Development and application of polarimetric methods for the study of optically active materials

The main thrust of this thesis will be to present new advances in the utilization of laser-based polarimetry.;A new method has been developed to measure the specific rotation of optically active compounds using flow injection as a sample introduction system. The theoretical and experimental considerations necessary to obtain a Gaussian analyte distribution will be discussed. The effect of peak shape asymmetry on the calculation of specific rotation by the peak height method (which is based on a Gaussian model) will be evaluated. It will be shown that under proper experimental conditions, the peak height method is not affected to any significant extend by an asymmetric analyte distribution. The application of laser-based polarimetry for the detection and identification of chiral molecules which differ only slightly structurally will be demonstrated by a study of the antibiotic gentamicin. Specific rotations for four structural analogs of gentamicin will be reported and used to differentiate these compounds which differ only minimally structurally. The determination of the specific rotation of several different amino acids and the corresponding peptides under different conditions of pH has been accomplished. These results have shown that specific rotation measurements can be used to monitor subtle changes in the structure of a complicated molecule.;One of the most common techniques for the separation of proteins is by HPLC. However, the structure and therefore the activity of the protein being separated may change after passage through the separation column. In order to avoid changes in the activity of the protein which might be caused by the separation conditions, a flow injection technique has been used to measure the specific rotation of a protein. Ultimately, this information can be used to determine the optimum conditions for the HPLC separation of a protein while at the same time minimizing denaturation.;The use of laser-based polarimeter as a selective detector for optically active compounds has been extended to the detection of achiral compounds. Achiral ketones are transformed to their corresponding "SAMP-hydrazones" by reaction with the enantiomerically pure hydrazone, 1-amino-2-(methoxy-1-methyl)prorolidine. The hydrazone is produced on-line by an FIA method, using a merged stream arrangement to mix the ketone and hydrozone. A similar scheme could be adapted to HPLC. The experimental parameters necessary to achieve sensitive detection of achiral materials by this chiral-derivatization scheme will be discussed.