The fate of bilirubin-IXα glucuronide in cholestasis and during storage in vitro. Intramolecular rearrangement to positional isomers of glucuronic acid

Abstract

In aqueous solution above pH 7 bilirubin-IXα 1-O-acylglucuronide rapidly isomerizes to the non-C-1 glucuronides by sequential migration of the bilirubin acyl group from position 1 to positions 2, 3 and 4 of the sugar moiety. The transformations are enhanced by increasing the pH. Compared with the rates at 37°C, the transformations are rather slow at 0°C. Virtually complete inhibition is observed at values below pH 6. The isomerization at 25°C and pH 7.4 is not affected by the presence in the solutions of a molar excess of human serum albumin. Isomerization in bile kept at 37°C at pH 7.7-7.8 is probably non-enzymic, as the rates of change are similar to those observed under comparable conditions for aqueous solutions of glucuronides of bilirubin-IXα and of azodipyrrole. Analysis without delay of normal biles of man and rats collected at 0°C over a maximum period of 10 mins shows that the bilirubin-IXα mono- and di-glucuronides consist exclusively of the 1-O-acyl isomers. The mixtures of the four positional isomers of bilirubin-IXα glucuronide found in freshly collected biles of man and rats with cholestasis probably originate from initially synthesized 1-O-acylglucuronide by the same mechanism of sequential migration as has been observed in aqueous solutions of conjugated bilirubin-IXa.