, air cooled Xe lamp, with quartz envelope. It emits a strong continuous spectra in the all wavelength range of the monochromator (163-1100 nm)
+
M1 is an elliptical mirror focusing the lamp arc on the entrance slit (S1) of the double monochromator
+
M0 is the mirror to collect back-emitted radiation from the source
+
S1,S2,S3 are the entrance, intermediate and exit slits of the monochromator, all linked together
+
M2-M5 are the spherical collimator mirrors of the double monochromator (Czerny Turner type)
+
P1,P2 are the quartz prisms of the monochromator, prisms are used to disperse radiation and also to get linear polarization output. E-ray (extraodinary ray with crossed polarization) is not getting +out
+
L is the focusing lens to get beam collimated in the sample compartment
+
F is a series of 45° quartz plates to clean further the monochromatic exit beam polarization
+
CDM is the photoelastic modulator crystal in its thermostatted housing, it acts (in CD mode) as a 1/4 wave achromatic plate, i.e. the linear polarized beam is deflected + and – 45° at the crystal +oscillation frequency (50 KHz)
+
SH is a mechanical shutter to remove radiation from sample (when needed)
+
PM is the photomultiplier tube detector
+ +
+
+=> As white light passes through the first prism it is split into
+its component wavelengths and the wavelength selected by the intermediate
+slit.
+
+=> The prism also separates the linear polarised components of light.
+The pointed ray is the vertically polarised component and the semi-solid ray is the
+horizontally polarised component
+
+=> What actually passes through the intermediate slit is a mixture
+of horizontally polarised light of the selected wavelength and vertically
+polarised light of different wavelength.
+
+=> The second prism separates out the contaminating vertically polarised
+beam, as well as straylight that has made it through the first monochromator.
+
+=> The unique combination of the two polarising quartz prisms provides
+a wide separation of polarised components, allowing wider slit widths, so
+more light, to be used.
+
+=> The polarised light is passed through a photoelastic modulator(CDM)
+which converts the beam into alternating left and right circularly
+polarised light.
+
+
+
+Polarisation modulation
+
+The measurement of CD requires the generation of a monochromatic probe light beam in each of the left circularly polarised (CPL) and right circularly polarised (CPR) states and a means of detecting the difference in absorbance of the two states caused by the introduction of a sample into the light beam. The differential absorbance of a typical sample with respect to the two circularly polarised states is typically in the order of 1 part in 104 or less. Therefore the differences to be measured in the CPL and CPR photometric signals are also of this order in relation to the transmission background of the sample. The most reliable mechanism developed to date for measuring such fine differences in sample absorbance utilises a modulation technique whereby the light beam from the monochromator is alternately switched at a high frequency (typically 50 kHz) between the CPL and CPR states using a photo-elastic modulator (CDM). Following transmission through the sample, the beam strikes a photomultiplier detector which converts the incident photon flux to a photometric current.Any sample CD results in a small 50 kHz AC component superimposed on a much larger DC background resulting from the standard transmission of the sample. This AC amplitude is detected using a tuned amplifier and demodulation circuit and the CD is derived from the ratio of the AC and DC amplitudes. This optical modulation approach has the benefit of allowing sensitive detection in a single channel and also inherent rejection of noise and drift away from the modulation frequency. +
++
+
+Initialization of the CD spectrometer
+
+(1)Purging the instrument with N2:
+The need to measure very small signals in the far-UV has been the biggest influence driving improvements in instrument specification over the years and significant increases to the limit of detection and far-UV performance have been achieved. Specifically, the most critical requirement for all CD instruments is to maximise the intensity and quality of circularly polarized light available, particularly in the far-UV, down to what is the experimentally practical limit of 170-175 nm. 190 nm and below is the most demanding region of the spectrum in which the instrument must operate.
+At this wavelength the instrument must be purged with high-purity oxygen-free nitrogen (approx 3–5 L min) for at least 20 min before starting the light source and while making measurements. Any oxygen present may be converted to ozone by far-UV light from the high intensity arc, and ozone will damage optical surfaces. Higher nitrogen flow rates should be used for measurements below 190 nm.
+
+(2)Controlling the temperature of the sample:External water bath and Peltier temperature control +Accurate and repeatable temperature control of the sample is a crucial need for any spectrometer system and even more important with CD due to the heavy use in measurement of biomolecules. +In the CD spectrometer generally the circulator water bath(temperature range 20-200°C) based peltier temperature control system is used to control the the temperature of sample placed inside the cell. +The Peltier thermostatted cell holder which enables temperature control using the temperature control program. It controls the sample temperature to measure the CD spectrum with change in temperature, generally in the range of -10 to +110°C . +
++
+Peltier cooling attachment is based on the Peltier effect(The Peltier effect occurs whenever electrical current flows through two dissimilar conductors(or metals). Depending on the direction of current flow, the junction of the two conductors will either absorb or release heat.) +
++Cooling, Heating: When DC voltage is applied to the module, the positive and negative charge carriers in the pellet array absorb heat energy from one substrate surface and release it to the substrate at the opposite side. The surface where heat energy is absorbed becomes cold; the opposite surface where heat energy is released, becomes hot. Using this simple approach to "heat pumping," thermoelectric technology is applied to many widely-varied applications—small laser diode coolers, portable refrigerators, scientific thermal conditioning, liquid coolers, and beyond. +
++
+ +Schematic of a simple Peltier cooler (Source:See Ref:6b) + |
+
+
+The ability to measure CD spectra while controlling the temperature of a sample offers important information about conformational change associated with temperature and is used in the study of biopolymers. The temperature control program measures the thermal denaturation curve (temperature scan) and CD spectra during temperature change based on user selected parameters. +
++
+(3)Solution preparation for CD spectrum measurement
+
+(i)Selection of protein:
+Samples should, of course, be of he highest possible purity. Near-UV CD signals,
+in particular, can be seriously distorted by the presence of relatively small
+amounts of protein impurities (if they have intense signals) and by the presence
+of nucleic acids, which have intense CD bands in this region.
+
+The protein should be free from other aggregates, as aggregated proteins can cause optical artefacts in CD such as differential light scattering and absorption flattening.
+
+Far-UV CD spectra of proteins (260–178 nm) are intense and small amounts of material are required to record them. Because all peptide bonds contribute to the spectrum the amount of material required is effectively the same for any protein. Typical quantities are 200 μL of a 0.1– 0.15 mg/mL solution with a 1-mm path length cuvet or 30 μL of a 1.0–1.5 mg/mL solution with a 0.1 mm (demountable) cuvet. The latter is preferable for good far-UV penetration but the material is not generally recoverable.
+
+
+Near-UV CD spectra (340–255 nm) are much less intense than far-UV spectra and recording them requires more material. Spectra are usually recorded under +conditions similar to those used for measuring a conventional absorption spectrum, e.g., use a 10-mm cuvet and aim for an absorbance at 280 nm in the range 0.7–1.0. Less-concentrated solutions may be used if the CD signals are intense. +
++
+CD signals will be seriously distorted if too little light reaches the photomultiplier. In practical terms, this means that one cannot make reliable measurements on samples with an absorbance (sample plus solvent) much greater than 1. The absorption spectrum of the sample should always be checked to see if this absorbance limit is exceeded. In far-UV measurements, the absorbance of the protein itself is generally rather small and the major problems arise from absorption by buffer components, almost all of which will limit far-UV penetration to some extent. +
+
+
+(ii)Selection of Solvent/buffer:
+The aim in recording CD spectra of good quality is to keep overall absorbance within reasonable bounds(less than 1). If the absorbance will be too high then the relatively small photon flux reaching the detector will lead to high errors in measuring the small values of absorbance and will generate high levels of noise relative to signal. Therefore to obtain better results the concentration of sample must be kept high as to that of the solvent system. It may also be necessary to maintain the ionic strength of the sample and to add reducing or chelating agents to improve the stability of protein. Oxygen absorbs light below 200 nm; for
+optimum transparency, buffers should therefore be prepared with glass-distilled water or the
+water should be degassed before use.
+
+
+
+It is essential to run a blank spectrum with buffer or solvent system alone in order to check that the absorbance and High Tension voltage is not excessive. If that happens then it may be possible to use cells of shorter path length, provided that the concentration of sample can be increased or lower teh concentration of species which is causing problem(e.g. Chloride ions). Selection of buffer provides denaturation of proteins and peptides. The buffers also show cutoffs at a certain point in low wavelength region where they start to absorb after certain point, therefore cutoff values of certain common buffers should be known. Use of optically active buffers should be avoided as it can provide inaccurate results. Therefore clear solutions are required and CD is taken in high transparency quartz cuvettes to ensure least interference. +
+Buffer components (especially those that contain carboxylates) that have a high absorbance below 200 nm must be avoided (e.g. Tris [tris(hydroxymethyl)aminomethane], dithiothreitol, imidazole, histidine, choloride, etc.). The following table describes the cutoff wavelength (wavelength in which absorbance = 1 for a 1 mm cuvette) for common solvents and buffer components. Cuvettes with short pathlengths are better, since the amount of buffer the light travels through is minimized. The disadvantage of short pathlength cuvettes is the higher concentration of protein which must be used (increased 10-fold relative to a 1 cm cell). Ideally, the buffer concentration is low (< 10 mM). +
++ |
++ |
+
| Water | +<185 | +
| Trifluoroethanol | +<185 | +
| Hexafluoroisopropanol | +<185 | +
| Acetonitrile | +185 | +
| Methanol | +195 | +
| Ethanol | +196 | +
| 2-Propanol | +196 | +
| Cyclohexane | +<185 | +
| Dioxane | +232 | +
| (NH4)2SO4 0.15 M | +191 | +
| NaCl 0.15 M | +196 | +
| NaClO4 0.15 M | +<185 | +
| NaNO3 0.15 M | +245 | +
| Phosphate 100 mM | +<185 | +
| Tris 100 mM | +195 | +
| Pipes 100 mM | +215 | +
| Mes 100 mM | +205 | +
| GdnHCl 4 M | +210 | +
| Urea 4 M | +210 | +
+
* For a one mm path length
+
+
+(Source:See Ref:4)
++
+
+
+
| Buffer component | +180nm | +190nm | +200nm | +210nm | +
| NaCl | +>0.5 | +>0.5 | +0.02 | +0 | +
| NaF | +0 | +0 | +0 | +0 | +
| Na borate(pH 9.1) | +0.3 | +0.09 | +0 | +0 | +
| Na2HPO4 | +>0.5 | +0.3 | +0.05 | +0 | +
| NaH2PO4 | +0.15 | +0.01 | +0 | +0 | +
| Na acetate | +>0.5 | +>0.5 | +0.17 | +0.03 | +
| Tris/H2SO4(pH 8.0) | +>0.5 | +0.24 | +0.13 | +0.02 | +
| HEPES/Na+ (pH 7.5) | +>0.5 | +>0.5 | +0.5 | +0.37 | +
(Source:See Ref:5)
+
++
+Chloride and carboxylate ions absorb strongly below 195nm. Therefore high concentrations of chloride ions arising from Tris/HCl buffers should be avoided and instead flouride and sulphate ions can be used to maintain ionic strength. +Buffers based on phosphate, borate or Tris are suitable for maintaining pH values in range from about 6.5-9.5 and buffers in pH range 4 to 6 are usually based on carboxylate groups or sulphonic acid derivatives and can only be used at low concentrations(<10mM). Organic solvents are used while dealing with very non-polar integral membrane proteins(generally in case of natural products or drugs). +
++
+(iii)Sample concentration to be used
+Solutions should be prepared according to the pathlength of cuvette
+
| +Cuvette path(cm) + | ++Concentration of sample(mg/ml) + | +
| 0.01-0.02 | +0.2-1.0 | +
| 0.1 | +0.05-0.2 | +
| 1 | +0.005-0.01 | +
+(Source:See Ref:3) +
+ ++
+The most accurate method of determining protein concentration is quantitative amino acid analysis, using the concentrations of the stable amino acids, e.g. alanine and lysine to calculate the concentration of the intact protein. This method is very sensitive and can be performed on actual CD samples. Concentration can be determined using published molar extinction coefficients if they are available; the protein should be dialyzed or desalted into the CD buffer immediately before the spectrum is obtained and filtered though 0.1 to 0.2 micron filters to reduce light scattering. The spectrum of the CD buffer alone must be subtracted from the +spectrum of the sample. Since CD is usually carried out on samples with relatively low concentrations, it is best to prepare a filtered stock solution of the protein of ~1 mg/ml in the CD buffer and determine its protein concentration and dilute the stock for the CD measurements. It is important to thoroughly dry the cuvette before using as water droplets in cuvette can alter the concentration of the sample. +
++
+(iV)Calibration of cell Pathlength
+The cells used for CD work should be manufactured to extremely high standards. Suprasil(quartz) is generally used for measurements. Refillable cells are available commercially with pathlengths in the range 0.01 to 10cm. Shorter pathlength calls (0.001cm or even shorter) are of demountable type consisting of 2 quartz plate and one of them is prepared to required depth.
+
+The path lengths can be established either on the basis of interference patterns caused by reflection of incident light from internal surfaces of the cells or on the absorbance of standard solutions.
+High transparency quartz cuvettes (cells) are used in CD spectrometer. Generally two types of cells are used, rectangular and
+cylindrical cells with path lengths ranging from 0.01 to 1 cm. Water jacketed
+cylindrical cells are available for CD machines that do not have temperature regulated cell holders. Rectangular cells with path lengths less than 0.1 cm often have a very small total sample volume and a very small surface area facing the light beam of the CD machine. It is important that the light beam should properly focus when these cells are used, since large artifacts are produced if
+the light does not go directly through the sample. In the near-UV region where path
+lengths from 1 mm down to 0.01 mm are required rectangular cells, in their standard, semimicro, or micro forms, are normally used. These are more economical than
+the cylindrical type, and allow mixing to be carried properly. For far-UV CD spectra, short-path-length cells from 1 mm down to 0.01 mm are
+necessary because of the higher absorbance and ellipticity in this region.
+
')
+
+ t1=content.index('')
+ print t1
+ f.seek(t1+13)
+ s1=s1+f.read()
+ #print s1
+ f.seek(t1+13)
+ f.write(s1)
+ f.seek(0)
+ content=f.read()
+ t= content.index('')
+ f.seek(t+13)
+ s='
' +while sectionNumber<=len(sectionno): + tag="" + att = ''+'experiment-article-section-'+str(sectionNumber)+'-heading' + tagger = soup.findAll('div', attrs={'id':att,'class':'heading'}) + if not tagger: + sectionNumber=sectionNumber+1 + continue + tag1=str(tagger[0].text) + tag1=tag1.strip() + print tag1 + if tag1=='Prerequisite S/W': + tag1='Prerequisites' + st+='
'+heading+'
'+s + s=s+f.read() + f.seek(t+13) + f.write(s) +f=open("template1.html",'r') +template=f.read() +f=open("content.html",'r') +srchtml=f.read() +f.close() +soup = BeautifulSoup(srchtml, 'html.parser') +sectionno=soup.find_all('section') +d=['introduction','theory','objective','experiment','manual','quizzes','further_readings','procedure','hh','ll'] +print len(sectionno) +sectionNumber=1 +st="" + +att = ''+'experiment-article-heading' +tagger = soup.findAll('header', attrs={'id':att,'class':'heading'}) +heading=str(tagger[0].text) +heading=heading.strip() +print heading +att = ''+'experiment-header-heading' +tagger = soup.findAll('div', attrs={'id':att,'class':'heading'}) +heading1=str(tagger[0].text) +print heading +heading1=heading1.strip() +breadcrumb='Chemical Sciences →'+heading1+' →List Of Experiments' +while sectionNumber<=len(sectionno): + tag="" + att = ''+'experiment-article-section-'+str(sectionNumber)+'-heading' + tagger = soup.findAll('div', attrs={'id':att,'class':'heading'}) + if not tagger: + sectionNumber=sectionNumber+1 + continue + tag1=str(tagger[0].text) + tag1=tag1.strip() + print tag1 + if tag1=='Prerequisite S/W': + tag1='Prerequisites' + st+='
'+tag1+'
' + sectionNumber=sectionNumber+1 +st+='Feedback
' +sectionNumber=1 +while sectionNumber<=len(sectionno): + tag="" + tag="" + att = ''+'experiment-article-section-'+str(sectionNumber)+'-heading' + tagger = soup.findAll('div', attrs={'id':att,'class':'heading'}) + if not tagger: + sectionNumber=sectionNumber+1 + continue + tag1=str(tagger[0].text) + tag1=tag1.strip() + if tag1=='Prerequisite S/W': + tag1='Prerequisites' + att = ''+'experiment-article-section-'+str(sectionNumber)+'-content' + tagger = soup.findAll('div', attrs={'id':att,'class':'content'}) + + tag+=str(tagger[0]) + #print tag + #print st + + + writefile(tag1+'.html',tag,st,heading) + #print sectionNumber + sectionNumber=sectionNumber+1 + +f=open("Feedback.html",'w+') +f.write(template) +f.seek(0) + +content = f.read() +content=content.replace('Disciplines and Domains',breadcrumb) +f.seek(0) +f.write(content) +#print content +f.seek(0) +content=f.read() +k=content.index('')
+
+t1=content.index('')
+print t1
+f.seek(t1+13)
+st=st+f.read()
+#print s1
+f.seek(t1+13)
+f.write(st)
+f.seek(0)
+content=f.read()
+t= content.index('')
+f.seek(t+13)
+s='Welcome to Virtual Labs - A MHRD Govt of india Initiative
+
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+Welcome to Virtual Labs - A MHRD Govt of india Initiative
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+Welcome to Virtual Labs - A MHRD Govt of india Initiative
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+Welcome to Virtual Labs - A MHRD Govt of india Initiative
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+Welcome to Virtual Labs - A MHRD Govt of india Initiative
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+Welcome to Virtual Labs - A MHRD Govt of india Initiative
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+Welcome to Virtual Labs - A MHRD Govt of india Initiative
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+Welcome to Virtual Labs - A MHRD Govt of india Initiative
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'+heading+'
'+'Feedback' +s=s+f.read() +f.seek(t+13) +f.write(s) diff --git a/src/lab/exp5/template1.html b/src/lab/exp5/template1.html new file mode 100644 index 0000000..382fb88 --- /dev/null +++ b/src/lab/exp5/template1.html @@ -0,0 +1,205 @@ + + + + + + + + + + +
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diff --git a/src/lab/exp6/Experiment.html b/src/lab/exp6/Experiment.html
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+ Disciplines and Domains
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diff --git a/src/lab/exp6/Feedback.html b/src/lab/exp6/Feedback.html
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+ Chemical Sciences →Circular Dichroism Virtual Lab →List Of Experiments
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+ Chemical Sciences →Circular Dichroism Virtual Lab →List Of Experiments
+
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+ To Study the effect of Secondary Structure elements on CD spectrum of proteins.
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diff --git a/src/lab/exp6/Introduction.html b/src/lab/exp6/Introduction.html
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+ Chemical Sciences →Circular Dichroism Virtual Lab →List Of Experiments
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+ Chemical Sciences →Circular Dichroism Virtual Lab →List Of Experiments
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+ To Study the effect of Secondary Structure elements on CD spectrum of proteins.
Feedback
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diff --git a/src/lab/exp6/Manual.html b/src/lab/exp6/Manual.html
new file mode 100644
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+ Chemical Sciences →Circular Dichroism Virtual Lab →List Of Experiments
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+ Chemical Sciences →Circular Dichroism Virtual Lab →List Of Experiments
+
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+ To Study the effect of Secondary Structure elements on CD spectrum of proteins.
+
+
+In the previous experiments, we have seen the effect of anisotropic medium on electromagnetic waves. +The rationale behind the emergence of the phenomenon of Circular birefringence and Circular Dichroism (CD) from such chiral +molecules has clearly been described in the earlier experiments. CD, being an intrinsic property of optically active molecules, +can therefore, be used as a method to identify and study the important optically active molecules of nature. One of the most +important optically active molecules is a protein molecule which is an essential biomolecule studied extensively across the +scientific community. Since, proteins are chiral in nature (due to their dextrorotary and levorotary components), they also +exhibit CD. As a consequence, CD can be used as a tool to study these molecules.
+
+In this experiment we will focus on the structure and function of protein molecules and their relationship with CD. As we proceed, we will be able to understand the different structural features of a protein and how do they affect the CD spectrum of proteins. We will see how CD spectroscopy can be used as a method for the characterization of different protein structures. Precisely, this experiment aims to observe and study characterization of protein structures and folds based on its CD spectrum. The capacity of CD to give a representative structural signature makes it a powerful tool in modern biochemistry with applications that can be found in virtually every field of study. +
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diff --git a/src/lab/exp6/Objective.html b/src/lab/exp6/Objective.html
new file mode 100644
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+ Chemical Sciences →Circular Dichroism Virtual Lab →List Of Experiments
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+ Chemical Sciences →Circular Dichroism Virtual Lab →List Of Experiments
+
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+ To Study the effect of Secondary Structure elements on CD spectrum of proteins.
+
+Step 1: The screen shows a CD spectrometer with a computer attached to it. There is a drop down menu named "Secondary Structure" and three control TABS on the right named "Measure CD", "Exit Simulation" and "N2 Purge".
+Step 2: You have to power on the CD spectrometer to activate the simulator. The power switch is at the left of the spectrometer.
+Step 3: Now you have to select one sample from the drop down menu named "Secondary Structure".
+Step 4: Now you may Purge the N2 and see what happens in the screen or directly "Measure CD".
