Structures of bilirubin conjugates synthesized in vitro from bilirubin and uridine diphosphate glucuronic acid, uridine diphosphate glucose or uridine diphosphate xylose by preparations from rat liver

Abstract

1. In incubation mixtures containing digitonin-activated or untreated preparations from rat liver, albumin-solubilized bilirubin as the acceptor substrate and (a) UDP-glucuronic acid, (b) UDP-glucose or (c) UDP-xylose as the sugar donor, formation of the following ester glycosides was demonstrated: with (a), bilirubin beta-d-monoglucuronoside, with (b), bilirubin beta-d-monoglucoside and with (c), bilirubin monoxyloside or mixtures of the mono-and di-xyloside. 2. With UDP-glucuronic acid prolonged incubation and variation of the composition of the incubation mixtures yielded equimolar amounts of azodipyrrole (I) and azodipyrrole beta-d-monoglucuronoside (II) after treatment of the incubation mixtures with the diazonium salt of ethyl anthranilate. The azo-derivatives were identified by t.l.c. by reference to known compounds and by the following chemical tests. After ammonolysis the conjugated azo-derivative (II) yielded d-glucuronic acid and the carboxylic acid amide of azodipyrrole, indicating transfer of a glucuronic acid residue to the carboxylic acid groups of bilirubin. The beta-d-configuration of the sugar moiety and binding at C-1 were demonstrated by enzymic hydrolysis tests. 3. Analogous evidence established the structure of the reaction product obtained with UDP-glucose as the sugar donor, as bilirubin beta-d-monoglucoside. 4. With UDP-xylose as the sugar donor xylosyl transfer to the carboxylic acid groups of bilirubin with attachment at C-1 was demonstrated in an analogous way. A beta-d-configuration is considered very likely, but requires confirmation. 5. Monoxyloside formation was predominant at pH7.4, whereas at decreasing pH values increasing fractions of the substrate were converted into the dixyloside. Prolonged incubation, low concentrations of bilirubin and high concentrations of UDP-xylose favoured diconjugate formation. The available evidence supports the synthesis sequence: bilirubin --> bilirubin monoxyloside --> bilirubin dixyloside.

Unconjugated Bilirubin and an Increased Proportion of Bilirubin Monoconjugates in the Bile of Patients with Gilbert's Syndrome and Crigler-Najjar Disease

“…1). Provided certain conditions are met, the azopigment fraction a0-EA (pH 2.7) multiplied by 2 equals the amount of monoconjugates expressed as a percentage of the total conjugated bilirubin-IXa (38,39). This ao-EA (pH 2.7) fraction can assume the extreme values of 0 (no monoconjugate) and 50% (only monoconjugates), but theoretically should never exceed 50%.…”

Separation by thin-layer chromatography and structure elucidation of bilirubin conjugates isolated from dog bile

“…Structure determination of these azopigments identifies a dipyrrole half with or without a conjugating group derived from the parent tetrapyrrole (e.g., see Fig. 2 of Fevery et al, 1972a). The procedure is valid provided neither fragments are lost nor changes occur during formation or purification of the azopigments.…”

“…Trivial References Compernolle et al (1970Compernolle et al ( , 1971; Jansen & Stoll (1971) Compernolle et al (1970); Jansen & Stoll (1971) Compernolle et al (1970) Compernolle et al (1971); Fevery et al (1971) Compemolle et al (1971; Fevery et al (1971; Compernolle et al (1970) Fevery et al (1972a) Table 2. Diazo-positive bile pigments isolatedfrom dog bile by t.l.c.…”

Diminished Enhancement of Hepatic UDP Glucuronyl Transferase (Bilirubin) by Phenobarbital during Pregnancy in the Rat

“…In vitro studies using rat liver homoge nates and microsomal fractions have demonstrated that bilirubin-monoxyloside or mixtures of mono-and dixyloside are also formed (8). During pregnancy a progressive shift in the conjugation of bilirubin could occur following PB ad ministration, such that UDPGT is not increased but rather UDP-glucosyl trans ferase and/or UDP-xylosil transferase are induced.…”