Application of 3 alpha-hydroxysteroid dehydrogenase column to the determination of bile acids fractionated by high-performance liquid chromatography: advantage of pretreating human bile acids with Seppak C18 and piperidinohydroxypropyl Sephadex LH-20.
The analysis of bile acids in human bile was tried by high-performance liquid chromatography (HPLC). Bile acids in human bile were first prefractionated into free, glycine- and taurine-conjugated bile acids using a Seppak C18 cartridge and a piperidinohydroxypropyl Sephadex LH-20 (PHP-LH-20) column. Each fraction was then processed through a HPLC system consisting of a Zorbax ODS column and a 3 alpha-hydroxysteroid dehydrogenase (3 alpha-HSD) column. By these procedures the major 15 bile acids were clearly separated, and each bile acid of 10-125 ng was accurately analyzed. More than 400 times of analysis could be repeated on one 3 alpha-HSD column without loss of sensitivity. Thus the pretreatment through Seppak C18 and PHP-LH-20 made the HPLC analysis of human bile acids accurate and applicable to clinically obtained materials.
D015096:3-Hydroxysteroid Dehydrogenases; D045002:3-alpha-Hydroxysteroid Dehydrogenase (B-Specific); D001646:Bile / Q000032:analysis; D001647:Bile Acids and Salts / Q000032:analysis*; D002851:Chromatography, High Pressure Liquid / Q000379:methods*; D003911:Dextrans; D004800:Enzymes, Immobilized; D006801:Humans; D012822:Silicon Dioxide
D001647:Bile Acids and Salts; D003911:Dextrans; D004800:Enzymes, Immobilized; D012822:Silicon Dioxide; C042953:octadecyl silane-bonded silica; C025614:sephadex; D015096:3-Hydroxysteroid Dehydrogenases; D045002:3-alpha-Hydroxysteroid Dehydrogenase (B-Specific)