+Step 5: If you select "Measure CD" then sample will be taken in special container. Now you have to open the spectrometer's lid . The sample will be automatically kept inside. Then you have close the lid. Opening and closing of the lid is done by clicking a white oval button below the lid.
+Step 6: You will now have the spectra of your sample. Click continue to "identify the secondary structural element" from the spectrum.
+Step 7: If it does not match with any existing spectra then you will be told that some thing has gone wrong. And you will be suggested to do the experiment again. While redoing the experiment you may try N2 purging to observe whether that makes any difference in your experimental result.
+Step 8: Once you receive a curve which matches with one of the spectrum of pure secondary structure of protein then you will be asked guess that structure.
+Step 9: Once you submit your result you will come to know whether your guess is right or wrong.
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diff --git a/src/lab/exp6/Quizzes.html b/src/lab/exp6/Quizzes.html
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+ Chemical Sciences →Circular Dichroism Virtual Lab →List Of Experiments
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+ Chemical Sciences →Circular Dichroism Virtual Lab →List Of Experiments
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+ To Study the effect of Secondary Structure elements on CD spectrum of proteins.
+
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+ +The purpose of the lab is to give students an idea of the structure of a protein and how CD spectroscopy technique is useful to detect the characteristics of secondary structures of proteins. By performing the experiment a student will
+
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-
+
- understand the level of complexity of protein structure. +
- observe CD spectrum of different secondary structures of the protein. +
- have an idea about the importance of the N2 purging during measuring the CD spectrometer. +
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diff --git a/src/lab/exp6/Theory.html b/src/lab/exp6/Theory.html
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+ Chemical Sciences →Circular Dichroism Virtual Lab →List Of Experiments
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+ Chemical Sciences →Circular Dichroism Virtual Lab →List Of Experiments
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+ To Study the effect of Secondary Structure elements on CD spectrum of proteins.
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+
+Here are two sets of interactive questions to help you to make your understanding more clear.
+After you go through the theory section you should try out the Pre-experiment quiz before you start the actual experiment.
+Once you perform the original experiment you should try out the Post-experiment quiz to have a better insight of the concepts.
+
+
Pre-experiment Quiz
+Click here to have a self evaluation.
+
+
Post-experiment Quiz
+Click here to have a self evaluation.
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diff --git a/src/lab/exp6/scraplab.py b/src/lab/exp6/scraplab.py
new file mode 100644
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+++ b/src/lab/exp6/scraplab.py
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+from bs4 import BeautifulSoup
+heading=""
+breadcrumb=""
+def writefile(fname,s,s1,heading):
+ f=open(fname, 'w+')
+ f.write(template)
+ f.seek(0)
+
+ content = f.read()
+ content=content.replace('Disciplines and Domains',breadcrumb)
+ f.seek(0)
+ f.write(content)
+ print content
+
+ f.seek(0)
+ content=f.read()
+ k=content.index('
+
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+ +
+ Chemical Sciences →Circular Dichroism Virtual Lab →List Of Experiments
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+ Chemical Sciences →Circular Dichroism Virtual Lab →List Of Experiments
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+ To Study the effect of Secondary Structure elements on CD spectrum of proteins.
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+Introduction to proteins: +
+
Biological Significance:+ + +
+ +Proteins are the most predominant organic molecules in all living cells with remarkable biological significance. +The structure, behaviour and unique qualities of all living beings are the consequence of the protein they contain. +Proteins perform the essential and specialized functions in living cells. They are primarily responsible for the +structure and strength of the body. According to their biological roles, proteins can be classified into various categories. +Some proteins act as enzymes, hormones, blood clotting factors, antibodies etc while some proteins are classified as +structural (e.g. collagen & keratin), contractile (e.g. actin & myosin), transport (e.g. haemoglobin & myoglobin) and +regulatory proteins (e.g. G-proteins). There are also nutrient and storage proteins which are vital for the growth and +survival of living cells.These are the dynamic functions of proteins. Due to their vital functions they are also known +as the working horses of cell. +
++
Structure of proteins:+
The building block of protein molecules is called as amino acid, which occur in 20 different naturally occurring forms. +A protein is a chain of amino acids linked together through peptide bonds. All proteins are made up of same set of these +20 amino acids arranged in a specific order. Various combinations of these amino acids account for a variety of proteins +occurring naturally. A typical amino acid consist of a primary amino group (-NH2), a carboxyl group (-COOH), a hydrogen atom +(-H) and a side-chain (-R) group attached to a central α-carbon atom (-Cα). The structure is as shown below. +
+|
+
+ |
+
+Structure of an amino acid
(source:http://www.bothbrainsandbeauty.com/page/3)
+
+The variation among the amino acids occurs at the R group, which is different in each amino acid displaying different +physiochemical properties (e.g. polarity, acidity, basicity, aromaticity, ability to from hydrogen bonds etc). +Only Proline differs from this basic structure and has its aliphatic side-chain bonded back onto the amino group +and is thus, instead an imino acid. This makes Proline a conformationally rigid molecule. +
++
+ +All of the 20 standard amino acids, except for glycine, have four different groups around the Cα +atom and thus are optically active and chiral in nature. These can be distinguished based on the difference +in rotation of the place polarised light. Only one of the forms of the isomer (L-isomer) +(see http://en.wikipedia.org/wiki/Chirality_(chemistry) for more details) is found in proteins. +
++
The peptide bond:
+As discussed above, a protein is a linear sequence of amino acids linked together by peptide bonds. +The peptide bond is a covalent bond between the α-amino group of one amino acid and the α-carboxyl +group of another. The formation of peptide bond involves removal of water molecule and hence, amino acids +are also referred to as residues. The figure below illustrates a peptide bond formation between two amino +acid residues with the removal of water molecule. +
++
|
+
+ |
+
Peptide bond formation
(source: http://www.bothbrainsandbeauty.com/page/3)
+
+
+The peptide bond has partial double bond character. When two amino acids are joined by a peptide bond,
+they form a dipeptide.A tripeptide refers to a chain of three amino acids while tetrapeptide is a sequence
+of four amino acid residues and so on. Addition of further amino acids results in long chains called as oligopeptides
+(upto 25 amino acid residues) and polypeptides (>25 amino acid residues). The backbone conformation of a polypeptide
+is specified by the rotational angles about the Cα - N bond(φ) and Cα - C (ψ) of each amino acid
+residues as shown below:
+
+
+ |
+
Torsional angles
(Source:http://biochem.co/page/3/)
+
+
+
+Since amino acids are chiral in nature, they can generate CD signals. In CD spectroscopy, +in the far-UV spectral region of wavelengths ranging from 190-250 nm, the chromophore is +the peptide bond while in the near-UV spectral region of wavelengths ranging from 250-350 nm, +the chromophores are the aromatic amino acids and disulfide bonds. Signals in the region from +250-270 nm are attributable to phenylalanine residues, signals from 270-290 nm are attributable +to tyrosine, and those from 280-300 nm are attributable to tryptophan. Disulfide bonds give rise +to broad weak signals throughout the near-UV spectrum. The signal strength in the near-UV CD region +is much weaker than that in the far-UV CD region. Near-UV CD spectra require about 1 ml of protein +solution with an OD at 280 nm of 0.5 to 1 (which corresponds to 0.25 to 2 mg/ml for most proteins). +
++
Primary Structure of proteins+
The primary level of structure in a protein is the linear sequence of amino acids as joined together by +peptide bonds. It refers to the number and order of the amino acids present in the protein. The convention +for the designation of the order of amino acids is that the N-terminal end (i.e. the end bearing the residue +with the free α-amino group) is to the left (and the number 1 amino acid) and the C-terminal end +(i.e. the end with the residue containing a free α-carboxyl group) is to the right. +
++
Secondary structure of proteins:+
The secondary level of structure in a protein refers to the folding of the local regions of the polypeptide chain. +Secondary structure arises when various functional groups of the constituting amino acids interact and arrange +themselves spatially. This causes the amino acid chain to twist and attain different configurations. +The most common types of secondary structure elements are the α-helices, β-pleated sheets and random coils. +
++
+The α-helix is a common secondary structure encountered in many proteins (e.g. haemoglobin). The formation of the α +-helix is spontaneous and is stabilized by H-bonding between amide hydrogen and carbonyl oxygen of peptide bonds spaced four +residues apart. This orientation of H-bonding produces a helical coiling of the peptide backbone such that the R-groups +lie on the exterior of the helix and perpendicular to its axis. In an α-helix, as shown below, there are 3.6 amino +acids per turn of the helix covering a distance of 5.4 Ao, and each amino acid residue represents an advance of 1.5 Ao along the axis of the helix. +
++
+
+ |
+
An Alpha-helix
(source: http://student.ccbcmd.edu/~gkaiser/biotutorials/proteins/fg4a.html)
+
+
+
+Not all amino acids favour the formation of the α-helix due to steric constraints of the R-groups. +Amino acids such as Alanine, Aspartic acid, Glutamic acid, Isoleucine, Leucine and Methionine favour +the formation of α-helices, whereas, Glycine and Proline disrupt the helix. This is particularly true for +Proline since it cannot form the correct pattern of hydrogen bonds due to lack of a hydrogen atom on its +nitrogen atom. For this reason, Proline is often found at the end of the α-helix, where it alters the direction +of the polypeptide chain and terminates the helix. + +
++
+ +The second common secondary structure in proteins is the β-pleated sheets. The β-sheets are +composed of two different regions of stretches of at least 5 - 10 amino acids called as +β-strands. They could be different polypeptide chains or in different sections of the +same polypeptide chain. The planarity of the peptide bond forces the polypeptide to be pleated, +with the side chains protruding above and below the sheet. The folding and alignment of +these strands aside one another to form β-sheets is stabilized by H-bonding between amide hydrogen and +carbonyl oxygen. Adjacent polypeptide chains in the β-sheets are either parallel or anti-parallel +depending on whether they run in the same direction or in opposite directions. In parallel sheets, +adjacent peptide chains proceed in the same direction (i.e. the direction of N-terminal to C-terminal ends is the same) +, whereas, in anti-parallel sheets adjacent chains are aligned in opposite directions. β-sheets can be depicted as shown below. +
++
+
+ |
+
+
+
+
+ |
Beta-sheet
(source:
+http://www.chembio.uoguelph.ca/educmat/phy456/456lec01.htm
+and http://www.cryst.bbk.ac.uk/pps97/course/section3/sheet.html)
+
+
+β-pleated sheets provide strength and rigidity in many structural proteins such as silk fibroin. +The ribbon depiction of β-pleated sheets is shown in the figure below. +
++
+
+ |
Ribbon depiction of β-pleated sheets.
+(source:http://themedicalbiochemistrypage.org/protein-structure.html)
+
+Turns are the third secondary structures that serve to reverse the direction of the polypeptide chain in order to +fold tightly into the compact shape. These turns are often found connecting the ends of anti-parallel β-pleated sheets. +They are located primarily on the protein surface and accordingly contain polar and charged residues. Antibody recognition, phosphorylation, glycosylation, and hydroxylation sites are found frequently at or adjacent to turns. +Regions of the polypeptide chain that are not in a regular secondary structure are said to have a coil or loop conformation. +
++We will now see how the presence of each of these structural elements in the protein can affect the CD spectrum of protein. We will also +observe the CD signal generated which is a representative of these structural elements that can be used to characterise a given protein. +
++
CD and Proteins:
++ +Having discussed the basic Physics behind CD spectroscopy in experiment 3 +we are now in a position to understand how naturally occurring molecules like protein shows Circular Dichroism.
+ ++ +We now know that in an optically active medium, chromophores cause differential absorption and in turn give rise to Circular +Dichroism. In case of proteins, the α-carbon acts as a chiral center and hence provides an optically active medium. Proteins +possess a number of chromophores (peptide bond, aromatic side chain, disulphide bond etc) which can give rise to CD signals. +In the far UV region (180-250nm), which corresponds to peptide bond absorption, the CD spectrum can be analysed to give the +content of regular secondary structural features such as α-helix and β-sheet. The CD spectrum in the near UV region (250-320 nm) +reflects the environments of the aromatic amino acid side chains and thus gives information about the tertiary structure of the protein. +
++To study secondary structures, amide bonds (peptide bonds) are the main source of CD spectra. Molecular Orbital +Theory(http://en.wikipedia.org/wiki/Molecular_orbital_theory) +tells us that each molecule has a set of molecular orbitals and electrons of its atoms are distributed among those orbitals. +For amide bond, in simplified Molecular Orbital Theory, the 2Px orbitals of the nitrogen, carbon and oxygen atoms of the amide +linkage are combined to form three orthogonal linear combinations, the π+, πo, and π- bonds. These three orbitals have the plane +of the peptide bond as there nodal plane. The four π electrons fill the first two molecular orbitals, the π+ which is strongly bonding and +πo which is almost nonbonding. The remaining two electrons are non-bonding electrons on the oxygen atoms of the amide bond which +are conventionally labeled the 2Py electrons. These electrons have energy which is close to that of the atomic orbitals of hydrogen. +
++
+Now under excitation there may be two types of electronic transition
+1. n - π* centered around 220 nm and
+2. π - π* centered around 190 nm
+
+
+
+ |
+*Peptide bond
+
+
+1. The n-π* transition involves the promotion of an electron from the n to the π- orbital.
+The transition is very weak because the ground and excited states have nodal planes which are perpendicular to one another.
+Thus the n-π* transition is said to be forbidden. The transition is forbidden electronically but permitted magnetically.
+ Thus the absorption band of the n-π* transition is very weak, but the CD bands can be very large.
+
+2. The π - π* transition involves the promotion of an electron from the πo to the π orbital and is strong with an
+ extinction coefficient equal to approximately 7100.
+
+
+ +In a system of N non interacting chromophores if one chromophore is excited by the incident light, the energies of the other +N non interacting molecules are unchanged. Thus, an array of N chromophores would have N-fold degeneracy, as any chromophores +could be equally excited by the light. But a folded protein the amide is in a continuous array. Therefore, their chromophores +are in ordered arrays. So, when electrons are excited they do interact. Now different secondary structures of a protein correspond +to such different types of arrays. And thus different secondary structures of protein give different types of CD spectra. +
++ +For example, we may have a series of amino acid Alanine (part of a protein) arranged in different forms. +They may appear as α-helix, β sheet or random coils. And for each of these secondary structures +we have different CD curves. The following figure shows that. +
++
+
+ |
*CD of Poly L Lysine for different secondary structure.
+
+
+
Let us now look at CD of different secondary structures+ +
+
+
+
+ |
*CD (blue) and absorption spectra of Random Coil +Electronic Transitions: p -> p*(positive at 212 nm) ; n -> p*(negative at 195 nm) +
++
+
+
+
+ |
*CD (blue) and absorption spectra of β Sheet +Electronic Transitions: n -> p*(positive at 195 nm) ; p -> p*(negative at 218 nm) + +
++
+
+
+
+ |
+
*CD (blue) and absorption (dotted) spectra of α Helix +Electronic Transitions: p -> p*(positive perpendicular at 192 nm); p -> p*(negative parallel at 209 +nm); n -> p*(negative at 222 nm is red shifted) +
++
+
+*Source:http://mach7.bluehill.com/proteinc/cd/cdspec.html +
++
Goal of The Experiment
++
+In this experiment we are going to measure CD spectrum of different unknown protein samples. These samples are pure secondary structures. User will have an experience to predict the nature of secondary structure from the shape of the CD spectrum. +
++
')
+
+ t1=content.index('')
+ print t1
+ f.seek(t1+13)
+ s1=s1+f.read()
+ #print s1
+ f.seek(t1+13)
+ f.write(s1)
+ f.seek(0)
+ content=f.read()
+ t= content.index('')
+ f.seek(t+13)
+ s='
' +while sectionNumber<=len(sectionno): + tag="" + att = ''+'experiment-article-section-'+str(sectionNumber)+'-heading' + tagger = soup.findAll('div', attrs={'id':att,'class':'heading'}) + if not tagger: + sectionNumber=sectionNumber+1 + continue + tag1=str(tagger[0].text) + tag1=tag1.strip() + print tag1 + if tag1=='Prerequisite S/W': + tag1='Prerequisites' + st+='
'+heading+'
'+s + s=s+f.read() + f.seek(t+13) + f.write(s) +f=open("template1.html",'r') +template=f.read() +f=open("content.html",'r') +srchtml=f.read() +f.close() +soup = BeautifulSoup(srchtml, 'html.parser') +sectionno=soup.find_all('section') +d=['introduction','theory','objective','experiment','manual','quizzes','further_readings','procedure','hh','ll'] +print len(sectionno) +sectionNumber=1 +st="" + +att = ''+'experiment-article-heading' +tagger = soup.findAll('header', attrs={'id':att,'class':'heading'}) +heading=str(tagger[0].text) +heading=heading.strip() +print heading +att = ''+'experiment-header-heading' +tagger = soup.findAll('div', attrs={'id':att,'class':'heading'}) +heading1=str(tagger[0].text) +print heading +heading1=heading1.strip() +breadcrumb='Chemical Sciences →'+heading1+' →List Of Experiments' +while sectionNumber<=len(sectionno): + tag="" + att = ''+'experiment-article-section-'+str(sectionNumber)+'-heading' + tagger = soup.findAll('div', attrs={'id':att,'class':'heading'}) + if not tagger: + sectionNumber=sectionNumber+1 + continue + tag1=str(tagger[0].text) + tag1=tag1.strip() + print tag1 + if tag1=='Prerequisite S/W': + tag1='Prerequisites' + st+='
'+tag1+'
' + sectionNumber=sectionNumber+1 +st+='Feedback
' +sectionNumber=1 +while sectionNumber<=len(sectionno): + tag="" + tag="" + att = ''+'experiment-article-section-'+str(sectionNumber)+'-heading' + tagger = soup.findAll('div', attrs={'id':att,'class':'heading'}) + if not tagger: + sectionNumber=sectionNumber+1 + continue + tag1=str(tagger[0].text) + tag1=tag1.strip() + if tag1=='Prerequisite S/W': + tag1='Prerequisites' + att = ''+'experiment-article-section-'+str(sectionNumber)+'-content' + tagger = soup.findAll('div', attrs={'id':att,'class':'content'}) + + tag+=str(tagger[0]) + #print tag + #print st + + + writefile(tag1+'.html',tag,st,heading) + #print sectionNumber + sectionNumber=sectionNumber+1 + +f=open("Feedback.html",'w+') +f.write(template) +f.seek(0) + +content = f.read() +content=content.replace('Disciplines and Domains',breadcrumb) +f.seek(0) +f.write(content) +#print content +f.seek(0) +content=f.read() +k=content.index('')
+
+t1=content.index('')
+print t1
+f.seek(t1+13)
+st=st+f.read()
+#print s1
+f.seek(t1+13)
+f.write(st)
+f.seek(0)
+content=f.read()
+t= content.index('')
+f.seek(t+13)
+s='Welcome to Virtual Labs - A MHRD Govt of india Initiative
+
+
+
+
+
+
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+Welcome to Virtual Labs - A MHRD Govt of india Initiative
+
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+Welcome to Virtual Labs - A MHRD Govt of india Initiative
+
+
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+Welcome to Virtual Labs - A MHRD Govt of india Initiative
+
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+Welcome to Virtual Labs - A MHRD Govt of india Initiative
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+Welcome to Virtual Labs - A MHRD Govt of india Initiative
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+Welcome to Virtual Labs - A MHRD Govt of india Initiative
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+Welcome to Virtual Labs - A MHRD Govt of india Initiative
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'+heading+'
'+'Feedback' +s=s+f.read() +f.seek(t+13) +f.write(s) diff --git a/src/lab/exp6/template1.html b/src/lab/exp6/template1.html new file mode 100644 index 0000000..382fb88 --- /dev/null +++ b/src/lab/exp6/template1.html @@ -0,0 +1,205 @@ + + + + + + + + + + +
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diff --git a/src/lab/exp7/Experiment.html b/src/lab/exp7/Experiment.html
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+ Disciplines and Domains
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diff --git a/src/lab/exp7/Feedback.html b/src/lab/exp7/Feedback.html
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+ +
+ Chemical Sciences →Circular Dichroism Virtual Lab →List Of Experiments
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+ Chemical Sciences →Circular Dichroism Virtual Lab →List Of Experiments
+
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+ To deconvolute the CD spectrum of a given protein solution and to classify it in terms of its secondary structure elements.
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diff --git a/src/lab/exp7/Introduction.html b/src/lab/exp7/Introduction.html
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+ Chemical Sciences →Circular Dichroism Virtual Lab →List Of Experiments
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+ Chemical Sciences →Circular Dichroism Virtual Lab →List Of Experiments
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+ To deconvolute the CD spectrum of a given protein solution and to classify it in terms of its secondary structure elements.
Feedback
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diff --git a/src/lab/exp7/Manual.html b/src/lab/exp7/Manual.html
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+ Chemical Sciences →Circular Dichroism Virtual Lab →List Of Experiments
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+ Chemical Sciences →Circular Dichroism Virtual Lab →List Of Experiments
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+ To deconvolute the CD spectrum of a given protein solution and to classify it in terms of its secondary structure elements.
