High-performance liquid chromatographic determination of ursodeoxycholic acid after solid phase extraction of blood serum and detection-oriented derivatization.
Ursodeoxycholic acid (3 alpha,7 beta-dihydroxy-5 beta-cholanoic acid, UDCA) is a therapeutically applicable bile acid widely used for the dissolution of cholesterol-rich gallstones and in the treatment of chronic liver diseases associated with cholestasis. UDCA is more hydrophilic and less toxic than another therapeutically valuable bile acid, chenodeoxycholic acid (CDCA), the 7 alpha-epimer of UDCA. Procedures for sample preparation and HPLC determination of UDCA in blood serum were developed and validated. A higher homologue of UDCA containing an additional methylene group in the side chain was synthetized and used as an internal standard (IS). Serum samples with IS were diluted with a buffer (pH=7). The bile acids and IS were captured using solid phase extraction (C18 cartridges). The carboxylic group of the analytes was derivatized using 2-bromo-2'-acetonaphthone (a detection-oriented derivatization), and reaction mixtures were analyzed (HPLC with UV 245 nm detection; a 125--4 mm column containing Lichrospher 100 C18, 5 microm; mobile phase: acetonitrile--water, 6:4 (v/v)). Following validation, this method was used for pharmacokinetic studies of UDCA in humans.
Nobilis
M
M
Institute of Experimental Biopharmaceutics, Joint Research Centre of PRO.MED.CS Praha a.s. and Academy of Sciences of the Czech Republic, Heyrovského 1207, CZ-500 02 Hradec Králové, Czech Republic. nobilis@uebf.cas.cz
Pour
M
M
Kunes
J
J
Kopecký
J
J
Kvĕtina
J
J
Svoboda
Z
Z
Sládková
K
K
Vortel
J
J
D000328:Adult; D002851:Chromatography, High Pressure Liquid / Q000379:methods*; D005260:Female; D006801:Humans; D008297:Male; D010865:Pilot Projects; D012015:Reference Standards; D012016:Reference Values; D015203:Reproducibility of Results; D013056:Spectrophotometry, Ultraviolet; D014580:Ursodeoxycholic Acid / Q000097:blood* / Q000493:pharmacokinetics
D014580:Ursodeoxycholic Acid