D-saccharic Acid Research Articles

Isolation and characterization of the major form of β-glucuronidase from human seminal plasma

Human seminal plasma contains two forms of β-glucuronidase (β- d-glucuronidase glucuronosohydrolase, EC 3.2.1.31) which are present in the ratio 4:1. The major form of β-glucuronidase with a slow moving band in electrophoresis was purified to homogeneity as revealed by polyacrylamide gel electrophoresis, double immunodiffusion and immunoelectrophoresis. The major form of β-glucuronidase shows dual optimum pH at 4.3 and 4.7 with a dip in the activity at pH 4.5. The K m of this form of β-glucuronidase is dependent on pH and was found to be 0.95 3.08 and 0.67 mM at pH 4.4, 4.5 and 4.7, respectively. The major form of β-glucuronidase from seminal plasma is stable at 55°C for 30 min but it denatures at 65°C. Heat denaturation is faster at acidic pH (4.7) than at alkaline pH (7.8). However, the activity of enzyme increase linearly with increased in temperature up to 70°C during incubation with substrate. Cu, Ag, Hg and Ni salts inhibited enzyme activity significantly at 0.1 and 1.0 mM concentration, but the inhibition by HgCl 2 was protected by cysteine. 1,4- d-Saccharic acid lactone and ascorbic acid inhibited seminal β-glucuronidase competitively, yielding k i values of 1.7 ·10 −3 mM and 10.3 mM, respectively. Through fructose and mannose also showed significant inhibition of β-glucuronidase at 10–100 mM, glucose did not show any effect. The molecular weight of the major form of β-glucuronidase was found to be 279 000, and it appears to be composed of four subunits each having a molecular weight of 74 000.

Hydrolysis of the biliary glucuronic acid conjugate of phenol by the intestinal mucus/flora of goldfish (Carassius auratus).

1. An investigation was carried out on the hydrolytic activity of the intestinal mucus/flora on phenylglucuronide in the bile of goldfish. 2. Approximately 79% of the total phenylglucuronide (3.4 mumol) in the bile was hydrolysed after 16 h incubation with the intestinal mucus/flora. 3. Of the total phenol in the aquarium water of goldfish exposed previously to phenol for 48 h, 41% was found to be phenylglucuronide when fish were placed in a phenol-free medium and were dosed hourly for 8 h with D-saccharic acid 1,4-lactone to inhibit beta-glucuronidase activity in the intestine.