Improved methods have been developed for separation of a-keto acid derivatives and for thiamine and its phosphate esters. Linear gradient reverse phase chromatography using ion-pair formation provides baseline resolution for a seven-component homologous series of a-keto acid dinitrophenylhydrazones having increasing carbon chain lengths. Branched-chain keto acids can also be separated. Aldehyde derivatives were examined as possible interfering compounds. Peak identification in biological material was confirmed for pyruvate by an enzymic peak shift technique. Monitoring near 365 nm permits low nanogram detection. Preliminary studies of keto acids in biological material illustrate the method's applicability. The same system with slight changes separates the thiamine phosphate esters and is compared with anion exchange chromatography. The thiamine method involves effluent oxidation to form the respective fluorescent thiochrome esters for selective fluorometric detection.
College and Department
Chemistry and Biochemistry
BYU ScholarsArchive Citation
Hemming, Bruce Clark, "High pressure liquid chromatographic methods for the separations of derivatized alpha-keto acids and thiamine phosphate esters" (1977). Theses and Dissertations. 8228.
Liquid chromatography, Acids, Esters