+
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+In Experiment 6 we have seen CD spectrum of pure secondary structures of a protein eg alpha helix,beta sheet, coil etc. +We have seen how these spectra have different characteristics. Following diagram shows the nature of such spectra. +
++
|
+
+source: http://www.photophysics.com/secondarystructure.php
+ |
+
+
+But in practice, proteins are a combination of different types of secondary structures. So CD spectrum of a protein sample will show a combined effect of different types of secondary structures present in it. Now it is important to extract information about presence of different secondary structures in the sample. For that we follow a method called deconvolution. By deconvoluting + CD spectra of a protein we can come to know which kind of secondary structure is present in what amount in the protein. +
++
+Now in the case of protein structure is conserved more than sequence. Moreover structure is the basis of functions of a protein. + So there is a need to order the vast amounts of structural data of protein available. And our common philosophy leads us to study groups rather than individuals. Hence, on the basis of the amount and the type of secondary structures present in a protein, it is classified into different classes. +
++
+In this experiment we will see the method of deconvolution and learn the classification rule of protein on the basis of secondary structure. +
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diff --git a/src/lab/exp7/Objective.html b/src/lab/exp7/Objective.html
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+ Chemical Sciences →Circular Dichroism Virtual Lab →List Of Experiments
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+ Chemical Sciences →Circular Dichroism Virtual Lab →List Of Experiments
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+ To deconvolute the CD spectrum of a given protein solution and to classify it in terms of its secondary structure elements.
+
+Step 1: Select any one sample from the list of four samples and click "continue"
+Step 2: The next step shows variation of ellipticity (in milli-degrees) with wavelength (in nm) of both the sample spectrum and base line. It also shows the sample and experimental environment details. Click "Substract Baseline From Sample" to go to the next step.
+Step 3: In this step a graph is shown where baseline is substracted from the sample spectrum. You can go "back" to review the previous step or "continue" to the next step.
+Step 4: In this step you are given 3 options. Among them you have to select one method of Deconvolution and "Run" the experiment.
+Step 5: This step shows the result of deconvolution. From the information given in the result window you have to decide in which class does the protein sample belong. If you are not sure you can "Repeat Deconvolution" with other methods. And then submit your “answer”.
+Step 6: This step will show whether your result is right or wrong.
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diff --git a/src/lab/exp7/Quizzes.html b/src/lab/exp7/Quizzes.html
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+ Chemical Sciences →Circular Dichroism Virtual Lab →List Of Experiments
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+ Chemical Sciences →Circular Dichroism Virtual Lab →List Of Experiments
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+ To deconvolute the CD spectrum of a given protein solution and to classify it in terms of its secondary structure elements.
+
+Purpose of this lab is to give students an idea of how CD spectrum of a protein sample is deconvoluted and how
+information about secondary structure of the protein sample can be obtained from it. By doing this lab an user will be able to understand,
+
+
-
+
- +Different techniques of deconvolution of CD spectrum of a protein sample. + +
- +How to get information about secondary structures of a protein from its CD spectra. + +
- +How to classify a protein sample in terms of its secondary structure elements. + +
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diff --git a/src/lab/exp7/Theory.html b/src/lab/exp7/Theory.html
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+ To deconvolute the CD spectrum of a given protein solution and to classify it in terms of its secondary structure elements.
+
+
+Here are two sets of interactive questions to help you to make your understanding more clear.
+After you go through the theory section you should try out the Pre-experiment quiz before you start the actual experiment.
+Once you perform the original experiment you should try out the Post-experiment quiz to have a better insight of the concepts.
+
+
Pre-experiment Quiz
+Click here to have a self evaluation.
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+
Post-experiment Quiz
+Click here to have a self evaluation.
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diff --git a/src/lab/exp7/scraplab.py b/src/lab/exp7/scraplab.py
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+from bs4 import BeautifulSoup
+heading=""
+breadcrumb=""
+def writefile(fname,s,s1,heading):
+ f=open(fname, 'w+')
+ f.write(template)
+ f.seek(0)
+
+ content = f.read()
+ content=content.replace('Disciplines and Domains',breadcrumb)
+ f.seek(0)
+ f.write(content)
+ print content
+
+ f.seek(0)
+ content=f.read()
+ k=content.index('
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+ Chemical Sciences →Circular Dichroism Virtual Lab →List Of Experiments
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+ Chemical Sciences →Circular Dichroism Virtual Lab →List Of Experiments
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+ To deconvolute the CD spectrum of a given protein solution and to classify it in terms of its secondary structure elements.
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Deconvolution
++
+Let us now consider the CD spectrum as a mathematical functional or signal. We will first see what is called convolution of such signals. +
++Convolution is a mathematical operation on two functions, producing a third function that is typically viewed as a modified version of one of the original functions. In this context these initial functions are CD spectra of known pure secondary structures which convolute to give rise to a complex spectrum of a real protein. +
++Deconvolution is an algorithm-based process used to reverse the effects of convolution on recorded data. Now to deconvolute a CD spectrum of a protein sample we have to: +
-
+
- +Obtain a proper set of basis. + +
- +Perform the data fitting procedure. + +
+
+A basis set is a set of components chosen in such a way that your experimental CD spectrum can be view as a linear combination of them. If these basis were obtained from a set of reference protein whose structure and CD is known, we can obtain structural information on our protein as well. +
++
+Different Processes of Deconvolution of CD spectrum +
++All methods of analyzing CD spectra assume that the spectrum of a protein can be represented +by a linear combination of the spectra of its secondary structural elements, plus a noise term, +which includes the contribution of aromatic chromophores and prosthetic groups. +
++
+
θλ=Σ εi Sλi + noise
+
++
+where θλ is the CD of the protein as a function of wavelength, εi is the fraction of each secondary +structure, i, and Sλi is the ellipticity at each wavelength of each ith secondary structural element. + In constrained fits, the sum of all the fractional weights, εi, +must equal one and all of the fractional contributions must be greater than or equal to zero. +
++
+There are two general classes of methods to evaluate protein conformation.
+
-
+
- + The first uses standards of polypeptides, with defined compositions in known conformations, which have been determined by X-ray scattering of films or by IR in solution. + +
- + The second uses the spectra of proteins which have been characterized by X-ray crystallography as standards. These are then compared to the spectra of unknown proteins using least squares analysis, ridge regression, singular value decomposition, single value decomposition with variable selection; the self consistent method; or neural network analysis. + +
+
+The protein-based analyses are superior when analyzing the conformation of globular, well-folded proteins. + So we will discuss three different methods of second type. +
++
+1. Ridge Regression based method CONTIN: +
+Principle:CONTIN fits the CD of unknown proteins by comparison to a linear combination of the spectra of a large database of proteins with known conformations. In this method the contribution of each reference spectrum is kept small unless it contributes to a good agreement between the theoretical best fit curve and the raw data. +
++
+Advantage: This method results in relatively good estimates of α-helices and β-sheets. Different references are used for every fit, which is an advantage for obtaining the best fits of the data. +
++
+Disadvantage: Complicates the quantitative analysis of the effect of a mutations or denaturant, since a different set of standards is used for each analysis. +
++
+(2) & (3) Neural Network based approach CDNN & K2D: +
++Process: A neural network is an artificial intelligence program used to find correlations in data. A neural network is first trained using a set of known proteins so that the input of the CD at each wavelength results in the output of the correct secondary structure. The trained network is then used to analyze unknown proteins. Two widely used programs are CDNN and K2D. +
++
+Advantage: CDNN analyzes data to determine helix, anti and parallel β- structure, turns and remainder and K2D determines helix, total β-structure and remainder. K2D gives a good estimate of the helical and sheet contents of both proteins and polypeptides. +
++
+Disadvantage: However, the K2D program does not estimate turns. CDNN is not suitable for the analysis of polypeptides and it currently is not being distributed. +
++
+Classification of Globular Proteins According to Secondary Structure: +
++As we have discussed in the introduction section, structure is more conservative than sequence and structure of a protein directly controls its functions. So it is important to classify proteins depending on its secondary structure and study proteins as a group having common structural properties. +
+ ++
+Here are four types of classes of proteins classified on the basis of their secondary structure. +
++
+All alpha: Proteins that contain only (in some exceptional instances there may be isolated beta-sheets) +alpha helical secondary structure. Myoglobin is an example of an all alpha protein. +
+
+ |
+
Source:http://en.wikipedia.org/wiki/File:Myoglobin.png +
++
+All beta: Protein that contain only (in some exceptional instances there may be isolated alpha helices) beta-sheet secondary structure. Tenascin is an example of an all beta protein. +
+
+ |
+
Source:http://en.wikipedia.org/wiki/File:1ten_fibronectin_type_III.png +
++
+Alpha / beta: Proteins that contain alternating alpha-helical and beta-sheet secondary structure elements. Triose Phosphate Isomerase is an example of an alpha/beta protein. This structure is known as a TIM barrel. The alternating helical and sheet segments form a closed barrel like structure. +
+
+ |
+
Source:http://en.wikipedia.org/wiki/File:Beta-barrel.png +
++
+Alpha + Beta: In these proteins the alpha helical and beta sheet regions occur in independent regions of the molecule. Ribonuclease A is an example of an alpha+beta protein. +
+
+ |
+
Source:http://en.wikipedia.org/wiki/File:RNase_A.png +
++
+Goal of The Experiment:
+In this experiment we will observe results of different deconvolution methods. Compare them and will try to classify proteins from those results.
+
+
+
')
+
+ t1=content.index('')
+ print t1
+ f.seek(t1+13)
+ s1=s1+f.read()
+ #print s1
+ f.seek(t1+13)
+ f.write(s1)
+ f.seek(0)
+ content=f.read()
+ t= content.index('')
+ f.seek(t+13)
+ s='
' +while sectionNumber<=len(sectionno): + tag="" + att = ''+'experiment-article-section-'+str(sectionNumber)+'-heading' + tagger = soup.findAll('div', attrs={'id':att,'class':'heading'}) + if not tagger: + sectionNumber=sectionNumber+1 + continue + tag1=str(tagger[0].text) + tag1=tag1.strip() + print tag1 + if tag1=='Prerequisite S/W': + tag1='Prerequisites' + st+='
'+heading+'
'+s + s=s+f.read() + f.seek(t+13) + f.write(s) +f=open("template1.html",'r') +template=f.read() +f=open("content.html",'r') +srchtml=f.read() +f.close() +soup = BeautifulSoup(srchtml, 'html.parser') +sectionno=soup.find_all('section') +d=['introduction','theory','objective','experiment','manual','quizzes','further_readings','procedure','hh','ll'] +print len(sectionno) +sectionNumber=1 +st="" + +att = ''+'experiment-article-heading' +tagger = soup.findAll('header', attrs={'id':att,'class':'heading'}) +heading=str(tagger[0].text) +heading=heading.strip() +print heading +att = ''+'experiment-header-heading' +tagger = soup.findAll('div', attrs={'id':att,'class':'heading'}) +heading1=str(tagger[0].text) +print heading +heading1=heading1.strip() +breadcrumb='Chemical Sciences →'+heading1+' →List Of Experiments' +while sectionNumber<=len(sectionno): + tag="" + att = ''+'experiment-article-section-'+str(sectionNumber)+'-heading' + tagger = soup.findAll('div', attrs={'id':att,'class':'heading'}) + if not tagger: + sectionNumber=sectionNumber+1 + continue + tag1=str(tagger[0].text) + tag1=tag1.strip() + print tag1 + if tag1=='Prerequisite S/W': + tag1='Prerequisites' + st+='
'+tag1+'
' + sectionNumber=sectionNumber+1 +st+='Feedback
' +sectionNumber=1 +while sectionNumber<=len(sectionno): + tag="" + tag="" + att = ''+'experiment-article-section-'+str(sectionNumber)+'-heading' + tagger = soup.findAll('div', attrs={'id':att,'class':'heading'}) + if not tagger: + sectionNumber=sectionNumber+1 + continue + tag1=str(tagger[0].text) + tag1=tag1.strip() + if tag1=='Prerequisite S/W': + tag1='Prerequisites' + att = ''+'experiment-article-section-'+str(sectionNumber)+'-content' + tagger = soup.findAll('div', attrs={'id':att,'class':'content'}) + + tag+=str(tagger[0]) + #print tag + #print st + + + writefile(tag1+'.html',tag,st,heading) + #print sectionNumber + sectionNumber=sectionNumber+1 + +f=open("Feedback.html",'w+') +f.write(template) +f.seek(0) + +content = f.read() +content=content.replace('Disciplines and Domains',breadcrumb) +f.seek(0) +f.write(content) +#print content +f.seek(0) +content=f.read() +k=content.index('')
+
+t1=content.index('')
+print t1
+f.seek(t1+13)
+st=st+f.read()
+#print s1
+f.seek(t1+13)
+f.write(st)
+f.seek(0)
+content=f.read()
+t= content.index('')
+f.seek(t+13)
+s='Welcome to Virtual Labs - A MHRD Govt of india Initiative
+
+
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+Welcome to Virtual Labs - A MHRD Govt of india Initiative
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+Welcome to Virtual Labs - A MHRD Govt of india Initiative
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+Welcome to Virtual Labs - A MHRD Govt of india Initiative
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+Welcome to Virtual Labs - A MHRD Govt of india Initiative
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+Welcome to Virtual Labs - A MHRD Govt of india Initiative
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+Welcome to Virtual Labs - A MHRD Govt of india Initiative
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+Welcome to Virtual Labs - A MHRD Govt of india Initiative
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'+heading+'
'+'Feedback' +s=s+f.read() +f.seek(t+13) +f.write(s) diff --git a/src/lab/exp7/template1.html b/src/lab/exp7/template1.html new file mode 100644 index 0000000..382fb88 --- /dev/null +++ b/src/lab/exp7/template1.html @@ -0,0 +1,205 @@ + + + + + + + + + + +
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diff --git a/src/lab/exp8/Experiment.html b/src/lab/exp8/Experiment.html
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+ Disciplines and Domains
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diff --git a/src/lab/exp8/Feedback.html b/src/lab/exp8/Feedback.html
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+ Chemical Sciences →Circular Dichroism Virtual Lab →List Of Experiments
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+ Chemical Sciences →Circular Dichroism Virtual Lab →List Of Experiments
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+ To Study the Thermal Stability of Proteins.
+
+Click here to start the experiment. + + +
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diff --git a/src/lab/exp8/Introduction.html b/src/lab/exp8/Introduction.html
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+ Chemical Sciences →Circular Dichroism Virtual Lab →List Of Experiments
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+ Chemical Sciences →Circular Dichroism Virtual Lab →List Of Experiments
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+ To Study the Thermal Stability of Proteins.
Feedback
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diff --git a/src/lab/exp8/Manual.html b/src/lab/exp8/Manual.html
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+ Chemical Sciences →Circular Dichroism Virtual Lab →List Of Experiments
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+ Chemical Sciences →Circular Dichroism Virtual Lab →List Of Experiments
+
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+ To Study the Thermal Stability of Proteins.
+
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+We all are familiar with the phenomena of boiling an egg. At higher temperature the egg white gets solidified. This phenomenon may be explained by the concept of thermal denaturation. The white portion of an egg is nothing but a protein called albumin. At higher temperatures the protein albumin unfolds and as a result the state of the protein changes from liquid to solid. +
++Another interesting observation is that this process of unfolding is irreversible. So if we decrease the temperature of a boiled egg, the albumin will not refold and form a liquid state again. This phenomenon is explained in the following video. +
+
+
+ +
+To understand the extent of unfolding in proteins at different temperatures it is important to estimate its secondary structure content at that particular temperature. In Experiment 6 and 7 we have seen how to extract information regarding secondary structure of a protein from circular dichroism spectroscopic data. So CD spectrometry may be used as a very effective and efficient tool for studying the secondary structure of a protein at different temperatures and inturn aids in studying thermal stability of the protein. +
+
In this experiment we will observe the phenomena of unfolding of a helical membrane protein, called Rhodopsin. We will see how Rhodopsin goes to an unfolded state from a folded state with increase in the temperature. At the same time we will understand how we can use CD spectrometry data to study the thermal stability of Rhodopsin and find out the transition temperature of unfolding.
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diff --git a/src/lab/exp8/Objective.html b/src/lab/exp8/Objective.html
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+ Chemical Sciences →Circular Dichroism Virtual Lab →List Of Experiments
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+ Chemical Sciences →Circular Dichroism Virtual Lab →List Of Experiments
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+ To Study the Thermal Stability of Proteins.
+
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+
1. Video Tutorial
+ + + ++
click here to ENLARGE.
++
2. Step-by-step Procedure
++The experiment has 3 steps â STEP1, STEP2 and STEP3. +
++
Click on START to proceed step by step. Or Click STEP1 or STEP2 or STEP3 to jump to mentioned steps.
++Click to START. Observe the Animation. Click BACK to return to the previous stem or click CONTINUE to go to the next step. +
+
+Click on the Protein Structure shown and study the structure. If you are done click to BACK to go back to the previous step or Click START to go to the next step. +
+
+Next step shows the experiment set up. Follow the instructions appearing on the TOP left corner of the page. Once you are done click on the monitor to go to the next step. You may click on RESET at any point of time to reinitialize the experiment. +
+
+Select the initial temperature, final temperature, and step size. Then click on Measure CD Spectra. +
+
+Observe the spectra of the starting temperature. Use CONTROLS to see spectra at different temperatures. Take your mouse on the graph. Select the MRE value corresponding to 222nm and fill up the TABLE. Click on Plot once you have finished filling up the TABLE. You can come back to this step by clicking Go Back. +
+
+Once you are done click Go To Analysis. +
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diff --git a/src/lab/exp8/Quizzes.html b/src/lab/exp8/Quizzes.html
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+ Chemical Sciences →Circular Dichroism Virtual Lab →List Of Experiments
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+ Chemical Sciences →Circular Dichroism Virtual Lab →List Of Experiments
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+
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+
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+ To Study the Thermal Stability of Proteins.
+
+
The purpose of this lab is to make students familiar with the concept that structure of a protein is highly dependent on the environmental temperature and use of CD spectroscopy method to study the effect of temperature on protein structure. By doing this experiment user will be able to,
++
-
+
- Make the student understand how temperature causes unfolding of a protein. +
- Understand how CD spectroscopy can be used to study unfolding. +
- Observe transition between folded and unfolded state. +
- Determine the transition temperature using CD spectra at different temperatures. +
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diff --git a/src/lab/exp8/Theory.html b/src/lab/exp8/Theory.html
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+ Chemical Sciences →Circular Dichroism Virtual Lab →List Of Experiments
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+ To Study the Thermal Stability of Proteins.
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diff --git a/src/lab/exp8/scraplab.py b/src/lab/exp8/scraplab.py
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+from bs4 import BeautifulSoup
+heading=""
+breadcrumb=""
+def writefile(fname,s,s1,heading):
+ f=open(fname, 'w+')
+ f.write(template)
+ f.seek(0)
+
+ content = f.read()
+ content=content.replace('Disciplines and Domains',breadcrumb)
+ f.seek(0)
+ f.write(content)
+ print content
+
+ f.seek(0)
+ content=f.read()
+ k=content.index('
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+ Chemical Sciences →Circular Dichroism Virtual Lab →List Of Experiments
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+ Chemical Sciences →Circular Dichroism Virtual Lab →List Of Experiments
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+ To Study the Thermal Stability of Proteins.
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Protein Folding - the process and its cause
+
Protein folding is the ultimate process where the information contained in genes is transformed into the final functional unit, an active folded protein from the unfolded primary amino acid sequence.
+The amino-acid sequence (or primary structure) of a protein determines its native conformation. A protein molecule folds spontaneously during or after biosynthesis. +For the most part, scientists have been able to study many identical molecules folding together en masse. At the coarsest level, it appears that in transitioning to the native state, a given amino acid sequence takes on roughly the same route and proceeds through roughly the same intermediates and transition states. Often folding involves first the establishment of regular secondary and super secondary structures, particularly alpha helices and beta sheets, and afterwards tertiary structure. Formation of quaternary structure usually involves the "assembly" or "coassembly" of subunits that have already folded. The regular alpha helix and beta sheet structures fold rapidly because they are stabilized by intramolecular hydrogen bonds, as was first characterized by Linus Pauling. Protein folding may involve covalent bonding in the form of disulfide bridges formed between two cysteine residues or the formation of metal clusters. Shortly before settling into their more energetically favorable native conformation, molecules may pass through an intermediate "molten globule" state.
+Folding is a spontaneous process independent of energy inputs from nucleoside triphosphates. The passage of the folded state is mainly guided by hydrophobic interactions, formation of intramolecular hydrogen bonds, and van der Waals forces, and it is opposed by conformational entropy.
+While these macromolecules may be regarded as "folding themselves", the process also depends on the solvent (water or lipid bilayer), the concentration of salts, the temperature, and the presence of molecular chaperones.
+The essential fact of folding, however, remains that the amino acid sequence of each protein contains the information that specifies both the native structure and the pathway to attain that state. This is not to say that nearly identical amino acid sequences always fold similarly. Conformations differ based on environmental factors as well; similar proteins fold differently based on where they are found.
+
Motivation:
+Misfolded proteins almost always fail to function correctly. Also excess misfolded protein can accumulate and interfere with the functioning of the cell. Consequently misfolded proteins are a feature of a large number of diseases, including Alzheimer's, Creutzfeldt-Jakob disease (CJD), cystic fibrosis, and many cancers. Understanding the difference in the folding process can bring insight into the causes of these diseases at the molecular level.
+
Secondary structure analysis by Circular Dichroism Spectroscopy
+
Circular Dichroism or CD Spectrocsopy measures the difference in absorption of right and left circularly polarized light. The difference arises if there is asymmetry present in a compact arrangement of chromophores that are within interacting distances. The typical chromophore in a protein or polypeptide is the peptide bond. The spatial juxtaposition of these peptide groups changes as a function of the backbone geometry. It is therefore expected that as the backbone conformation changes, the CD signal will also change. Therefore the CD spectra of polypeptides are highly sensitive to the overall conformation of a polypeptide chain.
