Characterization of cholyl-adenylate in rat liver microsomes by liquid chromatography/electrospray ionization-mass spectrometry.
The cholyl-adenylate, covalently bound 3alpha, 7alpha, 12alpha-trihydroxy-5beta-cholanoic acid (cholic acid) with adenosine 5'-monophosphate having an acid anhydride linkage, has been characterized by means of liquid chromatography/mass spectrometry in an incubation mixture with a rat liver microsomal fraction. The authentic specimen of cholyl-adenylate was synthesized using the carbodiimide method and the structure was confirmed by MS and nuclear magnetic resonance spectroscopy. After incubation of cholic acid with a hepatic microsomal fraction in the presence of adenosine 5'-triphosphate, bile acids were extracted and purified by solid-phase extraction on a Sep-Pak C18 cartridge and then subjected to a LC/MS analysis, where cholyl-adenylate and a CoA thioester of cholic acid (cholyl-CoA) were monitored with characteristic negative ions of m/z 736 and 577, respectively. Cholyl-adenylate was definitely characterized and preferential biotransformation into the acyl-adenylate prior to formation of cholyl-CoA was noted. The nonenzymatic formation of taurine-conjugated cholic acid by incubation of cholyl-adenylate with taurine in a buffer solution was also demonstrated.
Ikegawa
S
S
Faculty of Pharmaceutical Sciences, Tohoku University, Aobayama Sendai, 980-8578, Japan.
Ishikawa
H
H
Oiwa
H
H
Nagata
M
M
Goto
J
J
Kozaki
T
T
Gotowda
M
M
Asakawa
N
N
D000214:Acyl Coenzyme A / Q000138:chemical synthesis / Q000378:metabolism; D000249:Adenosine Monophosphate / Q000031:analogs & derivatives* / Q000032:analysis / Q000138:chemical synthesis / Q000737:chemistry; D000255:Adenosine Triphosphate / Q000737:chemistry; D000818:Animals; D019826:Cholic Acid / Q000737:chemistry; D002793:Cholic Acids / Q000032:analysis* / Q000138:chemical synthesis / Q000737:chemistry* / Q000378:metabolism; D002853:Chromatography, Liquid / Q000379:methods*; D008297:Male; D013058:Mass Spectrometry / Q000379:methods*; D008862:Microsomes, Liver / Q000737:chemistry* / Q000378:metabolism; D051381:Rats; D017208:Rats, Wistar; D013654:Taurine / Q000737:chemistry; D013656:Taurocholic Acid / Q000737:chemistry
D000214:Acyl Coenzyme A; D002793:Cholic Acids; C117584:cholyl-adenylate; C117585:cholyl-coenzyme A; D013654:Taurine; D000249:Adenosine Monophosphate; D013656:Taurocholic Acid; D000255:Adenosine Triphosphate; D019826:Cholic Acid