Novel type of ornithine-glutathione double conjugate excreted as a major metabolite into the bile of rats administered clebopride.
Rats orally given radioactive Clebopride [[14C]CP; N-(1'-benzyl-4'-piperidyl)-2-[14C]methoxy-4-amino-5-chlorobenzamide++ +], an antiulcer agent, excreted a novel type of ornithine (Orn)-GSH double conjugate in the bile as a major metabolite [( 14C]BMCP), corresponding to 18% of the dose. The present study provides the first evidence for Orn conjugation of a xenobiotic in mammals and demonstrates that the structure of the radioactive conjugate differs fundamentally from those known in birds and reptiles. The structure of the biliary metabolite, [14C]BMCP, purified to homogeneity by silica gel thin layer and reverse phase high pressure liquid chromatography, was elucidated as S-[2-ornithylamino-4-[14C]methoxy-5-(1'-methyl-4'-piperidylamin o) carboxyphenyl]glutathione, based mainly on the following facts: 1) BMCP showed a protonated molecular ion (M + H)+ peak at m/z 683 in the secondary ion mass spectrum and 2) [14C]BMCP afforded Orn, glutamic acid, glycine, S-(2-amino-4-[14C]methoxy-5-carboxyphenyl)cysteine [( 14C]AMCC), and 1-methyl-4-aminopiperidine (MAP) quantitatively, in an equal molar ratio, by complete hydrolysis with peptidase. Thus, BMCP was a metabolite with three enzymatically hydrolyzable amide bonds in addition to the one existing originally in the parent structure of the drug, which produces MAP by peptic digestion. Of the three additional amide bonds of BMCP, one was a novel type of bond formed by condensation of the alpha-carboxylic acid group of Orn with the primary aromatic amino group of the drug and the other two were in the S-glutathionyl residue, substituted for the chlorine atom vicinal to the Orn-conjugating primary amino group in the aromatic ring and affording glutamic acid, glycine, and the S-cysteine conjugate AMCC by hydrolysis of BMCP with the peptidase. Substitution of a methyl group for the benzyl group at the piperidine ring nitrogen atom, leading to the formation of MAP by peptic digestion, also occurred during metabolism of CP to BMCP.
Ishizuka
T
T
Toxicology Research Laboratory, Pharmaceutical Research Center, Meiji Seika Kaisha, Ltd., Yokohama, Japan.
Komiya
I
I
Hiratsuka
A
A
Watabe
T
T
D000818:Animals; D001549:Benzamides / Q000494:pharmacology*; D001646:Bile / Q000187:drug effects / Q000378:metabolism*; D002250:Carbon Radioisotopes; D005978:Glutathione / Q000031:analogs & derivatives* / Q000378:metabolism; D006868:Hydrolysis; D008297:Male; D008657:Metabolic Clearance Rate; D009952:Ornithine / Q000031:analogs & derivatives* / Q000378:metabolism; D051381:Rats; D011919:Rats, Inbred Strains; D013056:Spectrophotometry, Ultraviolet; D013329:Structure-Activity Relationship
D001549:Benzamides; D002250:Carbon Radioisotopes; C064706:S-(2-ornithylamino-4-methoxy-5-(1'-methyl-4'-piperidylamino)carboxyphenyl)glutathione; D009952:Ornithine; D005978:Glutathione; C014417:clebopride