Rearranged glucuronic acid conjugates of bilirubin-IX alpha in post-obstructive bile. Structure elucidation  of azopigments beta and gamma as ethyl anthranilate N-glycosides derived from 2-, 3- and 4-o-acyl glucuronides.
Azopigment analysis was performed on conjugates of bilirubin-IXalpha in bile of man and rats obtained after obstruction of the bile duct or in bile incubated under N2. The azopigments beta and gamma, formed by applying a pH 2.7 diazonium reagent containing an excess of ethyl anthranilate, correspond to rearranged ethyl athranilate N-glucuronides having the azodipyrrole acyl group on positions 2, 3 and 4 of the sugar. These assignments were verified, first by conversion of the structurally known 2-, 3- and 4-O-acyl glucuronide azopigments, unsubstituted at C-1, into ethyl anthranilate N-glucuronide reference compounds, and second, by mass spectrometry of trimethylsilyl ether methyl ester derivatives of unknown and reference compounds. The C-1 ethyl anthranilate group of the N-glucuronides triggers characteristics fragmentation reactions of the carbohydrate moiety revealing the position of the azodipyrrole O-acyl group.
D000818:Animals; D001391:Azo Compounds / Q000032:analysis; D001646:Bile / Q000032:analysis*; D001654:Bile Pigments / Q000032:analysis; D001663:Bilirubin / Q000031:analogs & derivatives*; D055598:Chemical Phenomena; D002621:Chemistry; D002855:Chromatography, Thin Layer; D005965:Glucuronates; D006801:Humans; D013058:Mass Spectrometry / Q000379:methods; D051381:Rats; D014297:Trimethylsilyl Compounds; D062367:ortho-Aminobenzoates
D001391:Azo Compounds; D001654:Bile Pigments; D005965:Glucuronates; D014297:Trimethylsilyl Compounds; D062367:ortho-Aminobenzoates; D001663:Bilirubin