TY - JOUR
T1 - Separation and characterization of carboxyl-linked glucuronides of bile acids in incubation mixture of rat liver microsomes
AU - Goto, Junichi
AU - Murao, Naoaki
AU - Nakada, Chifumi
AU - Motoyama, Takashi
AU - Oohashi, Junji
AU - Yanagihara, Takamitsu
AU - Niwa, Toshifumi
AU - Ikegawa, Shigeo
N1 - Funding Information:
This work was supported in part by a grant from the Ministry of Education, Science, Sports and Culture of Japan.
PY - 1998/4
Y1 - 1998/4
N2 - The carboxyl-linked 24-glucuronides of common bile acids have been identified by means of liquid chromatography (LC)/atmospheric pressure chemical ionization (APCI)-mass spectrometry (MS) in an incubation mixture with a male Wistar rat liver microsomal fraction. The authentic specimens of bile acid 24-glucuronide acetate-methyl esters were synthesized unequivocally using the Mitsunobu reaction, and the APCI-mass spectrometric properties of these glucuronide derivatives were also characterized. After incubation of common unconjugated bile acids with hepatic microsomes, glucuronides were extracted and purified with a Sep-Pak C18 cartridge and lipophilic ion exchange gel, piperidino-hydroxypropyl Sephadex LH-20, and then derivatized into the acetate-methyl esters. Subsequent resolution into α- and β- isomers at the glucuronosyl linkage was attained by LC on Cosmosil 5C8 and Sumichiral OA-2500 columns using 200 mM ammonium acetate (pH 7.0)-methanol (1:4, v/v), where 24-glucuronides were monitored with characteristic positive ions [M + NH4]+. The 24-glucuronides of lithocholic, chenodeoxycholic, deoxycholic, ursodeoxycholic and cholic acid were definitely characterized, in contrast to no formation of corresponding 3-glucuronides.
AB - The carboxyl-linked 24-glucuronides of common bile acids have been identified by means of liquid chromatography (LC)/atmospheric pressure chemical ionization (APCI)-mass spectrometry (MS) in an incubation mixture with a male Wistar rat liver microsomal fraction. The authentic specimens of bile acid 24-glucuronide acetate-methyl esters were synthesized unequivocally using the Mitsunobu reaction, and the APCI-mass spectrometric properties of these glucuronide derivatives were also characterized. After incubation of common unconjugated bile acids with hepatic microsomes, glucuronides were extracted and purified with a Sep-Pak C18 cartridge and lipophilic ion exchange gel, piperidino-hydroxypropyl Sephadex LH-20, and then derivatized into the acetate-methyl esters. Subsequent resolution into α- and β- isomers at the glucuronosyl linkage was attained by LC on Cosmosil 5C8 and Sumichiral OA-2500 columns using 200 mM ammonium acetate (pH 7.0)-methanol (1:4, v/v), where 24-glucuronides were monitored with characteristic positive ions [M + NH4]+. The 24-glucuronides of lithocholic, chenodeoxycholic, deoxycholic, ursodeoxycholic and cholic acid were definitely characterized, in contrast to no formation of corresponding 3-glucuronides.
KW - Atmospheric pressure chemical ionization
KW - Bile acid
KW - Glucuronide
KW - Glucuronosyltransferase
KW - LC/MS
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U2 - 10.1016/S0039-128X(98)00002-6
DO - 10.1016/S0039-128X(98)00002-6
M3 - Article
C2 - 9589552
AN - SCOPUS:0032056005
VL - 63
SP - 186
EP - 192
JO - Steroids
JF - Steroids
SN - 0039-128X
IS - 4
ER -