Magdalou J, Hammock B D
Department of Entomology, University of California, Davis 95616.
Toxicol Appl Pharmacol. 1987 Dec;91(3):439-49. doi: 10.1016/0041-008x(87)90065-2.
The transformation of the herbicide tridiphane (Tandem, Dowco 356, 2-(3,5-dichlorophenyl)-2(2,2,2-trichloroethyl)oxirane by the epoxide-metabolizing enzymes, epoxide hydrolases (EH) and glutathione S-transferases (GST), was investigated in mouse liver microsomes and cytosol. The microsomal EH catalyzed the formation of tridiphane diol. The production of this metabolite was prevented by cyclohexene oxide at 1 mM, a known inhibitor of microsomal EH. The structure of the diol was verified by comparison of retention time or Rf of the compound with those of an authentic standard using gas-liquid chromatography or thin-layer chromatography techniques. The diol formed a diester with 1-butane boronic acid or an aldehyde with lead tetraacetate. Mass spectral analysis supported the structural assignment. After optimization of the assay conditions, kinetic constants for the hydration of tridiphane by the microsomal EH were determined (Km = 65 microM and Vmax = 0.9 nmol/min/mg protein). Dietary exposure of mice to the hypolipidemic drug clofibrate at a dose of 0.5% (w/w) for 2 weeks increased by 173% the metabolism of tridiphane to tridiphane diol by the microsomal fraction. No diol could be detected following incubation of tridiphane with the cytosolic EH, even after induction by clofibrate. Tridiphane was also a substrate for GST, but administration of clofibrate did not change the specific activity for the formation of the glutathione conjugate. The herbicide was a rather weak inhibitor of the microsomal EH and the cytosolic GST activities measured with cis-stilbene oxide and trans-stilbene oxide as substrates with I50's of 3.0 x 10(-5) and 1.8 x 10(-4)M, respectively. Tridiphane diol was a poor inhibitor of the enzymes studied, and the glutathione conjugate of tridiphane caused marked inhibition of only the GST activity (I50, 2.0 x 10(-5)M). By contrast the activity of cytosolic EH (trans-stilbene oxide) was relatively insensitive to the addition of tridiphane or of tridiphane metabolites.
在小鼠肝脏微粒体和胞液中，研究了除草剂敌稗（Tandem，陶氏益农356，2-(3,5-二氯苯基)-2-(2,2,2-三氯乙基)环氧乙烷）被环氧化物代谢酶环氧水解酶（EH）和谷胱甘肽S-转移酶（GST）的转化情况。微粒体EH催化生成敌稗二醇。1 mM的环氧环己烷可抑制微粒体EH的活性，从而阻止该代谢产物的生成，环氧环己烷是一种已知的微粒体EH抑制剂。通过气液色谱或薄层色谱技术，将该化合物的保留时间或比移值与标准品进行比较，从而验证了二醇的结构。二醇与1-丁烷硼酸形成二酯，或与四乙酸铅形成醛。质谱分析支持了结构鉴定。在优化分析条件后，测定了微粒体EH催化敌稗水化反应的动力学常数（Km = 65 μM，Vmax = 0.9 nmol/min/mg蛋白）。给小鼠喂食0.5%（w/w）的降血脂药物氯贝丁酯，持续2周，微粒体部分将敌稗代谢为敌稗二醇的能力提高了173%。即使在用氯贝丁酯诱导后，将敌稗与胞液EH一起孵育，也检测不到二醇。敌稗也是GST的底物，但给予氯贝丁酯并没有改变生成谷胱甘肽缀合物的比活性。以顺式氧化茋和反式氧化茋为底物测定时，该除草剂对微粒体EH和胞液GST活性的抑制作用较弱，其半数抑制浓度（I50）分别为3.0×10⁻⁵和1.8×10⁻⁴ M。敌稗二醇对所研究的酶的抑制作用较弱，敌稗的谷胱甘肽缀合物仅对GST活性有显著抑制作用（I50，2.0×⁻⁵ M）。相比之下，胞液EH（反式氧化茋）的活性对敌稗或敌稗代谢产物的添加相对不敏感。
