Chiral immunoaffinity chromatography in miniaturized systems.

摘要

Miniaturized separation systems are an attractive option for the separation of a wide range of analytes. Smaller columns result in reduced consumption of stationary phase, solvents, and samples. This is particularly important in the case of chiral separations since chiral selectors and chiral analytes tend to be expensive. In this research, two miniaturized separation systems were established, namely, microbore and capillary separations which contain stereoselective antibodies as chiral selectors. Microbore columns were used for separations of amino acid enantiomers with both UV and mass spectrometric detection. The temperature of microbore columns could be controlled using a homemade water jacket connected to a thermostated waterbath. An additional study confirmed that aqueous buffers at physiological pH were suitable mobile phases; specifically, phosphate buffers were ideal for UV detection and ammonium bicarbonate buffer was ideal for mass spectrometric detection. Use of mass spectrometric detection was particularly advantageous since it enabled the simultaneous separation of multiple amino acid enantiomer pairs. Capillary columns containing stereoselective antibodies were also developed; several parameters were investigated to optimize preparation of capillary columns. For both microbore and capillary columns, separations improved on longer columns and at lower temperatures. Preliminary studies on chiral immunoaffinity capillary electrochromatography and on the development of monolithic immunoaffinity columns are also presented.