Hinchman C A, Matsumoto H, Simmons T W, Ballatori N
Department of Biophysics, University of Rochester School of Medicine, New York 14642.
J Biol Chem. 1991 Nov 25;266(33):22179-85.
Because of the low hepatic activity of gamma-glutamyl-transferase in the rat, the liver is generally considered to play only a minor role in the degradation of glutathione conjugates, a limiting step in mercapturic acid formation. Recent findings indicate, however, that the liver has a prominent role in glutathione catabolism, particularly in species other than rat. To examine the contributions of liver to mercapturic acid biosynthesis, mercapturate formation was compared in isolated perfused livers from rats and guinea pigs dosed with either 0.3 or 3.0 mumol of 1-chloro-2,4-dinitrobenzene (CDNB). Chemically synthesized glutathione conjugate, mercapturic acid, and intermediary metabolites of CDNB were used as standards in the high performance liquid chromatography analysis of bile and perfusate samples. Biliary excretion accounted for almost all of the recovered metabolites. A marked species difference was observed in the pattern of CDNB metabolism. Rat livers dosed with 0.3 mumol of CDNB excreted 55% of total biliary metabolites as the glutathione conjugate and 8.2% as the mercapturic acid, whereas guinea pig livers excreted only 4.8% as the glutathione conjugate and 47% as the mercapturate. Mercapturic formation was also dose-dependent, with a larger fraction formed at the 0.3- versus the 3.0-mumol dose (8.2 versus 3.7% in the rat; 47 versus 19% in the guinea pig). Hepatic conversion of the glutathione conjugate to the mercapturic acid was markedly inhibited in both species after retrograde intrabiliary infusion of acivicin, an inhibitor of gamma-glutamyltransferase activity. These findings provide direct evidence for intrahepatic biosynthesis of mercapturic acids. Thus, glutathione conjugates synthesized within hepatocytes are secreted into bile and broken down to cysteine conjugates; the latter are then presumably reabsorbed by the liver, N-acetylated to form the mercapturic acid and re-excreted into bile.
由于大鼠肝脏中γ-谷氨酰转移酶的活性较低，肝脏通常被认为在谷胱甘肽结合物的降解中仅起次要作用，而谷胱甘肽结合物的降解是巯基尿酸形成的限速步骤。然而，最近的研究结果表明，肝脏在谷胱甘肽分解代谢中起重要作用，尤其是在大鼠以外的物种中。为了研究肝脏对巯基尿酸生物合成的贡献，比较了分别给予0.3或3.0 μmol 1-氯-2,4-二硝基苯（CDNB）的大鼠和豚鼠离体灌注肝脏中巯基尿酸的形成情况。化学合成的谷胱甘肽结合物、巯基尿酸和CDNB的中间代谢产物被用作胆汁和灌注液样品高效液相色谱分析的标准品。胆汁排泄几乎占所有回收代谢产物的全部。在CDNB代谢模式中观察到明显的物种差异。给予0.3 μmol CDNB的大鼠肝脏将55%的总胆汁代谢产物以谷胱甘肽结合物的形式排泄，8.2%以巯基尿酸的形式排泄，而豚鼠肝脏仅以4.8%的谷胱甘肽结合物和47%的巯基尿酸形式排泄。巯基尿酸的形成也呈剂量依赖性，在0.3 μmol剂量下形成的比例高于3.0 μmol剂量（大鼠中分别为8.2%和3.7%；豚鼠中分别为47%和19%）。在逆行胆管内注入γ-谷氨酰转移酶活性抑制剂阿西维辛后，两种物种中谷胱甘肽结合物向巯基尿酸的肝脏转化均受到显著抑制。这些发现为巯基尿酸的肝内生物合成提供了直接证据。因此，肝细胞内合成的谷胱甘肽结合物分泌到胆汁中并分解为半胱氨酸结合物；后者可能随后被肝脏重新吸收，N-乙酰化形成巯基尿酸并重新排泄到胆汁中。
