Isolation of bile acid glucosides and N-acetylglucosaminides from human urine by ion-exchange chromatography and reversed-phase high-performance liquid chromatography.
A method for the isolation, separation and analysis of glucosides and N-acetylglucosaminides of non-amidated bile acids and of glycine- and taurine-conjugated bile acid glucosides from normal human urine is described. Total bile acids were extracted from 24-h collections of urine by repetitive use of Sep-Pak C18 cartridges. After elution with 80% aqueous methanol, a group separation into non-amidated, glycine- and taurine-conjugated bile acids was performed by ion-exchange chromatography on Lipidex-DEAP. The glycosylated compounds were then separated from the corresponding non-glycosylated ones by high-performance liquid chromatography (HPLC) using a reversed-phase system with a linear methanol gradient. The glycosylated compounds isolated by HPLC were analysed by fast atom bombardment mass spectrometry and, after derivatization, by gas chromatography-mass spectrometry. Information about the sugar moieties of the bile acid glycosides was also obtained by treatment with different glycosidases.
Marschall
H U
HU
Department of Physiological Chemistry, Karolinska Institutet, Stockholm, Sweden.
Green
G
G
Egestad
B
B
Sjövall
J
J
D000117:Acetylglucosamine / Q000031:analogs & derivatives* / Q000302:isolation & purification / Q000652:urine; D001647:Bile Acids and Salts / Q000302:isolation & purification / Q000652:urine*; D002849:Chromatography, Gas; D002851:Chromatography, High Pressure Liquid; D002852:Chromatography, Ion Exchange; D008401:Gas Chromatography-Mass Spectrometry; D005944:Glucosamine / Q000031:analogs & derivatives*; D006027:Glycosides / Q000302:isolation & purification / Q000652:urine; D006801:Humans; D006868:Hydrolysis; D013058:Mass Spectrometry; D012997:Solvents
D001647:Bile Acids and Salts; D006027:Glycosides; D012997:Solvents; D005944:Glucosamine; D000117:Acetylglucosamine