+CD values are measured as mean residue ellipticities in millidegree units and reported as MRE ([θ] = θ/10 n.l.c) where n is the number of peptide bonds, l is the cuvette pathlength in cm, c is the concentration in Molarity and θ is the observed ellipticity in mdeg. This normalizes concentration or number of peptide bond differences between two samples. The unit of [θ] is: deg.cm2.dmol-1.
+Theoretical calculations suggest that the random coil conformation gives rise to a negative minimum < 200nm. β-sheet conformation is characterized by one negative band around 216 nm and a positive band around 197 nm. The α- helix gives rise to two negative bands (double minima), around 222nm (n-π*) and 208nm (π-π* parallel) with almost equal intensity. There is also a strong positive band at around 192nm (π-π* perpendicular).
+
| Protein backbone. | +Aromatic amino acids. | +||
|---|---|---|---|
| CD sensitive to secondary structure change. | +CD sensitive to tertiary structure change. | +||
| 170nm | +Far-UV | +250nm | +NearUV | +
Structural types identified:
+
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+Structural types identified:
+
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+||
+
Changes in the circular dichroism spectra of bio-molecules represent changes in their secondary and tertiary structures. When this is coupled with the facts that (i) spectra can be recorded in minutes and (ii) single wavelength kinetics can be recorded from milliseconds onwards, it can be seen that CD is a particularly powerful tool to follow dynamic changes in protein structure. For instance, changes induced by changing temperature, pH, ligands, or denaturants are all commonly studied.
+
CD spectroscopy and its use to study thermal denaturation of protein:
+Continuing from what we have learnt in experiment 6 and 7, the far UV region of a protein circular dichroism (CD) spectrum shows spectral features related to the structure of the backbone of the protein, and is directly relevant to the secondary structural elements that make up the protein structure. The spectral signatures can be used to predict the secondary structure makeup of a particular proteins structure as discussed in experiments 6 and 7. We can also similarly analyze the protein's secondary structure against temperature as the parameters as sighted below i.e. the process of thermal denaturation of protein with CD spectra.
++
+
+ |
+
| This picture is taken from the website of Applied Photophysics Ltd, 21 Mole Business Park, Leatherhead,Surrey KT22 7AG, United Kingdom + |
+
+The above figure illustrates clearly the change in CD signature as a function of temperature. Comparison of known CD signatures with the data in this experiment suggests that the initial CD signature is a combination of α-helical and β-sheet structure which, on heating, loses much of its α-helical content to an unfolded state, whilst retaining some β-sheet character.
+
Protein Denaturation
+
Protein denaturation is commonly defined as any non covalent change in the structure of a protein. This change may alter the secondary, tertiary or quaternary structure of the molecules. When using this definition it should be noted that what constitutes denaturation is largely dependent upon the method utilized to observe the protein molecule. Some methods can detect very slight changes in structure while other requires rather large alterations in structure before changes are observed.
+Mechanism of protein denaturation:
+Denaturation of proteins involves the disruption and possible destruction of both the secondary and tertiary structures. Since denaturation reactions are not strong enough to break the peptide bonds, the primary structure (sequence of amino acids) remains the same after a denaturation process. Denaturation disrupts the normal alpha-helix and beta-sheets in a protein and uncoils it into a random shape.
+How denaturation occurs at levels of protein structure:
+-
+
- In quaternary structure denaturation, protein sub-units are dissociated and/or the spatial arrangement of protein subunits is disrupted. +
- Tertiary structure denaturation involves the disruption of:
+
-
+
- Covalent interactions between amino acid side-chains (such as disulfide bridges between cysteine groups) +
- Noncovalent dipole-dipole interactions between polar amino acid side-chains (and the surrounding solvent) +
- Van der Waals (induced dipole) interactions between nonpolar amino acid side-chains. +
+ - In secondary structure denaturation, proteins lose all regular repeating patterns such as alpha-helices and beta-pleated sheets, and adopt a random coil configuration. +
- Primary structure, such as the sequence of amino acids held together by covalent peptide bonds, is not disrupted by denaturation. +
+
Thermal Denaturation
+
Heat can be used to disrupt hydrogen bonds and non-polar hydrophobic interactions. This occurs because heat increases the kinetic energy and causes the molecules to vibrate so rapidly and violently that the bonds are disrupted. The proteins in eggs denature and coagulate during cooking. Other foods are cooked to denature the proteins to make it easier for enzymes to digest them. Medical supplies and instruments are sterilized by heating to denature proteins in bacteria and thus destroy the bacteria.
+When proteins are exposed to increasing temperature, losses of solubility or enzymatic activity occurs over a fairly narrow range. Depending upon the protein studied and the severity of the heating, these changes may or may not be reversible.
+Mechanism of thermal denaturation: Why proteins unfold at higher temperature?
+As the temperature is increased, a number of bonds in the protein molecule are weakened. The first affected are the long range interactions that are necessary for the presence of tertiary structure. As these bonds are first weakened and are broken, the protein obtains a more flexible structure and the groups are exposed to solvent. If heating ceases at this stage the protein should be able to readily refold to the native structure. As heating continues, some of the cooperative hydrogen bonds that stabilize helical structure will begin to break. As these bonds are broken, water can interact with and form new hydrogen bonds with the amide nitrogen and carbonyl oxygens of the peptide bonds. The presence of water further weakens nearby hydrogen bonds by causing an increase in the effective dielectric constant near them. As the helical structure is broken, hydrophobic groups are exposed to the solvent.
+The effect of exposure of new hydrogen bonding groups and of hydrophobic groups is to increase the amount of water bound by the protein molecules. The unfolding that occur increase the hydrodynamic radius of the molecule causing the viscosity of the solution to increase. The net result will be an attempt by the protein to minimize its free energy by burying as many hydrophobic groups while exposing as many polar groups as possible to the solvent. While this is analogous to what occurred when the protein folded originally, it is happening at a much higher temperature. This greatly weakens the short range interaction that initially direct protein folding and the structures that occur will often be vastly different from the native protein.
+The increased water binding noted in the early stages of denaturation may be retained following hydrophobic aggregations. The loss of solubility that occurs will greatly reduce the viscosity to a level below that of the native proteins.
+Under some conditions proteins will not fold into their biochemically functional forms. Temperatures above or below the range that cells tend to live in will cause thermally unstable proteins to unfold or "denature" (this is why boiling makes an egg white turn opaque). High concentrations of solutes, extremes of pH, mechanical forces, and the presence of chemical denaturants can do the same. Protein thermal stability is far from constant, however. For example, hyperthermophilic bacteria have been found that grow at temperatures as high as 122 °C, which of course requires that their full complement of vital proteins and protein assemblies be stable at that temperature or above.
+Exposure of most proteins to high temperatures results in irreversible denaturation. Some proteins, like caseins, however, contain little if any secondary structure and have managed to remove their hydrophobic groups from contact with the solvent without the need for extensive structure. This lack of secondary structure causes these proteins to be extremely resistant to thermal denaturation.
+Thermal denaturation of protein (by circular dichroism):
+This protocol is used to determine the stability of protein. The thermal denaturation of protein can be monitored by circular dichroism spectrum .As explained in this theory portion above, Circular dichroism is one of the most general and basic tools to study protein folding. Circular dichroism spectroscopy measures the absorption of circularly polarized light. In proteins, structures such as alpha helicies and beta sheets are chiral, and thus absorb such light. The absorption of this light acts as a marker of the degree of foldedness of the protein ensemble. This technique can be used to measure equilibrium unfolding of the protein by measuring the change in this absorption as a function of denaturant concentration or temperature. Usually, one wavelength (we use 222nm for our helical protein) is chosen to monitor. A denaturant melt measures the free energy of unfolding as well as the protein's Tm value.
++
+
Denaturation midpoint:
+
++The transition midpoint (Tm) is the temperature where 50% of the protein is in its native conformation and the other 50% is in maximum denatured / unfolded conformation. The higher the Tm, the more stable is the molecule. In this experiment, it is determined by measuring a series of CD spectra of the protein at various temperatures, the transition temperature is usually taken as the point where 50% of the protein structure is in maximally unfolded state. +
+This type of spectroscopy can also be combined with fast-mixing devices, such as stopped flow, to measure protein folding kinetics and to generate chevron plots.In this experiment we will study the effect of temperature on protein denaturation using Rhodopsin as a model system.
+
RHODOPSIN
+
Rhodopsin, also known as visual purple, is a pigment of the retina that is responsible for both the formation of the photoreceptor cells and the first events in the perception of light. Rhodopsins belong to the G-protein coupled receptor family and are extremely sensitive to light, enabling vision in low-light conditions. Exposed to light, the pigment immediately photobleaches, and it takes about 30 minutes to regenerate fully in humans.
+Property
+Proteins interact with other molecules---substrates, inhibitors, ions, nucleic acids, carbohydrates, lipids, and other proteins--as ways in which they carry out their various functions. By the time rhodopsin reaches its membrane location in the outer segment, it has already interacted with dozens of proteins involved in such activities as its biosynthesis, acetylation, glycosylation, and transport. It has bound two molecules of palmitate and one molecule of 11-cis retinal and has become embedded in a membrane lipid bilayer. Rhodopsin is now ready to carry out its role in phototransduction.
+After photoactivation, rhodopsin exerts its effect on rod cell biochemistry by interacting with rod cell proteins through its cytoplasmic surface. As described previously, photoactivated rhodopsin (R*) binds and activates the G-protein, transducin, initiating the phototransduction cascade. R* is then phosphorylated by rhodopsin kinase, and the resulting R*P is bound by arrestin, preventing further activation of transducin. Thus, rhodopsin interacts with three major proteins---transducin, rhodopsin kinase, and arrestin--and is then dephosphorylated by a phosphatase, as part of its functional cycle.
+Importance
+Mutation of the rhodopsin gene is a major contributor to various retinopathies such as
Structure
+Rhodopsin consists of the protein moiety opsin and a reversibly covalently bound cofactor, retinal.Opsin, a bundle of seven transmembrane helices connected to each other by protein loops, binds retinal (a photoreactive chromophore), which is located in a central pocket on the seventh helix at a lysine residue. Retinal lies horizontally with relation to the membrane. Each outer segment disc contains thousands of visual pigment molecules. About half the opsin is within the lipid bilayer. Retinal is produced in the retina from Vitamin A, from dietary beta-carotene. Isomerization of 11-cis-retinal into all-trans-retinal by light induces a conformational change (bleaching) in opsin continuing with metarhodopsin II, which activates the associated G protein transducin and triggers a second messenger cascade.
+The structure of rhodopsin has been studied in detail via x-ray crystallography on rhodopsin crystals. Several models (e.g. the bicycle-pedal mechanism, hula-twist mechanism) attempt to explain how the retinal group can change its conformation without clashing with the enveloping rhodopsin protein pocket.
+Recent data supports that it is a functional monomer as opposed to a dimer, which was the paradigm of G-coupled protein receptors for many years.
+
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+
+ t1=content.index('')
+ print t1
+ f.seek(t1+13)
+ s1=s1+f.read()
+ #print s1
+ f.seek(t1+13)
+ f.write(s1)
+ f.seek(0)
+ content=f.read()
+ t= content.index('')
+ f.seek(t+13)
+ s='
' +while sectionNumber<=len(sectionno): + tag="" + att = ''+'experiment-article-section-'+str(sectionNumber)+'-heading' + tagger = soup.findAll('div', attrs={'id':att,'class':'heading'}) + if not tagger: + sectionNumber=sectionNumber+1 + continue + tag1=str(tagger[0].text) + tag1=tag1.strip() + print tag1 + if tag1=='Prerequisite S/W': + tag1='Prerequisites' + st+='
'+heading+'
'+s + s=s+f.read() + f.seek(t+13) + f.write(s) +f=open("template1.html",'r') +template=f.read() +f=open("content.html",'r') +srchtml=f.read() +f.close() +soup = BeautifulSoup(srchtml, 'html.parser') +sectionno=soup.find_all('section') +d=['introduction','theory','objective','experiment','manual','quizzes','further_readings','procedure','hh','ll'] +print len(sectionno) +sectionNumber=1 +st="" + +att = ''+'experiment-article-heading' +tagger = soup.findAll('header', attrs={'id':att,'class':'heading'}) +heading=str(tagger[0].text) +heading=heading.strip() +print heading +att = ''+'experiment-header-heading' +tagger = soup.findAll('div', attrs={'id':att,'class':'heading'}) +heading1=str(tagger[0].text) +print heading +heading1=heading1.strip() +breadcrumb='Chemical Sciences →'+heading1+' →List Of Experiments' +while sectionNumber<=len(sectionno): + tag="" + att = ''+'experiment-article-section-'+str(sectionNumber)+'-heading' + tagger = soup.findAll('div', attrs={'id':att,'class':'heading'}) + if not tagger: + sectionNumber=sectionNumber+1 + continue + tag1=str(tagger[0].text) + tag1=tag1.strip() + print tag1 + if tag1=='Prerequisite S/W': + tag1='Prerequisites' + st+='
'+tag1+'
' + sectionNumber=sectionNumber+1 +st+='Feedback
' +sectionNumber=1 +while sectionNumber<=len(sectionno): + tag="" + tag="" + att = ''+'experiment-article-section-'+str(sectionNumber)+'-heading' + tagger = soup.findAll('div', attrs={'id':att,'class':'heading'}) + if not tagger: + sectionNumber=sectionNumber+1 + continue + tag1=str(tagger[0].text) + tag1=tag1.strip() + if tag1=='Prerequisite S/W': + tag1='Prerequisites' + att = ''+'experiment-article-section-'+str(sectionNumber)+'-content' + tagger = soup.findAll('div', attrs={'id':att,'class':'content'}) + + tag+=str(tagger[0]) + #print tag + #print st + + + writefile(tag1+'.html',tag,st,heading) + #print sectionNumber + sectionNumber=sectionNumber+1 + +f=open("Feedback.html",'w+') +f.write(template) +f.seek(0) + +content = f.read() +content=content.replace('Disciplines and Domains',breadcrumb) +f.seek(0) +f.write(content) +#print content +f.seek(0) +content=f.read() +k=content.index('')
+
+t1=content.index('')
+print t1
+f.seek(t1+13)
+st=st+f.read()
+#print s1
+f.seek(t1+13)
+f.write(st)
+f.seek(0)
+content=f.read()
+t= content.index('')
+f.seek(t+13)
+s='Welcome to Virtual Labs - A MHRD Govt of india Initiative
+
+
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+Welcome to Virtual Labs - A MHRD Govt of india Initiative
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+Welcome to Virtual Labs - A MHRD Govt of india Initiative
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+Welcome to Virtual Labs - A MHRD Govt of india Initiative
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+Welcome to Virtual Labs - A MHRD Govt of india Initiative
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+Welcome to Virtual Labs - A MHRD Govt of india Initiative
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+Welcome to Virtual Labs - A MHRD Govt of india Initiative
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+Welcome to Virtual Labs - A MHRD Govt of india Initiative
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'+heading+'
'+'Feedback' +s=s+f.read() +f.seek(t+13) +f.write(s) diff --git a/src/lab/exp8/template1.html b/src/lab/exp8/template1.html new file mode 100644 index 0000000..382fb88 --- /dev/null +++ b/src/lab/exp8/template1.html @@ -0,0 +1,205 @@ + + + + + + + + + + +
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diff --git a/src/lab/exp9/Experiment.html b/src/lab/exp9/Experiment.html
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+ Disciplines and Domains
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diff --git a/src/lab/exp9/Feedback.html b/src/lab/exp9/Feedback.html
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+ Chemical Sciences →Circular Dichroism Virtual Lab →List Of Experiments
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+ Chemical Sciences →Circular Dichroism Virtual Lab →List Of Experiments
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+ To Study The Effect of Chemical Denaturants on Protein Stability.
+
+Click here to start the experiment. + + +
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diff --git a/src/lab/exp9/Introduction.html b/src/lab/exp9/Introduction.html
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+ Chemical Sciences →Circular Dichroism Virtual Lab →List Of Experiments
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+ Chemical Sciences →Circular Dichroism Virtual Lab →List Of Experiments
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+ To Study The Effect of Chemical Denaturants on Protein Stability.
Feedback
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diff --git a/src/lab/exp9/Manual.html b/src/lab/exp9/Manual.html
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+ Chemical Sciences →Circular Dichroism Virtual Lab →List Of Experiments
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+ Chemical Sciences →Circular Dichroism Virtual Lab →List Of Experiments
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+ To Study The Effect of Chemical Denaturants on Protein Stability.
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+In Experiment 8 we have seen how a protein denatures at high temperature. But temperature is not the sole factor that causes denaturation of proteins. The chemical environment is another factor that is responsible for denaturation of protein. Change of pH of the environment or presence of denaturing chemical in high concentration in the medium causes denaturation of protein.
++In our daily life we have seen many such examples. Formation of paneer from milk is one of them. The following video shows how egg protein albumin denatures in a highly acidic medium.
+
+
+In chemical denaturation the secondary bonds holding the protein segments together are disrupted by some chemicals capable of forming equally strong or stronger bonds with the groups holding the conformation together. For disrupting the hydrogen bonds, urea or guanidine hydrochloride are used. At high concentrations of these substances (e.g, 8M urea or 5M guanidine hydrochloride) many proteins adopt a highly unfolded conformation in solution. Proteins of multiple subunits are likely to be separated into their constituent polypeptide chains. Other proteins aggregate upon denaturation in urea or guanidine hydrochloride which is frequently due to the formation of disulphide bridges between sulphydryl groups made accessible by the unfolding of the polypeptide chains. +
+
+This experiment has two parts. In the first part we will observe how helical membrane protein Rhodopsin looses its native structure and unfolds in presence of some denaturing chemicals like urea, SDS, GuHCl and TFA. We will compare their denaturing capability. In the second part of the experiment we will observe how the extend of unfolding depends on the concentration of the chemical. We will try to see the transition between folded and unfolded state and find out the concentration at which the transition takes place.
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diff --git a/src/lab/exp9/Objective.html b/src/lab/exp9/Objective.html
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+ Chemical Sciences →Circular Dichroism Virtual Lab →List Of Experiments
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+ Chemical Sciences →Circular Dichroism Virtual Lab →List Of Experiments
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+ To Study The Effect of Chemical Denaturants on Protein Stability.
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1. Video Tutorial
++ + +
++
click here to ENLARGE.
++
2. Step-by-step Procedure
+The experiment section has three sections – Animation explaining extent of protein unfolding depending on chemical denaturant concentration, part 1 and part 2.
+
Section 1
+
Click on first section. Observe the animation showing MD simulation of unfolding of a helical protein at different concentration (2M,4M,6M and 8M).
+
+Click the button BACK to go back to the first page and perform the experiment.
+
Section 2
+
Click on PART1. A list of chemicals will appear along with their molecular level picture and physiochemical descriptions. Select any number chemicals whose denaturing capability you want to study. Click PROCEED to go to experiment setup. +
+
Steps to follow in this page will appear on top left corner of the page. Follow the instructions. After you are done click on the monitor to see the results.
++
Go through the NOTE carefully and then click Result.
++
Select the chemical from bottom left panel and the CD spectra. Secondary structure content are listed on the right top panel.
++
Once you are done click to “Go to analysis”.
++
Answer the questions and submit your answer.
++
Section 3
Click on PART2. This will lead to the experimental set-up. Steps to follow in this page will appear on top left corner of the page. Follow the instructions. After you are done click on the monitor to see the results.
+
+See the CD spectrum of Rhodopsin at different concentrations. Use CONTROLS to see spectrum at different concentrations. Take your mouse on the graph. Select the MRE value corresponding to 222nm and fill up the TABLE. Click on Plot once you have finished filling up the TABLE. You can come back to this step by clicking Go Back.
+
+Once you are done click Go To Analysis.
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diff --git a/src/lab/exp9/Quizzes.html b/src/lab/exp9/Quizzes.html
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+ Chemical Sciences →Circular Dichroism Virtual Lab →List Of Experiments
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+ To Study The Effect of Chemical Denaturants on Protein Stability.
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The purpose of this lab is to make students familiar with the concept that structure of a protein is highly dependent on the chemical environmental. By performing this experiment a student will also get familiar with the use of CD spectroscopy method to study the effect of chemical denaturant on protein structure. By doing this experiment a student will be able to,
++
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+
- Make the student understand how a chemical environment denatures a protein. +
- Have an idea of the effect of different types of chemicals on the protein structure. +
- Have an idea of the effect of different concentrations of chemical environment on the protein structure. +
- Understand how CD spectroscopy can be used to study unfolding of proteins. +
- Observe transition between folded and unfolded state. +
- Determine the critical concentration of transition from folded to unfolded structure using CD spectra at different concentrations. +
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diff --git a/src/lab/exp9/Theory.html b/src/lab/exp9/Theory.html
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+ Chemical Sciences →Circular Dichroism Virtual Lab →List Of Experiments
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+ To Study The Effect of Chemical Denaturants on Protein Stability.
+
+Click here for pre-experiment quiz.
