Circular Dichroism Spectroscopy Lab
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
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.
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.