Synthesisof p-Nitrophenyl β-D-Glucopyranosiduronic Acid and Its Utilization as a Substrate for the Assay of β-Glucuronidase Activity

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1. ρ-Nitrophenyl β-D-glucosiduronic acid is proposed as a substrate for the assay of the hydrolytic activity of β-glucuronidase. The liberated p-nitrophenol is measured photocolorimetrically in alkaline solution. 2. The chemical synthesis of p-nitropenyl β-D-glucopyranosiduronic acid was accomplished by alkaline hydrolysis of methyl (p-nitrophenyl 2,3,4-tri-O-acetyl-β-D-glucopyranosid) uronate which was prepared by the condensation of methyl 1-bromo-2,3,4-tri-O-acetyl-α-D-glucopyranuronate and p-nitrophenol using acetonitrile and freshly prepared silver oxide. 3. The kinetics of the hydrolysis of p-nitrophenyl β-D-glucopyranosiduronic acid by β-glucuronidase were investigated. The optimum pH is 3.8 in 0.2M acetate buffer at 38°. The reaction velocity is constant with time and varies linearly with enzyme concentrations in the presence of excess substrate. The Michaelis-Menten constant is 2×10^<-4>M.
社団法人日本薬学会の論文
1960-03-25