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diff --git a/src/lab/exp9/scraplab.py b/src/lab/exp9/scraplab.py
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+from bs4 import BeautifulSoup
+heading=""
+breadcrumb=""
+def writefile(fname,s,s1,heading):
+ f=open(fname, 'w+')
+ f.write(template)
+ f.seek(0)
+
+ content = f.read()
+ content=content.replace('Disciplines and Domains',breadcrumb)
+ f.seek(0)
+ f.write(content)
+ print content
+
+ f.seek(0)
+ content=f.read()
+ k=content.index('
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Previously, we have already discussed in details in experiment 8 the causes and the effects of "protein folding" and "protein denaturation", the denatured state and the mechanism behind the same. Continuing from where we had left our study with temperature related effects (thermal denaturation), now we look to extend our search further penetrating into the effects due to chemical denaturants in this experiment. So, without referring again the details of the earlier explained points we limit our discussion to focus in the perspective of the chemical denaturants and their contribution to our cause.
+Protein Stability
+
To be biologically active, proteins must adopt specific folded three-dimensional, tertiary structures. Yet the genetic information for the protein specifies only the primary structure that is the linear sequence of amino acids in the polypeptide backbone. Many purified proteins can spontaneously refold in vitro after being completely unfolded, so the three-dimensional structure must be determined by the primary structure (Anfinsen, 1973). Different conformations of a protein differ only in the angle of rotation about the bonds of the backbone and amino acid side-chains. It may, therefore, appear surprising that a protein folds into a single unique conformation from all the possible rotational conformations available around single bonds in the primary structure of a protein. The question arises as to when a protein folds up to its native conformation, does this structure represent a local or a global energy minimum? The native structure of the protein may correspond to a metastable state with a very long lifetime. If proteins are only metastable, their structures must be grossly different from the most stable one. The polypeptide may adopt a structure corresponding to the global minimum.
+This led to the idea of a biased random search resulting in faster folding since a proportion of the conformations would be sterically disallowed. A framework model proposed (Baldwin, 1989; Kim and Baldwin, 1990) that the folded structure was formed by packing together pre-existing individual elements of secondary structure which had significant stability in the unfolded protein. It became evident through some experiments that the equilibrium of unfolding of proteins does not always follow a simple two-state model in which only the native and fully unfolded states are significantly populated (Wong and Tanford, 1973). An intermediate compact structure known as the molten globule which is different from the native structure and whose formation is determined mainly by non-specific interactions of amino acid residues with their environment was presented. Specific interactions could direct the folding pathway by stabilizing folded conformations. The best studied example is bovine pancreatic trypsin inhibitor (Creighton, 1978). For this protein it was shown that formation of a disulphide bridge stabilizes secondary structure elements, and the protein refolds by a specific pathway of disulphide bond formation and rearrangement (Directed folding model). Since non covalent forces act on the primary structure to cause a protein to fold into a unique conformational structure and then stabilize the native structure against denaturation processes, it is of importance to understand the properties of these forces.
+
Chemical Denaturation
+
Protein denaturation has been defined in several ways, for example as a change in solubility (Mirsky, 1941) or by simultaneous changes in chemical, physical and biological properties (Neurath et al., 1944; Langmuir, 1938) under some standard reference set of conditions (Timasheff and Gibbs, 1957).
+The term chemical denaturation denotes the response of the native protein to acid, alkali, and a variety of other chemical agents which cause marked changes in the protein structure. Therefore chemical denaturation may be defined in very general terms as any modification of the secondary, tertiary or quaternary structure of the protein molecule, excluding any breaking of covalent bond (Joly, 1965).
++Denaturation midpoint is defined as that denaturant concentration (Cm) at which both the folded and unfolded states of a protein are equally populated at equilibrium. It is usually determined by recording CD spectra of protein at various concentrations of the denaturant. The mid point of transition is taken as the point where 50% of the protein’s secondary structure is in conformation 1 and the other 50% is in conformation 2. +
++
Mathematical model behind chemical denaturation
+In the less expensive technique of equilibrium unfolding, the fractions of folded and unfolded molecules (denoted as pN and pU, respectively) are measured as the solution conditions are gradually changed from those favoring the native state to those favoring the unfolded state, e.g., by adding a denaturant such as guanidinium hydrochloride or urea. (In equilibrium folding, the reverse process is carried out.) Given that the fractions must sum to one and their ratio must be given by the Boltzmann factor, we have
++
+
+
+ |
+
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+ |
+
+Protein stabilities are typically found to vary linearly with the denaturant concentration. A number of models have been proposed to explain this observation prominent among them being the denaturant binding model, solvent-exchange model and the Linear Energy Model (LEM). All of the models assume that only two thermodynamic states are populated/de-populated upon denaturation. They could be extended to interpret more complicated reaction schemes.
+The denaturant binding model assumes that there are specific but independent sites on the protein molecule (folded or unfolded) to which the denaturant binds with an effective (average) binding constant k. The equilibrium shifts towards the unfolded state at high denaturant concentrations as it has more binding sites for the denaturant relative to the folded state (Δn). In other words, the increased number of potential sites exposed in the unfolded state is seen as the reason for denaturation transitions. An elementary treatment results in the following functional form:
+
+
+ |
+
where ΔGw is the stability of the protein in water and [D] is the denaturant concentration. Thus the analysis of denaturation data with this model requires 7 parameters: ΔGw, Δn, k, and the slopes and intercepts of the folded and unfolded state baselines.
+The solvent exchange model (also called the 'weak binding model' or 'selective solvation') of Schellman invokes the idea of an equilibrium between the water molecules bound to independent sites on protein and the denaturant molecules in solution. It has the form:
+
+
+ |
+
where K is the equilibrium constant for the exchange reaction and Xd is the mole-fraction of the denaturant in solution. This model tries to answer the question of whether the denaturant molecules actually bind to the protein or they seem to be bound just because denaturants occupy about 20-30 % of the total solution volume at high concentrations used in experiments, i.e. non-specific effects - and hence the term 'weak binding'. As in the denaturant-binding model, fitting to this model also requires 7 parameters. One common theme obtained from both these models is that the binding constants (in the molar scale) for urea and guanidinium hydrochloride are small: ~ 0.2 M-1 for urea and 0.6M-1 for GuHCl.
+Intuitively, the difference in the number of binding sites between the folded and unfolded states is directly proportional to the differences in the accessible surface area. This forms the basis for the LEM which assumes a simple linear dependence of stability on the denaturant concentration. The resulting slope of the plot of stability versus the denaturant concentration is called the m-value. In pure mathematical terms, m-value is the derivative of the change in stabilization free energy upon the addition of denaturant. However, a strong correlation between the accessible surface area (ASA) exposed upon unfolding, i.e. difference in the ASA between the unfolded and folded state of the studied protein (dASA), and the m-value has been documented by Pace and co-workers. In view of this observation, the m-values are typically interpreted as being proportional to the dASA. There is no physical basis for the LEM and is purely empirical, though it is widely used in interpreting solvent-denaturation data. It has the general form:
+
+
+ |
+
+where the slope m is called the "m-value"(> 0 for the above definition) and [D]1/2 (also called Cm) represents the denaturant concentration at which 50% of the molecules are folded (the denaturation midpoint of the transition, where pN = pU = 1 / 2).
+In practice, the observed experimental data at different denaturant concentrations are fit to a two-state model with this functional form for ΔG, together with linear baselines for the folded and unfolded states. The m and [D]1/2 are two fitting parameters, along with four others for the linear baselines (slope and intercept for each line); in some cases, the slopes are assumed to be zero, giving four fitting parameters in total. The conformational stability ΔG can be calculated for any denaturant concentration (including the stability at zero denaturant) from the fitted parameters m and [D]1/2. When combined with kinetic data on folding, the m-value can be used to roughly estimate the amount of buried hydrophobic surface in the folding transition state.
+
Chemical Denaturants
+
Chemical Denaturants are classified into numerous categories:
+
-
+
- Acids +
- Acetic acid +
- Trichloroacetic acid 12% in water +
- Sulfosalicylic acid +
- Dichloroacetic acid +
- Solvents +
- Ethanol +
- Methanol +
- Formamide +
- Dimethyl Formamide +
- Cross-linking reagents +
- Formaldehyde +
- Glutaraldehyde +
- Chaotropic agents +
- Urea 6 - 8 mol/l +
- Guanidinium chloride 6 mol/l +
- Lithium perchlorate 4.5 mol/l +
- Sodium dodecyl sulphate +
- Disulfide bond reducers +
- 2-Mercaptoethanol +
- Dithiothreitol +
- TCEP (tris(2-carboxyethyl)phosphine) +
- Inorganic salts +
Acidic protein denaturants include:
+-
+
Most organic solvents are denaturing, including:
+-
+
Cross-linking agents for proteins include:
+-
+
Chaotropic agents include:
+-
+
Agents that break disulfide bonds by reduction include:
+-
+
Lithium bromide, potassium thiocyanate, and sodium iodide.
+ ++
The process of chemical denaturation of a protein:
+In chemical denaturation the secondary bonds holding the protein segments together are disrupted by some chemicals capable of forming equally strong or stronger bonds with the groups holding the conformation together. For disrupting the hydrogen bonds, urea or guanidine hydrochlorides are used. At high concentrations of these substances (e.g., 8M urea or 5M guanidine hydrochloride) many proteins adopt a highly unfolded conformation in solution. Proteins of multiple subunits are likely to be separated into their constituent polypeptide chains. Other proteins aggregate upon denaturation in urea or guanidine hydrochloride which is frequently due to the formation of disulphide bridges between sulphydryl groups made accessible by the unfolding of the polypeptide chains. Such reactions may be inhibited by the addition of iodoacetate (Friedli, PhD thesis, chapter 3 and 5). Under these conditions, the denatured molecules remain in solution and may revert into native molecules if the denaturing agent is slowly dialysed away. Powerful detergents like SDS disrupt both hydrophobic and hydrogen bonds and effectively solvate the denatured molecule. Beta-Mercaptoethanol and dithiothreitol (DTT) disrupt disulphide bonds and can be used in conjunction with urea or SDS to fully solubilize protein molecules.
++
+ |
+
An example of chemical denaturantion:
++Membrane proteins are not soluble. The first work on the folding of membrane proteins was carried out with bacteriorhodopsin (bR) which was successfully refolded into phospholipids via transfer into sodium dodecyl sulfate (SDS) micelles after denaturation by trifluoroacetic acid (TFA). This has established bR as a model system for studying the folding of helical membrane proteins. Kinetic studies of refolding of bR from an SDS-denatured state into lipid/detergent micelles have enabled detection of folding intermediates in its folding pathway. +
++Recently, studies with rhodopsin in phospholipid bicelles revealed a significant decrease of 50% of its helical content in the presence of urea, but it could not be refolded from such a largely unfolded state.
+Different chemical denaturants like 4 M TFA, 8 M GuHCl, 10 M urea, 3% SDS + 8Murea, and 30% SDS have their effect on the secondary structure of Rhodopsin to different extents. Some of them cause aggregation in the unfolded structure. So to study the unfolded structure of Rhodopsin in the presence of chemical denaturants it is important to reinvestigate whether aggregation is taking place or not.
+
Goal of The Experiment
+In this experiment we will see how different denaturing agents are effecting secondary structure of Rhodopsin differently and at the same time we will study the secondary structure of Rhodopsin as a function of the concentration of the denaturant.
+
')
+
+ t1=content.index('')
+ print t1
+ f.seek(t1+13)
+ s1=s1+f.read()
+ #print s1
+ f.seek(t1+13)
+ f.write(s1)
+ f.seek(0)
+ content=f.read()
+ t= content.index('')
+ f.seek(t+13)
+ s='
' +while sectionNumber<=len(sectionno): + tag="" + att = ''+'experiment-article-section-'+str(sectionNumber)+'-heading' + tagger = soup.findAll('div', attrs={'id':att,'class':'heading'}) + if not tagger: + sectionNumber=sectionNumber+1 + continue + tag1=str(tagger[0].text) + tag1=tag1.strip() + print tag1 + if tag1=='Prerequisite S/W': + tag1='Prerequisites' + st+='
'+heading+'
'+s + s=s+f.read() + f.seek(t+13) + f.write(s) +f=open("template1.html",'r') +template=f.read() +f=open("content.html",'r') +srchtml=f.read() +f.close() +soup = BeautifulSoup(srchtml, 'html.parser') +sectionno=soup.find_all('section') +d=['introduction','theory','objective','experiment','manual','quizzes','further_readings','procedure','hh','ll'] +print len(sectionno) +sectionNumber=1 +st="" + +att = ''+'experiment-article-heading' +tagger = soup.findAll('header', attrs={'id':att,'class':'heading'}) +heading=str(tagger[0].text) +heading=heading.strip() +print heading +att = ''+'experiment-header-heading' +tagger = soup.findAll('div', attrs={'id':att,'class':'heading'}) +heading1=str(tagger[0].text) +print heading +heading1=heading1.strip() +breadcrumb='Chemical Sciences →'+heading1+' →List Of Experiments' +while sectionNumber<=len(sectionno): + tag="" + att = ''+'experiment-article-section-'+str(sectionNumber)+'-heading' + tagger = soup.findAll('div', attrs={'id':att,'class':'heading'}) + if not tagger: + sectionNumber=sectionNumber+1 + continue + tag1=str(tagger[0].text) + tag1=tag1.strip() + print tag1 + if tag1=='Prerequisite S/W': + tag1='Prerequisites' + st+='
'+tag1+'
' + sectionNumber=sectionNumber+1 +st+='Feedback
' +sectionNumber=1 +while sectionNumber<=len(sectionno): + tag="" + tag="" + att = ''+'experiment-article-section-'+str(sectionNumber)+'-heading' + tagger = soup.findAll('div', attrs={'id':att,'class':'heading'}) + if not tagger: + sectionNumber=sectionNumber+1 + continue + tag1=str(tagger[0].text) + tag1=tag1.strip() + if tag1=='Prerequisite S/W': + tag1='Prerequisites' + att = ''+'experiment-article-section-'+str(sectionNumber)+'-content' + tagger = soup.findAll('div', attrs={'id':att,'class':'content'}) + + tag+=str(tagger[0]) + #print tag + #print st + + + writefile(tag1+'.html',tag,st,heading) + #print sectionNumber + sectionNumber=sectionNumber+1 + +f=open("Feedback.html",'w+') +f.write(template) +f.seek(0) + +content = f.read() +content=content.replace('Disciplines and Domains',breadcrumb) +f.seek(0) +f.write(content) +#print content +f.seek(0) +content=f.read() +k=content.index('')
+
+t1=content.index('')
+print t1
+f.seek(t1+13)
+st=st+f.read()
+#print s1
+f.seek(t1+13)
+f.write(st)
+f.seek(0)
+content=f.read()
+t= content.index('')
+f.seek(t+13)
+s='Welcome to Virtual Labs - A MHRD Govt of india Initiative
+
+
+
+
+
+
+
+
+
+
+
+
+
+
+
+
+
+
+
'+heading+'
'+'Feedback' +s=s+f.read() +f.seek(t+13) +f.write(s) diff --git a/src/lab/exp9/template1.html b/src/lab/exp9/template1.html new file mode 100644 index 0000000..382fb88 --- /dev/null +++ b/src/lab/exp9/template1.html @@ -0,0 +1,205 @@ + + + + + + + + + + +
+
+
+
+
+
+
+
+
+
+
+
+
+
+
+
+
+
+
+
+
+
+
+
+
+
diff --git a/src/lab/scrap.py b/src/lab/scrap.py
new file mode 100644
index 0000000..d63fba8
--- /dev/null
+++ b/src/lab/scrap.py
@@ -0,0 +1,125 @@
+from bs4 import BeautifulSoup
+count=1
+heading=""
+breadcrumb=""
+def writefile(fname,s,s1,heading):
+ f=open(fname, 'w+')
+ f.write(template)
+ f.seek(0)
+
+ content = f.read()
+ content=content.replace('Disciplines and Domains',breadcrumb)
+ f.seek(0)
+ f.write(content)
+ #print content
+ f.seek(0)
+ content=f.read()
+ k=content.index('
+
+
+
+
+
+
+
+
+
+
+
+
+
+ +
+
+
+
+
+
+
+ Disciplines and Domains
+ +
+
+
+
+
+
+
+
+ ')
+
+ t1=content.index('')
+ print t1
+ f.seek(t1+13)
+ s1=s1+f.read()
+ #print s1
+ f.seek(t1+13)
+ f.write(s1)
+ f.seek(0)
+ content=f.read()
+ t= content.index('')
+ f.seek(t+13)
+ s='
' +while sectionNumber<=len(sectionno): + tag="" + att = ''+'lab-article-section-'+str(sectionNumber)+'-heading' + tagger = soup.findAll('div', attrs={'id':att,'class':'heading'}) + if not tagger: + sectionNumber=sectionNumber+1 + continue + tag1=str(tagger[0].text) + tag1=tag1.strip() + print tag1 + if tag1=='Prerequisite S/W': + tag1='Prerequisites' + st+='
'+heading+'
'+s + s=s+f.read() + f.seek(t+13) + f.write(s) + f.seek(0) + content=f.read() + #print content + filedata=content.replace('index.html','Introduction.html?domain= Chemical Sciences&lab='+heading) + f.seek(0) + f.write(filedata) +f=open("template.html",'r') +template=f.read() +f=open("content.html",'r') +srchtml=f.read() +f.close() +soup = BeautifulSoup(srchtml, 'html.parser') +sectionno=soup.find_all('section') +d=['introduction','theory','objective','experiment','manual','quizzes','further_readings','procedure','hh','ll'] +print len(sectionno) +sectionNumber=1 +st="" +att = ''+'lab-header-heading' +tagger = soup.findAll('div', attrs={'id':att,'class':'heading'}) +heading=str(tagger[0].text) +print heading +heading=heading.strip() +print heading +breadcrumb=' Chemical Sciences' +while sectionNumber<=len(sectionno): + tag="" + att = ''+'lab-article-section-'+str(sectionNumber)+'-heading' + tagger = soup.findAll('div', attrs={'id':att,'class':'heading'}) + if not tagger: + sectionNumber=sectionNumber+1 + continue + tag1=str(tagger[0].text) + tag1=tag1.strip() + print tag1 + if tag1=='Prerequisite S/W': + tag1='Prerequisites' + st+='
'+tag1+'
' + sectionNumber=sectionNumber+1 +st+='Feedback
' +sectionNumber=1 +while sectionNumber<=len(sectionno): + tag="" + tag="" + att = ''+'lab-article-section-'+str(sectionNumber)+'-heading' + tagger = soup.findAll('div', attrs={'id':att,'class':'heading'}) + if not tagger: + sectionNumber=sectionNumber+1 + continue + tag1=str(tagger[0].text) + tag1=tag1.strip() + if tag1=='Prerequisite S/W': + tag1='Prerequisites' + att ='lab-article-section-'+str(sectionNumber)+'-content' + tagger = soup.findAll('div', attrs={'id':att,'class':'content'}) + + tag+=str(tagger[0]) + tag=tag.replace('','
')
+ #print tag
+ #print st
+
+ writefile(tag1+'.html',tag,st,heading)
+ #print sectionNumber
+ sectionNumber=sectionNumber+1
+f=open("Feedback.html",'w+')
+f.write(template)
+f.seek(0)
+
+content = f.read()
+content=content.replace('Disciplines and Domains',breadcrumb)
+f.seek(0)
+f.write(content)
+#print content
+f.seek(0)
+content=f.read()
+k=content.index('
')
+
+t1=content.index('')
+print t1
+f.seek(t1+13)
+st=st+f.read()
+#print s1
+f.seek(t1+13)
+f.write(st)
+f.seek(0)
+content=f.read()
+t= content.index('')
+f.seek(t+13)
+s='
'+heading+'
'+'Feedback' +s=s+f.read() +f.seek(t+13) +f.write(s) + diff --git a/src/lab/scraplab.py b/src/lab/scraplab.py new file mode 100644 index 0000000..289d9d9 --- /dev/null +++ b/src/lab/scraplab.py @@ -0,0 +1,124 @@ +from bs4 import BeautifulSoup +heading="" +breadcrumb="" +def writefile(fname,s,s1,heading): + f=open(fname, 'w+') + f.write(template) + f.seek(0) + + content = f.read() + content=content.replace('Disciplines and Domains',breadcrumb) + f.seek(0) + f.write(content) + print content + + f.seek(0) + content=f.read() + k=content.index('')
+
+ t1=content.index('')
+ print t1
+ f.seek(t1+13)
+ s1=s1+f.read()
+ #print s1
+ f.seek(t1+13)
+ f.write(s1)
+ f.seek(0)
+ content=f.read()
+ t= content.index('')
+ f.seek(t+13)
+ s='
' +while sectionNumber<=len(sectionno): + tag="" + att = ''+'lab-article-section-'+str(sectionNumber)+'-heading' + tagger = soup.findAll('div', attrs={'id':att,'class':'heading'}) + if not tagger: + sectionNumber=sectionNumber+1 + continue + tag1=str(tagger[0].text) + tag1=tag1.strip() + print tag1 + if tag1=='Prerequisite S/W': + tag1='Prerequisites' + st+='
'+heading+'
'+s + s=s+f.read() + f.seek(t+13) + f.write(s) +f=open("template1.html",'r') +template=f.read() +f=open("content.html",'r') +srchtml=f.read() +f.close() +soup = BeautifulSoup(srchtml, 'html.parser') +sectionno=soup.find_all('section') +d=['introduction','theory','objective','experiment','manual','quizzes','further_readings','procedure','hh','ll'] +print len(sectionno) +sectionNumber=1 +st="" + +att = ''+'lab-article-heading' +tagger = soup.findAll('div', attrs={'id':att,'class':'heading'}) +heading=str(tagger[0].text) +heading=heading.strip() +print heading +att = ''+'lab-header-heading' +tagger = soup.findAll('div', attrs={'id':att,'class':'heading'}) +heading1=str(tagger[0].text) +print heading +heading1=heading1.strip() +breadcrumb='Chemical Sciences →'+heading1+' →List Of Experiments' +while sectionNumber<=len(sectionno): + tag="" + att = ''+'lab-article-section-'+str(sectionNumber)+'-heading' + tagger = soup.findAll('div', attrs={'id':att,'class':'heading'}) + if not tagger: + sectionNumber=sectionNumber+1 + continue + tag1=str(tagger[0].text) + tag1=tag1.strip() + print tag1 + if tag1=='Prerequisite S/W': + tag1='Prerequisites' + st+='
'+tag1+'
' + sectionNumber=sectionNumber+1 +st+='Feedback
' +sectionNumber=1 +while sectionNumber<=len(sectionno): + tag="" + tag="" + att = ''+'lab-article-section-'+str(sectionNumber)+'-heading' + tagger = soup.findAll('div', attrs={'id':att,'class':'heading'}) + if not tagger: + sectionNumber=sectionNumber+1 + continue + tag1=str(tagger[0].text) + tag1=tag1.strip() + if tag1=='Prerequisite S/W': + tag1='Prerequisites' + att = ''+'lab-article-section-'+str(sectionNumber)+'-content' + tagger = soup.findAll('div', attrs={'id':att,'class':'content'}) + + tag+=str(tagger[0]) + #print tag + #print st + + + writefile(tag1+'.html',tag,st,heading) + #print sectionNumber + sectionNumber=sectionNumber+1 + +f=open("Feedback.html",'w+') +f.write(template) +f.seek(0) + +content = f.read() +content=content.replace('Disciplines and Domains',breadcrumb) +f.seek(0) +f.write(content) +#print content +f.seek(0) +content=f.read() +k=content.index('')
+
+t1=content.index('')
+print t1
+f.seek(t1+13)
+st=st+f.read()
+#print s1
+f.seek(t1+13)
+f.write(st)
+f.seek(0)
+content=f.read()
+t= content.index('')
+f.seek(t+13)
+s='Welcome to Virtual Labs - A MHRD Govt of india Initiative
+
+
+
+
+
+
+
+
+
+
+
+
+
+
+
+
+
+
+Welcome to Virtual Labs - A MHRD Govt of india Initiative
+
+
+
+
+
+
+
+
+
+
+
+
+
+
+
+
+
+
+
'+heading+'
'+'Feedback' +s=s+f.read() +f.seek(t+13) +f.write(s) diff --git a/src/lab/template.html b/src/lab/template.html new file mode 100644 index 0000000..c7d51f2 --- /dev/null +++ b/src/lab/template.html @@ -0,0 +1,205 @@ + + + + + + + + + + +
+
+
+
+
+
+
+
+
+
+
+
+
+
+
+
+
+
+
+
+
+
+
+
+
+
diff --git a/src/lab/template1.html b/src/lab/template1.html
new file mode 100644
index 0000000..382fb88
--- /dev/null
+++ b/src/lab/template1.html
@@ -0,0 +1,205 @@
+
+
+
+
+
+
+
+
+
+
+
+
+
+
+
+
+
+
+
+
+
+
+
+
+ +
+
+
+
+
+
+
+ Disciplines and Domains
+ +
+
+
+
+
+
+
+
+
+
+
+
+
+
+
+
+
+
+
+
+
+
+
+
+
+
+
+
+
+
+
+
+
+
diff --git a/vendors/font-awesome/css/font-awesome.css b/vendors/font-awesome/css/font-awesome.css
new file mode 100644
index 0000000..2dcdc22
--- /dev/null
+++ b/vendors/font-awesome/css/font-awesome.css
@@ -0,0 +1,1801 @@
+/*!
