FUNDAMENTAL STUDIES ON ULTRAFAST CHIRAL AND ACHIRAL SEPARATIONS IN LIQUID CHROMATOGRAPHY AND SUB/SUPERCRITICAL FLUID CHROMATOGRAPHY

Abstract

This thesis is based on fundamental contributions in the field of ultrafast chiral and achiral separations using liquid and sub/supercritical fluid chromatography. The high efficiency sub-2 μm particles of narrow particle size distribution (NPSD) and 2.7 μm superficially porous particles (SPP) were used as a support for bonding various chiral selectors. The chiral selectors used in this thesis are teicoplanin (T), teicoplanin aglycone (TAG) and vancomycin (V) cyclofructan-6 derivatized isopropyl carbamate (CF6-P), cyclofructan-7 dimethylphenyl carbamate (CF7-DMP), tert-butyl carbomoylated quinine and hydroxypropyl-β cyclodextrin (HPRSP) . These high efficiency particles were packed in 0.5, 2.0, 3.0 and 5 cm long columns

As we enter the ultrafast chromatography regime various interesting phenomena manifest themselves which are usually not observed in routine chromatography. State of the art UHPLCs were shown to be insufficient for the narrow peaks obtained in the ultrafast chromatography in terms of extra-column volumes. Optimized instruments with reduced extra column volume, highest possible sampling frequencies and optimum digital filters were needed to preserve the “true” peak shape. Using short columns, chiral, achiral and shape selective separations were demonstrated in the sub-second to sub-minute range. Frictional heating was shown to be beneficial for both SPP and NPSD particles in chiral chromatography, unlike reversed phase chromatography. Additionally the instrumental idiosyncrasies of SFC are discussed such as the connection tubing diameter effect, noise analysis of backpressure regulator and peak shape analysis (upsampling effects on peak shapes).

Real world applications of pharmaceuticals and intermediates were developed for reverse phase liquid chromatography (RPLC) and SFC mode for 50 compounds belonging to various classes. This research culminated by exploring two dimensional liquid chromatography (2D-LC). Multiple achiral × chiral and chiral × chiral 2D-LC examples (single and multiple heart-cutting, high-resolution sampling, and comprehensive) using ultrafast chiral chromatography in the second dimension were successfully applied to the separation and analysis of complex mixtures of closely related pharmaceuticals and synthetic intermediates, including chiral, achiral drugs, metabolites, constitutional isomers, stereoisomers, and organohalogenated species.