+ * Font Awesome 4.3.0 by @davegandy - http://fontawesome.io - @fontawesome
+ * License - http://fontawesome.io/license (Font: SIL OFL 1.1, CSS: MIT License)
+ */
+/* FONT PATH
+ * -------------------------- */
+@font-face {
+ font-family: 'FontAwesome';
+ src: url('../fonts/fontawesome-webfont.eot?v=4.3.0');
+ src: url('../fonts/fontawesome-webfont.eot?#iefix&v=4.3.0') format('embedded-opentype'), url('../fonts/fontawesome-webfont.woff2?v=4.3.0') format('woff2'), url('../fonts/fontawesome-webfont.woff?v=4.3.0') format('woff'), url('../fonts/fontawesome-webfont.ttf?v=4.3.0') format('truetype'), url('../fonts/fontawesome-webfont.svg?v=4.3.0#fontawesomeregular') format('svg');
+ font-weight: normal;
+ font-style: normal;
+}
+.fa {
+ display: inline-block;
+ font: normal normal normal 14px/1 FontAwesome;
+ font-size: inherit;
+ text-rendering: auto;
+ -webkit-font-smoothing: antialiased;
+ -moz-osx-font-smoothing: grayscale;
+ transform: translate(0, 0);
+}
+/* makes the font 33% larger relative to the icon container */
+.fa-lg {
+ font-size: 1.33333333em;
+ line-height: 0.75em;
+ vertical-align: -15%;
+}
+.fa-2x {
+ font-size: 2em;
+}
+.fa-3x {
+ font-size: 3em;
+}
+.fa-4x {
+ font-size: 4em;
+}
+.fa-5x {
+ font-size: 5em;
+}
+.fa-fw {
+ width: 1.28571429em;
+ text-align: center;
+}
+.fa-ul {
+ padding-left: 0;
+ margin-left: 2.14285714em;
+ list-style-type: none;
+}
+.fa-ul > li {
+ position: relative;
+}
+.fa-li {
+ position: absolute;
+ left: -2.14285714em;
+ width: 2.14285714em;
+ top: 0.14285714em;
+ text-align: center;
+}
+.fa-li.fa-lg {
+ left: -1.85714286em;
+}
+.fa-border {
+ padding: .2em .25em .15em;
+ border: solid 0.08em #eeeeee;
+ border-radius: .1em;
+}
+.pull-right {
+ float: right;
+}
+.pull-left {
+ float: left;
+}
+.fa.pull-left {
+ margin-right: .3em;
+}
+.fa.pull-right {
+ margin-left: .3em;
+}
+.fa-spin {
+ -webkit-animation: fa-spin 2s infinite linear;
+ animation: fa-spin 2s infinite linear;
+}
+.fa-pulse {
+ -webkit-animation: fa-spin 1s infinite steps(8);
+ animation: fa-spin 1s infinite steps(8);
+}
+@-webkit-keyframes fa-spin {
+ 0% {
+ -webkit-transform: rotate(0deg);
+ transform: rotate(0deg);
+ }
+ 100% {
+ -webkit-transform: rotate(359deg);
+ transform: rotate(359deg);
+ }
+}
+@keyframes fa-spin {
+ 0% {
+ -webkit-transform: rotate(0deg);
+ transform: rotate(0deg);
+ }
+ 100% {
+ -webkit-transform: rotate(359deg);
+ transform: rotate(359deg);
+ }
+}
+.fa-rotate-90 {
+ filter: progid:DXImageTransform.Microsoft.BasicImage(rotation=1);
+ -webkit-transform: rotate(90deg);
+ -ms-transform: rotate(90deg);
+ transform: rotate(90deg);
+}
+.fa-rotate-180 {
+ filter: progid:DXImageTransform.Microsoft.BasicImage(rotation=2);
+ -webkit-transform: rotate(180deg);
+ -ms-transform: rotate(180deg);
+ transform: rotate(180deg);
+}
+.fa-rotate-270 {
+ filter: progid:DXImageTransform.Microsoft.BasicImage(rotation=3);
+ -webkit-transform: rotate(270deg);
+ -ms-transform: rotate(270deg);
+ transform: rotate(270deg);
+}
+.fa-flip-horizontal {
+ filter: progid:DXImageTransform.Microsoft.BasicImage(rotation=0, mirror=1);
+ -webkit-transform: scale(-1, 1);
+ -ms-transform: scale(-1, 1);
+ transform: scale(-1, 1);
+}
+.fa-flip-vertical {
+ filter: progid:DXImageTransform.Microsoft.BasicImage(rotation=2, mirror=1);
+ -webkit-transform: scale(1, -1);
+ -ms-transform: scale(1, -1);
+ transform: scale(1, -1);
+}
+:root .fa-rotate-90,
+:root .fa-rotate-180,
+:root .fa-rotate-270,
+:root .fa-flip-horizontal,
+:root .fa-flip-vertical {
+ filter: none;
+}
+.fa-stack {
+ position: relative;
+ display: inline-block;
+ width: 2em;
+ height: 2em;
+ line-height: 2em;
+ vertical-align: middle;
+}
+.fa-stack-1x,
+.fa-stack-2x {
+ position: absolute;
+ left: 0;
+ width: 100%;
+ text-align: center;
+}
+.fa-stack-1x {
+ line-height: inherit;
+}
+.fa-stack-2x {
+ font-size: 2em;
+}
+.fa-inverse {
+ color: #ffffff;
+}
+/* Font Awesome uses the Unicode Private Use Area (PUA) to ensure screen
+ readers do not read off random characters that represent icons */
+.fa-glass:before {
+ content: "\f000";
+}
+.fa-music:before {
+ content: "\f001";
+}
+.fa-search:before {
+ content: "\f002";
+}
+.fa-envelope-o:before {
+ content: "\f003";
+}
+.fa-heart:before {
+ content: "\f004";
+}
+.fa-star:before {
+ content: "\f005";
+}
+.fa-star-o:before {
+ content: "\f006";
+}
+.fa-user:before {
+ content: "\f007";
+}
+.fa-film:before {
+ content: "\f008";
+}
+.fa-th-large:before {
+ content: "\f009";
+}
+.fa-th:before {
+ content: "\f00a";
+}
+.fa-th-list:before {
+ content: "\f00b";
+}
+.fa-check:before {
+ content: "\f00c";
+}
+.fa-remove:before,
+.fa-close:before,
+.fa-times:before {
+ content: "\f00d";
+}
+.fa-search-plus:before {
+ content: "\f00e";
+}
+.fa-search-minus:before {
+ content: "\f010";
+}
+.fa-power-off:before {
+ content: "\f011";
+}
+.fa-signal:before {
+ content: "\f012";
+}
+.fa-gear:before,
+.fa-cog:before {
+ content: "\f013";
+}
+.fa-trash-o:before {
+ content: "\f014";
+}
+.fa-home:before {
+ content: "\f015";
+}
+.fa-file-o:before {
+ content: "\f016";
+}
+.fa-clock-o:before {
+ content: "\f017";
+}
+.fa-road:before {
+ content: "\f018";
+}
+.fa-download:before {
+ content: "\f019";
+}
+.fa-arrow-circle-o-down:before {
+ content: "\f01a";
+}
+.fa-arrow-circle-o-up:before {
+ content: "\f01b";
+}
+.fa-inbox:before {
+ content: "\f01c";
+}
+.fa-play-circle-o:before {
+ content: "\f01d";
+}
+.fa-rotate-right:before,
+.fa-repeat:before {
+ content: "\f01e";
+}
+.fa-refresh:before {
+ content: "\f021";
+}
+.fa-list-alt:before {
+ content: "\f022";
+}
+.fa-lock:before {
+ content: "\f023";
+}
+.fa-flag:before {
+ content: "\f024";
+}
+.fa-headphones:before {
+ content: "\f025";
+}
+.fa-volume-off:before {
+ content: "\f026";
+}
+.fa-volume-down:before {
+ content: "\f027";
+}
+.fa-volume-up:before {
+ content: "\f028";
+}
+.fa-qrcode:before {
+ content: "\f029";
+}
+.fa-barcode:before {
+ content: "\f02a";
+}
+.fa-tag:before {
+ content: "\f02b";
+}
+.fa-tags:before {
+ content: "\f02c";
+}
+.fa-book:before {
+ content: "\f02d";
+}
+.fa-bookmark:before {
+ content: "\f02e";
+}
+.fa-print:before {
+ content: "\f02f";
+}
+.fa-camera:before {
+ content: "\f030";
+}
+.fa-font:before {
+ content: "\f031";
+}
+.fa-bold:before {
+ content: "\f032";
+}
+.fa-italic:before {
+ content: "\f033";
+}
+.fa-text-height:before {
+ content: "\f034";
+}
+.fa-text-width:before {
+ content: "\f035";
+}
+.fa-align-left:before {
+ content: "\f036";
+}
+.fa-align-center:before {
+ content: "\f037";
+}
+.fa-align-right:before {
+ content: "\f038";
+}
+.fa-align-justify:before {
+ content: "\f039";
+}
+.fa-list:before {
+ content: "\f03a";
+}
+.fa-dedent:before,
+.fa-outdent:before {
+ content: "\f03b";
+}
+.fa-indent:before {
+ content: "\f03c";
+}
+.fa-video-camera:before {
+ content: "\f03d";
+}
+.fa-photo:before,
+.fa-image:before,
+.fa-picture-o:before {
+ content: "\f03e";
+}
+.fa-pencil:before {
+ content: "\f040";
+}
+.fa-map-marker:before {
+ content: "\f041";
+}
+.fa-adjust:before {
+ content: "\f042";
+}
+.fa-tint:before {
+ content: "\f043";
+}
+.fa-edit:before,
+.fa-pencil-square-o:before {
+ content: "\f044";
+}
+.fa-share-square-o:before {
+ content: "\f045";
+}
+.fa-check-square-o:before {
+ content: "\f046";
+}
+.fa-arrows:before {
+ content: "\f047";
+}
+.fa-step-backward:before {
+ content: "\f048";
+}
+.fa-fast-backward:before {
+ content: "\f049";
+}
+.fa-backward:before {
+ content: "\f04a";
+}
+.fa-play:before {
+ content: "\f04b";
+}
+.fa-pause:before {
+ content: "\f04c";
+}
+.fa-stop:before {
+ content: "\f04d";
+}
+.fa-forward:before {
+ content: "\f04e";
+}
+.fa-fast-forward:before {
+ content: "\f050";
+}
+.fa-step-forward:before {
+ content: "\f051";
+}
+.fa-eject:before {
+ content: "\f052";
+}
+.fa-chevron-left:before {
+ content: "\f053";
+}
+.fa-chevron-right:before {
+ content: "\f054";
+}
+.fa-plus-circle:before {
+ content: "\f055";
+}
+.fa-minus-circle:before {
+ content: "\f056";
+}
+.fa-times-circle:before {
+ content: "\f057";
+}
+.fa-check-circle:before {
+ content: "\f058";
+}
+.fa-question-circle:before {
+ content: "\f059";
+}
+.fa-info-circle:before {
+ content: "\f05a";
+}
+.fa-crosshairs:before {
+ content: "\f05b";
+}
+.fa-times-circle-o:before {
+ content: "\f05c";
+}
+.fa-check-circle-o:before {
+ content: "\f05d";
+}
+.fa-ban:before {
+ content: "\f05e";
+}
+.fa-arrow-left:before {
+ content: "\f060";
+}
+.fa-arrow-right:before {
+ content: "\f061";
+}
+.fa-arrow-up:before {
+ content: "\f062";
+}
+.fa-arrow-down:before {
+ content: "\f063";
+}
+.fa-mail-forward:before,
+.fa-share:before {
+ content: "\f064";
+}
+.fa-expand:before {
+ content: "\f065";
+}
+.fa-compress:before {
+ content: "\f066";
+}
+.fa-plus:before {
+ content: "\f067";
+}
+.fa-minus:before {
+ content: "\f068";
+}
+.fa-asterisk:before {
+ content: "\f069";
+}
+.fa-exclamation-circle:before {
+ content: "\f06a";
+}
+.fa-gift:before {
+ content: "\f06b";
+}
+.fa-leaf:before {
+ content: "\f06c";
+}
+.fa-fire:before {
+ content: "\f06d";
+}
+.fa-eye:before {
+ content: "\f06e";
+}
+.fa-eye-slash:before {
+ content: "\f070";
+}
+.fa-warning:before,
+.fa-exclamation-triangle:before {
+ content: "\f071";
+}
+.fa-plane:before {
+ content: "\f072";
+}
+.fa-calendar:before {
+ content: "\f073";
+}
+.fa-random:before {
+ content: "\f074";
+}
+.fa-comment:before {
+ content: "\f075";
+}
+.fa-magnet:before {
+ content: "\f076";
+}
+.fa-chevron-up:before {
+ content: "\f077";
+}
+.fa-chevron-down:before {
+ content: "\f078";
+}
+.fa-retweet:before {
+ content: "\f079";
+}
+.fa-shopping-cart:before {
+ content: "\f07a";
+}
+.fa-folder:before {
+ content: "\f07b";
+}
+.fa-folder-open:before {
+ content: "\f07c";
+}
+.fa-arrows-v:before {
+ content: "\f07d";
+}
+.fa-arrows-h:before {
+ content: "\f07e";
+}
+.fa-bar-chart-o:before,
+.fa-bar-chart:before {
+ content: "\f080";
+}
+.fa-twitter-square:before {
+ content: "\f081";
+}
+.fa-facebook-square:before {
+ content: "\f082";
+}
+.fa-camera-retro:before {
+ content: "\f083";
+}
+.fa-key:before {
+ content: "\f084";
+}
+.fa-gears:before,
+.fa-cogs:before {
+ content: "\f085";
+}
+.fa-comments:before {
+ content: "\f086";
+}
+.fa-thumbs-o-up:before {
+ content: "\f087";
+}
+.fa-thumbs-o-down:before {
+ content: "\f088";
+}
+.fa-star-half:before {
+ content: "\f089";
+}
+.fa-heart-o:before {
+ content: "\f08a";
+}
+.fa-sign-out:before {
+ content: "\f08b";
+}
+.fa-linkedin-square:before {
+ content: "\f08c";
+}
+.fa-thumb-tack:before {
+ content: "\f08d";
+}
+.fa-external-link:before {
+ content: "\f08e";
+}
+.fa-sign-in:before {
+ content: "\f090";
+}
+.fa-trophy:before {
+ content: "\f091";
+}
+.fa-github-square:before {
+ content: "\f092";
+}
+.fa-upload:before {
+ content: "\f093";
+}
+.fa-lemon-o:before {
+ content: "\f094";
+}
+.fa-phone:before {
+ content: "\f095";
+}
+.fa-square-o:before {
+ content: "\f096";
+}
+.fa-bookmark-o:before {
+ content: "\f097";
+}
+.fa-phone-square:before {
+ content: "\f098";
+}
+.fa-twitter:before {
+ content: "\f099";
+}
+.fa-facebook-f:before,
+.fa-facebook:before {
+ content: "\f09a";
+}
+.fa-github:before {
+ content: "\f09b";
+}
+.fa-unlock:before {
+ content: "\f09c";
+}
+.fa-credit-card:before {
+ content: "\f09d";
+}
+.fa-rss:before {
+ content: "\f09e";
+}
+.fa-hdd-o:before {
+ content: "\f0a0";
+}
+.fa-bullhorn:before {
+ content: "\f0a1";
+}
+.fa-bell:before {
+ content: "\f0f3";
+}
+.fa-certificate:before {
+ content: "\f0a3";
+}
+.fa-hand-o-right:before {
+ content: "\f0a4";
+}
+.fa-hand-o-left:before {
+ content: "\f0a5";
+}
+.fa-hand-o-up:before {
+ content: "\f0a6";
+}
+.fa-hand-o-down:before {
+ content: "\f0a7";
+}
+.fa-arrow-circle-left:before {
+ content: "\f0a8";
+}
+.fa-arrow-circle-right:before {
+ content: "\f0a9";
+}
+.fa-arrow-circle-up:before {
+ content: "\f0aa";
+}
+.fa-arrow-circle-down:before {
+ content: "\f0ab";
+}
+.fa-globe:before {
+ content: "\f0ac";
+}
+.fa-wrench:before {
+ content: "\f0ad";
+}
+.fa-tasks:before {
+ content: "\f0ae";
+}
+.fa-filter:before {
+ content: "\f0b0";
+}
+.fa-briefcase:before {
+ content: "\f0b1";
+}
+.fa-arrows-alt:before {
+ content: "\f0b2";
+}
+.fa-group:before,
+.fa-users:before {
+ content: "\f0c0";
+}
+.fa-chain:before,
+.fa-link:before {
+ content: "\f0c1";
+}
+.fa-cloud:before {
+ content: "\f0c2";
+}
+.fa-flask:before {
+ content: "\f0c3";
+}
+.fa-cut:before,
+.fa-scissors:before {
+ content: "\f0c4";
+}
+.fa-copy:before,
+.fa-files-o:before {
+ content: "\f0c5";
+}
+.fa-paperclip:before {
+ content: "\f0c6";
+}
+.fa-save:before,
+.fa-floppy-o:before {
+ content: "\f0c7";
+}
+.fa-square:before {
+ content: "\f0c8";
+}
+.fa-navicon:before,
+.fa-reorder:before,
+.fa-bars:before {
+ content: "\f0c9";
+}
+.fa-list-ul:before {
+ content: "\f0ca";
+}
+.fa-list-ol:before {
+ content: "\f0cb";
+}
+.fa-strikethrough:before {
+ content: "\f0cc";
+}
+.fa-underline:before {
+ content: "\f0cd";
+}
+.fa-table:before {
+ content: "\f0ce";
+}
+.fa-magic:before {
+ content: "\f0d0";
+}
+.fa-truck:before {
+ content: "\f0d1";
+}
+.fa-pinterest:before {
+ content: "\f0d2";
+}
+.fa-pinterest-square:before {
+ content: "\f0d3";
+}
+.fa-google-plus-square:before {
+ content: "\f0d4";
+}
+.fa-google-plus:before {
+ content: "\f0d5";
+}
+.fa-money:before {
+ content: "\f0d6";
+}
+.fa-caret-down:before {
+ content: "\f0d7";
+}
+.fa-caret-up:before {
+ content: "\f0d8";
+}
+.fa-caret-left:before {
+ content: "\f0d9";
+}
+.fa-caret-right:before {
+ content: "\f0da";
+}
+.fa-columns:before {
+ content: "\f0db";
+}
+.fa-unsorted:before,
+.fa-sort:before {
+ content: "\f0dc";
+}
+.fa-sort-down:before,
+.fa-sort-desc:before {
+ content: "\f0dd";
+}
+.fa-sort-up:before,
+.fa-sort-asc:before {
+ content: "\f0de";
+}
+.fa-envelope:before {
+ content: "\f0e0";
+}
+.fa-linkedin:before {
+ content: "\f0e1";
+}
+.fa-rotate-left:before,
+.fa-undo:before {
+ content: "\f0e2";
+}
+.fa-legal:before,
+.fa-gavel:before {
+ content: "\f0e3";
+}
+.fa-dashboard:before,
+.fa-tachometer:before {
+ content: "\f0e4";
+}
+.fa-comment-o:before {
+ content: "\f0e5";
+}
+.fa-comments-o:before {
+ content: "\f0e6";
+}
+.fa-flash:before,
+.fa-bolt:before {
+ content: "\f0e7";
+}
+.fa-sitemap:before {
+ content: "\f0e8";
+}
+.fa-umbrella:before {
+ content: "\f0e9";
+}
+.fa-paste:before,
+.fa-clipboard:before {
+ content: "\f0ea";
+}
+.fa-lightbulb-o:before {
+ content: "\f0eb";
+}
+.fa-exchange:before {
+ content: "\f0ec";
+}
+.fa-cloud-download:before {
+ content: "\f0ed";
+}
+.fa-cloud-upload:before {
+ content: "\f0ee";
+}
+.fa-user-md:before {
+ content: "\f0f0";
+}
+.fa-stethoscope:before {
+ content: "\f0f1";
+}
+.fa-suitcase:before {
+ content: "\f0f2";
+}
+.fa-bell-o:before {
+ content: "\f0a2";
+}
+.fa-coffee:before {
+ content: "\f0f4";
+}
+.fa-cutlery:before {
+ content: "\f0f5";
+}
+.fa-file-text-o:before {
+ content: "\f0f6";
+}
+.fa-building-o:before {
+ content: "\f0f7";
+}
+.fa-hospital-o:before {
+ content: "\f0f8";
+}
+.fa-ambulance:before {
+ content: "\f0f9";
+}
+.fa-medkit:before {
+ content: "\f0fa";
+}
+.fa-fighter-jet:before {
+ content: "\f0fb";
+}
+.fa-beer:before {
+ content: "\f0fc";
+}
+.fa-h-square:before {
+ content: "\f0fd";
+}
+.fa-plus-square:before {
+ content: "\f0fe";
+}
+.fa-angle-double-left:before {
+ content: "\f100";
+}
+.fa-angle-double-right:before {
+ content: "\f101";
+}
+.fa-angle-double-up:before {
+ content: "\f102";
+}
+.fa-angle-double-down:before {
+ content: "\f103";
+}
+.fa-angle-left:before {
+ content: "\f104";
+}
+.fa-angle-right:before {
+ content: "\f105";
+}
+.fa-angle-up:before {
+ content: "\f106";
+}
+.fa-angle-down:before {
+ content: "\f107";
+}
+.fa-desktop:before {
+ content: "\f108";
+}
+.fa-laptop:before {
+ content: "\f109";
+}
+.fa-tablet:before {
+ content: "\f10a";
+}
+.fa-mobile-phone:before,
+.fa-mobile:before {
+ content: "\f10b";
+}
+.fa-circle-o:before {
+ content: "\f10c";
+}
+.fa-quote-left:before {
+ content: "\f10d";
+}
+.fa-quote-right:before {
+ content: "\f10e";
+}
+.fa-spinner:before {
+ content: "\f110";
+}
+.fa-circle:before {
+ content: "\f111";
+}
+.fa-mail-reply:before,
+.fa-reply:before {
+ content: "\f112";
+}
+.fa-github-alt:before {
+ content: "\f113";
+}
+.fa-folder-o:before {
+ content: "\f114";
+}
+.fa-folder-open-o:before {
+ content: "\f115";
+}
+.fa-smile-o:before {
+ content: "\f118";
+}
+.fa-frown-o:before {
+ content: "\f119";
+}
+.fa-meh-o:before {
+ content: "\f11a";
+}
+.fa-gamepad:before {
+ content: "\f11b";
+}
+.fa-keyboard-o:before {
+ content: "\f11c";
+}
+.fa-flag-o:before {
+ content: "\f11d";
+}
+.fa-flag-checkered:before {
+ content: "\f11e";
+}
+.fa-terminal:before {
+ content: "\f120";
+}
+.fa-code:before {
+ content: "\f121";
+}
+.fa-mail-reply-all:before,
+.fa-reply-all:before {
+ content: "\f122";
+}
+.fa-star-half-empty:before,
+.fa-star-half-full:before,
+.fa-star-half-o:before {
+ content: "\f123";
+}
+.fa-location-arrow:before {
+ content: "\f124";
+}
+.fa-crop:before {
+ content: "\f125";
+}
+.fa-code-fork:before {
+ content: "\f126";
+}
+.fa-unlink:before,
+.fa-chain-broken:before {
+ content: "\f127";
+}
+.fa-question:before {
+ content: "\f128";
+}
+.fa-info:before {
+ content: "\f129";
+}
+.fa-exclamation:before {
+ content: "\f12a";
+}
+.fa-superscript:before {
+ content: "\f12b";
+}
+.fa-subscript:before {
+ content: "\f12c";
+}
+.fa-eraser:before {
+ content: "\f12d";
+}
+.fa-puzzle-piece:before {
+ content: "\f12e";
+}
+.fa-microphone:before {
+ content: "\f130";
+}
+.fa-microphone-slash:before {
+ content: "\f131";
+}
+.fa-shield:before {
+ content: "\f132";
+}
+.fa-calendar-o:before {
+ content: "\f133";
+}
+.fa-fire-extinguisher:before {
+ content: "\f134";
+}
+.fa-rocket:before {
+ content: "\f135";
+}
+.fa-maxcdn:before {
+ content: "\f136";
+}
+.fa-chevron-circle-left:before {
+ content: "\f137";
+}
+.fa-chevron-circle-right:before {
+ content: "\f138";
+}
+.fa-chevron-circle-up:before {
+ content: "\f139";
+}
+.fa-chevron-circle-down:before {
+ content: "\f13a";
+}
+.fa-html5:before {
+ content: "\f13b";
+}
+.fa-css3:before {
+ content: "\f13c";
+}
+.fa-anchor:before {
+ content: "\f13d";
+}
+.fa-unlock-alt:before {
+ content: "\f13e";
+}
+.fa-bullseye:before {
+ content: "\f140";
+}
+.fa-ellipsis-h:before {
+ content: "\f141";
+}
+.fa-ellipsis-v:before {
+ content: "\f142";
+}
+.fa-rss-square:before {
+ content: "\f143";
+}
+.fa-play-circle:before {
+ content: "\f144";
+}
+.fa-ticket:before {
+ content: "\f145";
+}
+.fa-minus-square:before {
+ content: "\f146";
+}
+.fa-minus-square-o:before {
+ content: "\f147";
+}
+.fa-level-up:before {
+ content: "\f148";
+}
+.fa-level-down:before {
+ content: "\f149";
+}
+.fa-check-square:before {
+ content: "\f14a";
+}
+.fa-pencil-square:before {
+ content: "\f14b";
+}
+.fa-external-link-square:before {
+ content: "\f14c";
+}
+.fa-share-square:before {
+ content: "\f14d";
+}
+.fa-compass:before {
+ content: "\f14e";
+}
+.fa-toggle-down:before,
+.fa-caret-square-o-down:before {
+ content: "\f150";
+}
+.fa-toggle-up:before,
+.fa-caret-square-o-up:before {
+ content: "\f151";
+}
+.fa-toggle-right:before,
+.fa-caret-square-o-right:before {
+ content: "\f152";
+}
+.fa-euro:before,
+.fa-eur:before {
+ content: "\f153";
+}
+.fa-gbp:before {
+ content: "\f154";
+}
+.fa-dollar:before,
+.fa-usd:before {
+ content: "\f155";
+}
+.fa-rupee:before,
+.fa-inr:before {
+ content: "\f156";
+}
+.fa-cny:before,
+.fa-rmb:before,
+.fa-yen:before,
+.fa-jpy:before {
+ content: "\f157";
+}
+.fa-ruble:before,
+.fa-rouble:before,
+.fa-rub:before {
+ content: "\f158";
+}
+.fa-won:before,
+.fa-krw:before {
+ content: "\f159";
+}
+.fa-bitcoin:before,
+.fa-btc:before {
+ content: "\f15a";
+}
+.fa-file:before {
+ content: "\f15b";
+}
+.fa-file-text:before {
+ content: "\f15c";
+}
+.fa-sort-alpha-asc:before {
+ content: "\f15d";
+}
+.fa-sort-alpha-desc:before {
+ content: "\f15e";
+}
+.fa-sort-amount-asc:before {
+ content: "\f160";
+}
+.fa-sort-amount-desc:before {
+ content: "\f161";
+}
+.fa-sort-numeric-asc:before {
+ content: "\f162";
+}
+.fa-sort-numeric-desc:before {
+ content: "\f163";
+}
+.fa-thumbs-up:before {
+ content: "\f164";
+}
+.fa-thumbs-down:before {
+ content: "\f165";
+}
+.fa-youtube-square:before {
+ content: "\f166";
+}
+.fa-youtube:before {
+ content: "\f167";
+}
+.fa-xing:before {
+ content: "\f168";
+}
+.fa-xing-square:before {
+ content: "\f169";
+}
+.fa-youtube-play:before {
+ content: "\f16a";
+}
+.fa-dropbox:before {
+ content: "\f16b";
+}
+.fa-stack-overflow:before {
+ content: "\f16c";
+}
+.fa-instagram:before {
+ content: "\f16d";
+}
+.fa-flickr:before {
+ content: "\f16e";
+}
+.fa-adn:before {
+ content: "\f170";
+}
+.fa-bitbucket:before {
+ content: "\f171";
+}
+.fa-bitbucket-square:before {
+ content: "\f172";
+}
+.fa-tumblr:before {
+ content: "\f173";
+}
+.fa-tumblr-square:before {
+ content: "\f174";
+}
+.fa-long-arrow-down:before {
+ content: "\f175";
+}
+.fa-long-arrow-up:before {
+ content: "\f176";
+}
+.fa-long-arrow-left:before {
+ content: "\f177";
+}
+.fa-long-arrow-right:before {
+ content: "\f178";
+}
+.fa-apple:before {
+ content: "\f179";
+}
+.fa-windows:before {
+ content: "\f17a";
+}
+.fa-android:before {
+ content: "\f17b";
+}
+.fa-linux:before {
+ content: "\f17c";
+}
+.fa-dribbble:before {
+ content: "\f17d";
+}
+.fa-skype:before {
+ content: "\f17e";
+}
+.fa-foursquare:before {
+ content: "\f180";
+}
+.fa-trello:before {
+ content: "\f181";
+}
+.fa-female:before {
+ content: "\f182";
+}
+.fa-male:before {
+ content: "\f183";
+}
+.fa-gittip:before,
+.fa-gratipay:before {
+ content: "\f184";
+}
+.fa-sun-o:before {
+ content: "\f185";
+}
+.fa-moon-o:before {
+ content: "\f186";
+}
+.fa-archive:before {
+ content: "\f187";
+}
+.fa-bug:before {
+ content: "\f188";
+}
+.fa-vk:before {
+ content: "\f189";
+}
+.fa-weibo:before {
+ content: "\f18a";
+}
+.fa-renren:before {
+ content: "\f18b";
+}
+.fa-pagelines:before {
+ content: "\f18c";
+}
+.fa-stack-exchange:before {
+ content: "\f18d";
+}
+.fa-arrow-circle-o-right:before {
+ content: "\f18e";
+}
+.fa-arrow-circle-o-left:before {
+ content: "\f190";
+}
+.fa-toggle-left:before,
+.fa-caret-square-o-left:before {
+ content: "\f191";
+}
+.fa-dot-circle-o:before {
+ content: "\f192";
+}
+.fa-wheelchair:before {
+ content: "\f193";
+}
+.fa-vimeo-square:before {
+ content: "\f194";
+}
+.fa-turkish-lira:before,
+.fa-try:before {
+ content: "\f195";
+}
+.fa-plus-square-o:before {
+ content: "\f196";
+}
+.fa-space-shuttle:before {
+ content: "\f197";
+}
+.fa-slack:before {
+ content: "\f198";
+}
+.fa-envelope-square:before {
+ content: "\f199";
+}
+.fa-wordpress:before {
+ content: "\f19a";
+}
+.fa-openid:before {
+ content: "\f19b";
+}
+.fa-institution:before,
+.fa-bank:before,
+.fa-university:before {
+ content: "\f19c";
+}
+.fa-mortar-board:before,
+.fa-graduation-cap:before {
+ content: "\f19d";
+}
+.fa-yahoo:before {
+ content: "\f19e";
+}
+.fa-google:before {
+ content: "\f1a0";
+}
+.fa-reddit:before {
+ content: "\f1a1";
+}
+.fa-reddit-square:before {
+ content: "\f1a2";
+}
+.fa-stumbleupon-circle:before {
+ content: "\f1a3";
+}
+.fa-stumbleupon:before {
+ content: "\f1a4";
+}
+.fa-delicious:before {
+ content: "\f1a5";
+}
+.fa-digg:before {
+ content: "\f1a6";
+}
+.fa-pied-piper:before {
+ content: "\f1a7";
+}
+.fa-pied-piper-alt:before {
+ content: "\f1a8";
+}
+.fa-drupal:before {
+ content: "\f1a9";
+}
+.fa-joomla:before {
+ content: "\f1aa";
+}
+.fa-language:before {
+ content: "\f1ab";
+}
+.fa-fax:before {
+ content: "\f1ac";
+}
+.fa-building:before {
+ content: "\f1ad";
+}
+.fa-child:before {
+ content: "\f1ae";
+}
+.fa-paw:before {
+ content: "\f1b0";
+}
+.fa-spoon:before {
+ content: "\f1b1";
+}
+.fa-cube:before {
+ content: "\f1b2";
+}
+.fa-cubes:before {
+ content: "\f1b3";
+}
+.fa-behance:before {
+ content: "\f1b4";
+}
+.fa-behance-square:before {
+ content: "\f1b5";
+}
+.fa-steam:before {
+ content: "\f1b6";
+}
+.fa-steam-square:before {
+ content: "\f1b7";
+}
+.fa-recycle:before {
+ content: "\f1b8";
+}
+.fa-automobile:before,
+.fa-car:before {
+ content: "\f1b9";
+}
+.fa-cab:before,
+.fa-taxi:before {
+ content: "\f1ba";
+}
+.fa-tree:before {
+ content: "\f1bb";
+}
+.fa-spotify:before {
+ content: "\f1bc";
+}
+.fa-deviantart:before {
+ content: "\f1bd";
+}
+.fa-soundcloud:before {
+ content: "\f1be";
+}
+.fa-database:before {
+ content: "\f1c0";
+}
+.fa-file-pdf-o:before {
+ content: "\f1c1";
+}
+.fa-file-word-o:before {
+ content: "\f1c2";
+}
+.fa-file-excel-o:before {
+ content: "\f1c3";
+}
+.fa-file-powerpoint-o:before {
+ content: "\f1c4";
+}
+.fa-file-photo-o:before,
+.fa-file-picture-o:before,
+.fa-file-image-o:before {
+ content: "\f1c5";
+}
+.fa-file-zip-o:before,
+.fa-file-archive-o:before {
+ content: "\f1c6";
+}
+.fa-file-sound-o:before,
+.fa-file-audio-o:before {
+ content: "\f1c7";
+}
+.fa-file-movie-o:before,
+.fa-file-video-o:before {
+ content: "\f1c8";
+}
+.fa-file-code-o:before {
+ content: "\f1c9";
+}
+.fa-vine:before {
+ content: "\f1ca";
+}
+.fa-codepen:before {
+ content: "\f1cb";
+}
+.fa-jsfiddle:before {
+ content: "\f1cc";
+}
+.fa-life-bouy:before,
+.fa-life-buoy:before,
+.fa-life-saver:before,
+.fa-support:before,
+.fa-life-ring:before {
+ content: "\f1cd";
+}
+.fa-circle-o-notch:before {
+ content: "\f1ce";
+}
+.fa-ra:before,
+.fa-rebel:before {
+ content: "\f1d0";
+}
+.fa-ge:before,
+.fa-empire:before {
+ content: "\f1d1";
+}
+.fa-git-square:before {
+ content: "\f1d2";
+}
+.fa-git:before {
+ content: "\f1d3";
+}
+.fa-hacker-news:before {
+ content: "\f1d4";
+}
+.fa-tencent-weibo:before {
+ content: "\f1d5";
+}
+.fa-qq:before {
+ content: "\f1d6";
+}
+.fa-wechat:before,
+.fa-weixin:before {
+ content: "\f1d7";
+}
+.fa-send:before,
+.fa-paper-plane:before {
+ content: "\f1d8";
+}
+.fa-send-o:before,
+.fa-paper-plane-o:before {
+ content: "\f1d9";
+}
+.fa-history:before {
+ content: "\f1da";
+}
+.fa-genderless:before,
+.fa-circle-thin:before {
+ content: "\f1db";
+}
+.fa-header:before {
+ content: "\f1dc";
+}
+.fa-paragraph:before {
+ content: "\f1dd";
+}
+.fa-sliders:before {
+ content: "\f1de";
+}
+.fa-share-alt:before {
+ content: "\f1e0";
+}
+.fa-share-alt-square:before {
+ content: "\f1e1";
+}
+.fa-bomb:before {
+ content: "\f1e2";
+}
+.fa-soccer-ball-o:before,
+.fa-futbol-o:before {
+ content: "\f1e3";
+}
+.fa-tty:before {
+ content: "\f1e4";
+}
+.fa-binoculars:before {
+ content: "\f1e5";
+}
+.fa-plug:before {
+ content: "\f1e6";
+}
+.fa-slideshare:before {
+ content: "\f1e7";
+}
+.fa-twitch:before {
+ content: "\f1e8";
+}
+.fa-yelp:before {
+ content: "\f1e9";
+}
+.fa-newspaper-o:before {
+ content: "\f1ea";
+}
+.fa-wifi:before {
+ content: "\f1eb";
+}
+.fa-calculator:before {
+ content: "\f1ec";
+}
+.fa-paypal:before {
+ content: "\f1ed";
+}
+.fa-google-wallet:before {
+ content: "\f1ee";
+}
+.fa-cc-visa:before {
+ content: "\f1f0";
+}
+.fa-cc-mastercard:before {
+ content: "\f1f1";
+}
+.fa-cc-discover:before {
+ content: "\f1f2";
+}
+.fa-cc-amex:before {
+ content: "\f1f3";
+}
+.fa-cc-paypal:before {
+ content: "\f1f4";
+}
+.fa-cc-stripe:before {
+ content: "\f1f5";
+}
+.fa-bell-slash:before {
+ content: "\f1f6";
+}
+.fa-bell-slash-o:before {
+ content: "\f1f7";
+}
+.fa-trash:before {
+ content: "\f1f8";
+}
+.fa-copyright:before {
+ content: "\f1f9";
+}
+.fa-at:before {
+ content: "\f1fa";
+}
+.fa-eyedropper:before {
+ content: "\f1fb";
+}
+.fa-paint-brush:before {
+ content: "\f1fc";
+}
+.fa-birthday-cake:before {
+ content: "\f1fd";
+}
+.fa-area-chart:before {
+ content: "\f1fe";
+}
+.fa-pie-chart:before {
+ content: "\f200";
+}
+.fa-line-chart:before {
+ content: "\f201";
+}
+.fa-lastfm:before {
+ content: "\f202";
+}
+.fa-lastfm-square:before {
+ content: "\f203";
+}
+.fa-toggle-off:before {
+ content: "\f204";
+}
+.fa-toggle-on:before {
+ content: "\f205";
+}
+.fa-bicycle:before {
+ content: "\f206";
+}
+.fa-bus:before {
+ content: "\f207";
+}
+.fa-ioxhost:before {
+ content: "\f208";
+}
+.fa-angellist:before {
+ content: "\f209";
+}
+.fa-cc:before {
+ content: "\f20a";
+}
+.fa-shekel:before,
+.fa-sheqel:before,
+.fa-ils:before {
+ content: "\f20b";
+}
+.fa-meanpath:before {
+ content: "\f20c";
+}
+.fa-buysellads:before {
+ content: "\f20d";
+}
+.fa-connectdevelop:before {
+ content: "\f20e";
+}
+.fa-dashcube:before {
+ content: "\f210";
+}
+.fa-forumbee:before {
+ content: "\f211";
+}
+.fa-leanpub:before {
+ content: "\f212";
+}
+.fa-sellsy:before {
+ content: "\f213";
+}
+.fa-shirtsinbulk:before {
+ content: "\f214";
+}
+.fa-simplybuilt:before {
+ content: "\f215";
+}
+.fa-skyatlas:before {
+ content: "\f216";
+}
+.fa-cart-plus:before {
+ content: "\f217";
+}
+.fa-cart-arrow-down:before {
+ content: "\f218";
+}
+.fa-diamond:before {
+ content: "\f219";
+}
+.fa-ship:before {
+ content: "\f21a";
+}
+.fa-user-secret:before {
+ content: "\f21b";
+}
+.fa-motorcycle:before {
+ content: "\f21c";
+}
+.fa-street-view:before {
+ content: "\f21d";
+}
+.fa-heartbeat:before {
+ content: "\f21e";
+}
+.fa-venus:before {
+ content: "\f221";
+}
+.fa-mars:before {
+ content: "\f222";
+}
+.fa-mercury:before {
+ content: "\f223";
+}
+.fa-transgender:before {
+ content: "\f224";
+}
+.fa-transgender-alt:before {
+ content: "\f225";
+}
+.fa-venus-double:before {
+ content: "\f226";
+}
+.fa-mars-double:before {
+ content: "\f227";
+}
+.fa-venus-mars:before {
+ content: "\f228";
+}
+.fa-mars-stroke:before {
+ content: "\f229";
+}
+.fa-mars-stroke-v:before {
+ content: "\f22a";
+}
+.fa-mars-stroke-h:before {
+ content: "\f22b";
+}
+.fa-neuter:before {
+ content: "\f22c";
+}
+.fa-facebook-official:before {
+ content: "\f230";
+}
+.fa-pinterest-p:before {
+ content: "\f231";
+}
+.fa-whatsapp:before {
+ content: "\f232";
+}
+.fa-server:before {
+ content: "\f233";
+}
+.fa-user-plus:before {
+ content: "\f234";
+}
+.fa-user-times:before {
+ content: "\f235";
+}
+.fa-hotel:before,
+.fa-bed:before {
+ content: "\f236";
+}
+.fa-viacoin:before {
+ content: "\f237";
+}
+.fa-train:before {
+ content: "\f238";
+}
+.fa-subway:before {
+ content: "\f239";
+}
+.fa-medium:before {
+ content: "\f23a";
+}
diff --git a/vendors/font-awesome/css/font-awesome.min.css b/vendors/font-awesome/css/font-awesome.min.css
new file mode 100644
index 0000000..24fcc04
--- /dev/null
+++ b/vendors/font-awesome/css/font-awesome.min.css
@@ -0,0 +1,4 @@
+/*!
+ * Font Awesome 4.3.0 by @davegandy - http://fontawesome.io - @fontawesome
+ * License - http://fontawesome.io/license (Font: SIL OFL 1.1, CSS: MIT License)
+ */@font-face{font-family:'FontAwesome';src:url('../fonts/fontawesome-webfont.eot?v=4.3.0');src:url('../fonts/fontawesome-webfont.eot?#iefix&v=4.3.0') format('embedded-opentype'),url('../fonts/fontawesome-webfont.woff2?v=4.3.0') format('woff2'),url('../fonts/fontawesome-webfont.woff?v=4.3.0') format('woff'),url('../fonts/fontawesome-webfont.ttf?v=4.3.0') format('truetype'),url('../fonts/fontawesome-webfont.svg?v=4.3.0#fontawesomeregular') format('svg');font-weight:normal;font-style:normal}.fa{display:inline-block;font:normal normal normal 14px/1 FontAwesome;font-size:inherit;text-rendering:auto;-webkit-font-smoothing:antialiased;-moz-osx-font-smoothing:grayscale;transform:translate(0, 0)}.fa-lg{font-size:1.33333333em;line-height:.75em;vertical-align:-15%}.fa-2x{font-size:2em}.fa-3x{font-size:3em}.fa-4x{font-size:4em}.fa-5x{font-size:5em}.fa-fw{width:1.28571429em;text-align:center}.fa-ul{padding-left:0;margin-left:2.14285714em;list-style-type:none}.fa-ul>li{position:relative}.fa-li{position:absolute;left:-2.14285714em;width:2.14285714em;top:.14285714em;text-align:center}.fa-li.fa-lg{left:-1.85714286em}.fa-border{padding:.2em .25em .15em;border:solid .08em #eee;border-radius:.1em}.pull-right{float:right}.pull-left{float:left}.fa.pull-left{margin-right:.3em}.fa.pull-right{margin-left:.3em}.fa-spin{-webkit-animation:fa-spin 2s infinite linear;animation:fa-spin 2s infinite linear}.fa-pulse{-webkit-animation:fa-spin 1s infinite steps(8);animation:fa-spin 1s infinite steps(8)}@-webkit-keyframes fa-spin{0%{-webkit-transform:rotate(0deg);transform:rotate(0deg)}100%{-webkit-transform:rotate(359deg);transform:rotate(359deg)}}@keyframes fa-spin{0%{-webkit-transform:rotate(0deg);transform:rotate(0deg)}100%{-webkit-transform:rotate(359deg);transform:rotate(359deg)}}.fa-rotate-90{filter:progid:DXImageTransform.Microsoft.BasicImage(rotation=1);-webkit-transform:rotate(90deg);-ms-transform:rotate(90deg);transform:rotate(90deg)}.fa-rotate-180{filter:progid:DXImageTransform.Microsoft.BasicImage(rotation=2);-webkit-transform:rotate(180deg);-ms-transform:rotate(180deg);transform:rotate(180deg)}.fa-rotate-270{filter:progid:DXImageTransform.Microsoft.BasicImage(rotation=3);-webkit-transform:rotate(270deg);-ms-transform:rotate(270deg);transform:rotate(270deg)}.fa-flip-horizontal{filter:progid:DXImageTransform.Microsoft.BasicImage(rotation=0, mirror=1);-webkit-transform:scale(-1, 1);-ms-transform:scale(-1, 1);transform:scale(-1, 1)}.fa-flip-vertical{filter:progid:DXImageTransform.Microsoft.BasicImage(rotation=2, mirror=1);-webkit-transform:scale(1, -1);-ms-transform:scale(1, -1);transform:scale(1, -1)}:root .fa-rotate-90,:root .fa-rotate-180,:root .fa-rotate-270,:root .fa-flip-horizontal,:root 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diff --git a/vendors/font-awesome/fonts/fontawesome-webfont.eot b/vendors/font-awesome/fonts/fontawesome-webfont.eot
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diff --git a/vendors/font-awesome/fonts/fontawesome-webfont.svg b/vendors/font-awesome/fonts/fontawesome-webfont.svg
new file mode 100644
index 0000000..1ee89d4
--- /dev/null
+++ b/vendors/font-awesome/fonts/fontawesome-webfont.svg
@@ -0,0 +1,565 @@
+
+
+
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diff --git a/vendors/font-awesome/fonts/fontawesome-webfont.ttf b/vendors/font-awesome/fonts/fontawesome-webfont.ttf
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diff --git a/vendors/owl-carousel/AjaxLoader.gif b/vendors/owl-carousel/AjaxLoader.gif
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diff --git a/vendors/owl-carousel/grabbing.png b/vendors/owl-carousel/grabbing.png
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diff --git a/vendors/owl-carousel/owl.carousel.css b/vendors/owl-carousel/owl.carousel.css
new file mode 100644
index 0000000..4e3c17c
--- /dev/null
+++ b/vendors/owl-carousel/owl.carousel.css
@@ -0,0 +1,71 @@
+/*
+ * Core Owl Carousel CSS File
+ * v1.3.3
+ */
+
+/* clearfix */
+.owl-carousel .owl-wrapper:after {
+ content: ".";
+ display: block;
+ clear: both;
+ visibility: hidden;
+ line-height: 0;
+ height: 0;
+}
+/* display none until init */
+.owl-carousel{
+ display: none;
+ position: relative;
+ width: 100%;
+ -ms-touch-action: pan-y;
+}
+.owl-carousel .owl-wrapper{
+ display: none;
+ position: relative;
+ -webkit-transform: translate3d(0px, 0px, 0px);
+}
+.owl-carousel .owl-wrapper-outer{
+ overflow: hidden;
+ position: relative;
+ width: 100%;
+}
+.owl-carousel .owl-wrapper-outer.autoHeight{
+ -webkit-transition: height 500ms ease-in-out;
+ -moz-transition: height 500ms ease-in-out;
+ -ms-transition: height 500ms ease-in-out;
+ -o-transition: height 500ms ease-in-out;
+ transition: height 500ms ease-in-out;
+}
+
+.owl-carousel .owl-item{
+ float: left;
+}
+.owl-controls .owl-page,
+.owl-controls .owl-buttons div{
+ cursor: pointer;
+}
+.owl-controls {
+ -webkit-user-select: none;
+ -khtml-user-select: none;
+ -moz-user-select: none;
+ -ms-user-select: none;
+ user-select: none;
+ -webkit-tap-highlight-color: rgba(0, 0, 0, 0);
+}
+
+/* mouse grab icon */
+.grabbing {
+ cursor:url(grabbing.png) 8 8, move;
+}
+
+/* fix */
+.owl-carousel .owl-wrapper,
+.owl-carousel .owl-item{
+ -webkit-backface-visibility: hidden;
+ -moz-backface-visibility: hidden;
+ -ms-backface-visibility: hidden;
+ -webkit-transform: translate3d(0,0,0);
+ -moz-transform: translate3d(0,0,0);
+ -ms-transform: translate3d(0,0,0);
+}
+
diff --git a/vendors/owl-carousel/owl.carousel.js b/vendors/owl-carousel/owl.carousel.js
new file mode 100644
index 0000000..a5d9ac1
--- /dev/null
+++ b/vendors/owl-carousel/owl.carousel.js
@@ -0,0 +1,1512 @@
+/*
+ * jQuery OwlCarousel v1.3.3
+ *
+ * Copyright (c) 2013 Bartosz Wojciechowski
+ * http://www.owlgraphic.com/owlcarousel/
+ *
+ * Licensed under MIT
+ *
+ */
+
+/*JS Lint helpers: */
+/*global dragMove: false, dragEnd: false, $, jQuery, alert, window, document */
+/*jslint nomen: true, continue:true */
+
+if (typeof Object.create !== "function") {
+ Object.create = function (obj) {
+ function F() {}
+ F.prototype = obj;
+ return new F();
+ };
+}
+(function ($, window, document) {
+
+ var Carousel = {
+ init : function (options, el) {
+ var base = this;
+
+ base.$elem = $(el);
+ base.options = $.extend({}, $.fn.owlCarousel.options, base.$elem.data(), options);
+
+ base.userOptions = options;
+ base.loadContent();
+ },
+
+ loadContent : function () {
+ var base = this, url;
+
+ function getData(data) {
+ var i, content = "";
+ if (typeof base.options.jsonSuccess === "function") {
+ base.options.jsonSuccess.apply(this, [data]);
+ } else {
+ for (i in data.owl) {
+ if (data.owl.hasOwnProperty(i)) {
+ content += data.owl[i].item;
+ }
+ }
+ base.$elem.html(content);
+ }
+ base.logIn();
+ }
+
+ if (typeof base.options.beforeInit === "function") {
+ base.options.beforeInit.apply(this, [base.$elem]);
+ }
+
+ if (typeof base.options.jsonPath === "string") {
+ url = base.options.jsonPath;
+ $.getJSON(url, getData);
+ } else {
+ base.logIn();
+ }
+ },
+
+ logIn : function () {
+ var base = this;
+
+ base.$elem.data("owl-originalStyles", base.$elem.attr("style"));
+ base.$elem.data("owl-originalClasses", base.$elem.attr("class"));
+
+ base.$elem.css({opacity: 0});
+ base.orignalItems = base.options.items;
+ base.checkBrowser();
+ base.wrapperWidth = 0;
+ base.checkVisible = null;
+ base.setVars();
+ },
+
+ setVars : function () {
+ var base = this;
+ if (base.$elem.children().length === 0) {return false; }
+ base.baseClass();
+ base.eventTypes();
+ base.$userItems = base.$elem.children();
+ base.itemsAmount = base.$userItems.length;
+ base.wrapItems();
+ base.$owlItems = base.$elem.find(".owl-item");
+ base.$owlWrapper = base.$elem.find(".owl-wrapper");
+ base.playDirection = "next";
+ base.prevItem = 0;
+ base.prevArr = [0];
+ base.currentItem = 0;
+ base.customEvents();
+ base.onStartup();
+ },
+
+ onStartup : function () {
+ var base = this;
+ base.updateItems();
+ base.calculateAll();
+ base.buildControls();
+ base.updateControls();
+ base.response();
+ base.moveEvents();
+ base.stopOnHover();
+ base.owlStatus();
+
+ if (base.options.transitionStyle !== false) {
+ base.transitionTypes(base.options.transitionStyle);
+ }
+ if (base.options.autoPlay === true) {
+ base.options.autoPlay = 5000;
+ }
+ base.play();
+
+ base.$elem.find(".owl-wrapper").css("display", "block");
+
+ if (!base.$elem.is(":visible")) {
+ base.watchVisibility();
+ } else {
+ base.$elem.css("opacity", 1);
+ }
+ base.onstartup = false;
+ base.eachMoveUpdate();
+ if (typeof base.options.afterInit === "function") {
+ base.options.afterInit.apply(this, [base.$elem]);
+ }
+ },
+
+ eachMoveUpdate : function () {
+ var base = this;
+
+ if (base.options.lazyLoad === true) {
+ base.lazyLoad();
+ }
+ if (base.options.autoHeight === true) {
+ base.autoHeight();
+ }
+ base.onVisibleItems();
+
+ if (typeof base.options.afterAction === "function") {
+ base.options.afterAction.apply(this, [base.$elem]);
+ }
+ },
+
+ updateVars : function () {
+ var base = this;
+ if (typeof base.options.beforeUpdate === "function") {
+ base.options.beforeUpdate.apply(this, [base.$elem]);
+ }
+ base.watchVisibility();
+ base.updateItems();
+ base.calculateAll();
+ base.updatePosition();
+ base.updateControls();
+ base.eachMoveUpdate();
+ if (typeof base.options.afterUpdate === "function") {
+ base.options.afterUpdate.apply(this, [base.$elem]);
+ }
+ },
+
+ reload : function () {
+ var base = this;
+ window.setTimeout(function () {
+ base.updateVars();
+ }, 0);
+ },
+
+ watchVisibility : function () {
+ var base = this;
+
+ if (base.$elem.is(":visible") === false) {
+ base.$elem.css({opacity: 0});
+ window.clearInterval(base.autoPlayInterval);
+ window.clearInterval(base.checkVisible);
+ } else {
+ return false;
+ }
+ base.checkVisible = window.setInterval(function () {
+ if (base.$elem.is(":visible")) {
+ base.reload();
+ base.$elem.animate({opacity: 1}, 200);
+ window.clearInterval(base.checkVisible);
+ }
+ }, 500);
+ },
+
+ wrapItems : function () {
+ var base = this;
+ base.$userItems.wrapAll("
+
+
+
+
+
+
+
+
+
+
+
+
+
+ +
+
+
+
+
+
+
+ Disciplines and Domains
+ +
+
+
+
+
+
+
+
+ ").wrap("");
+ base.$elem.find(".owl-wrapper").wrap("
");
+ base.wrapperOuter = base.$elem.find(".owl-wrapper-outer");
+ base.$elem.css("display", "block");
+ },
+
+ baseClass : function () {
+ var base = this,
+ hasBaseClass = base.$elem.hasClass(base.options.baseClass),
+ hasThemeClass = base.$elem.hasClass(base.options.theme);
+
+ if (!hasBaseClass) {
+ base.$elem.addClass(base.options.baseClass);
+ }
+
+ if (!hasThemeClass) {
+ base.$elem.addClass(base.options.theme);
+ }
+ },
+
+ updateItems : function () {
+ var base = this, width, i;
+
+ if (base.options.responsive === false) {
+ return false;
+ }
+ if (base.options.singleItem === true) {
+ base.options.items = base.orignalItems = 1;
+ base.options.itemsCustom = false;
+ base.options.itemsDesktop = false;
+ base.options.itemsDesktopSmall = false;
+ base.options.itemsTablet = false;
+ base.options.itemsTabletSmall = false;
+ base.options.itemsMobile = false;
+ return false;
+ }
+
+ width = $(base.options.responsiveBaseWidth).width();
+
+ if (width > (base.options.itemsDesktop[0] || base.orignalItems)) {
+ base.options.items = base.orignalItems;
+ }
+ if (base.options.itemsCustom !== false) {
+ //Reorder array by screen size
+ base.options.itemsCustom.sort(function (a, b) {return a[0] - b[0]; });
+
+ for (i = 0; i < base.options.itemsCustom.length; i += 1) {
+ if (base.options.itemsCustom[i][0] <= width) {
+ base.options.items = base.options.itemsCustom[i][1];
+ }
+ }
+
+ } else {
+
+ if (width <= base.options.itemsDesktop[0] && base.options.itemsDesktop !== false) {
+ base.options.items = base.options.itemsDesktop[1];
+ }
+
+ if (width <= base.options.itemsDesktopSmall[0] && base.options.itemsDesktopSmall !== false) {
+ base.options.items = base.options.itemsDesktopSmall[1];
+ }
+
+ if (width <= base.options.itemsTablet[0] && base.options.itemsTablet !== false) {
+ base.options.items = base.options.itemsTablet[1];
+ }
+
+ if (width <= base.options.itemsTabletSmall[0] && base.options.itemsTabletSmall !== false) {
+ base.options.items = base.options.itemsTabletSmall[1];
+ }
+
+ if (width <= base.options.itemsMobile[0] && base.options.itemsMobile !== false) {
+ base.options.items = base.options.itemsMobile[1];
+ }
+ }
+
+ //if number of items is less than declared
+ if (base.options.items > base.itemsAmount && base.options.itemsScaleUp === true) {
+ base.options.items = base.itemsAmount;
+ }
+ },
+
+ response : function () {
+ var base = this,
+ smallDelay,
+ lastWindowWidth;
+
+ if (base.options.responsive !== true) {
+ return false;
+ }
+ lastWindowWidth = $(window).width();
+
+ base.resizer = function () {
+ if ($(window).width() !== lastWindowWidth) {
+ if (base.options.autoPlay !== false) {
+ window.clearInterval(base.autoPlayInterval);
+ }
+ window.clearTimeout(smallDelay);
+ smallDelay = window.setTimeout(function () {
+ lastWindowWidth = $(window).width();
+ base.updateVars();
+ }, base.options.responsiveRefreshRate);
+ }
+ };
+ $(window).resize(base.resizer);
+ },
+
+ updatePosition : function () {
+ var base = this;
+ base.jumpTo(base.currentItem);
+ if (base.options.autoPlay !== false) {
+ base.checkAp();
+ }
+ },
+
+ appendItemsSizes : function () {
+ var base = this,
+ roundPages = 0,
+ lastItem = base.itemsAmount - base.options.items;
+
+ base.$owlItems.each(function (index) {
+ var $this = $(this);
+ $this
+ .css({"width": base.itemWidth})
+ .data("owl-item", Number(index));
+
+ if (index % base.options.items === 0 || index === lastItem) {
+ if (!(index > lastItem)) {
+ roundPages += 1;
+ }
+ }
+ $this.data("owl-roundPages", roundPages);
+ });
+ },
+
+ appendWrapperSizes : function () {
+ var base = this,
+ width = base.$owlItems.length * base.itemWidth;
+
+ base.$owlWrapper.css({
+ "width": width * 2,
+ "left": 0
+ });
+ base.appendItemsSizes();
+ },
+
+ calculateAll : function () {
+ var base = this;
+ base.calculateWidth();
+ base.appendWrapperSizes();
+ base.loops();
+ base.max();
+ },
+
+ calculateWidth : function () {
+ var base = this;
+ base.itemWidth = Math.round(base.$elem.width() / base.options.items);
+ },
+
+ max : function () {
+ var base = this,
+ maximum = ((base.itemsAmount * base.itemWidth) - base.options.items * base.itemWidth) * -1;
+ if (base.options.items > base.itemsAmount) {
+ base.maximumItem = 0;
+ maximum = 0;
+ base.maximumPixels = 0;
+ } else {
+ base.maximumItem = base.itemsAmount - base.options.items;
+ base.maximumPixels = maximum;
+ }
+ return maximum;
+ },
+
+ min : function () {
+ return 0;
+ },
+
+ loops : function () {
+ var base = this,
+ prev = 0,
+ elWidth = 0,
+ i,
+ item,
+ roundPageNum;
+
+ base.positionsInArray = [0];
+ base.pagesInArray = [];
+
+ for (i = 0; i < base.itemsAmount; i += 1) {
+ elWidth += base.itemWidth;
+ base.positionsInArray.push(-elWidth);
+
+ if (base.options.scrollPerPage === true) {
+ item = $(base.$owlItems[i]);
+ roundPageNum = item.data("owl-roundPages");
+ if (roundPageNum !== prev) {
+ base.pagesInArray[prev] = base.positionsInArray[i];
+ prev = roundPageNum;
+ }
+ }
+ }
+ },
+
+ buildControls : function () {
+ var base = this;
+ if (base.options.navigation === true || base.options.pagination === true) {
+ base.owlControls = $("
").toggleClass("clickable", !base.browser.isTouch).appendTo(base.$elem);
+ }
+ if (base.options.pagination === true) {
+ base.buildPagination();
+ }
+ if (base.options.navigation === true) {
+ base.buildButtons();
+ }
+ },
+
+ buildButtons : function () {
+ var base = this,
+ buttonsWrapper = $("