Shimada T, Sawabe Y, Nakano Y
Arch Toxicol. 1985 Oct;58(1):20-6. doi: 10.1007/BF00292611.
Incubation of 14C-labeled 3,4,3',4'-tetrachlorobiphenyl (TCB) with a reconstituted monooxygenase system containing cytochrome P-450 isolated from 3-methylcholanthrene (MC)-treated rats caused an increase in incorporation of radioactivity to added rat erythrocytes and resulted in covalent binding of reactive metabolites to hemoglobin. Both increases in incorporation and covalent binding to the proteins were dependent on incubation time and required cytochrome P-450-linked monooxygenase activity. In the absence of erythrocytes, the reactive TCB metabolites bound to cytochrome P-450, NADPH-cytochrome P-450 reductase, and high-molecular weight aggregates, as determined by sodium dodecyl sulfate (SDS)-polyacrylamide gel electrophoresis, whereas inclusion of the cells in the reaction mixture caused marked alkylation of hemoglobin and a decrease in binding to the electron transfer proteins. Several metabolites other than protein-bound metabolites were detected by thin-layer chromatography; some were found to be incorporated into the erythrocytes while others were not. These results suggest that several TCB metabolites formed by cytochrome P-450 can interact with rat erythrocytes in vitro, and that these interactions may be responsible for the persistence of TCB metabolites in blood observed in vivo.
将14C标记的3,4,3',4'-四氯联苯（TCB）与一种重组单加氧酶系统一起温育，该系统含有从经3-甲基胆蒽（MC）处理的大鼠中分离出的细胞色素P-450，结果使添加的大鼠红细胞中放射性的掺入增加，并导致活性代谢产物与血红蛋白发生共价结合。掺入量的增加和与蛋白质的共价结合都依赖于温育时间，并且需要细胞色素P-450相关的单加氧酶活性。在没有红细胞的情况下，通过十二烷基硫酸钠（SDS）-聚丙烯酰胺凝胶电泳测定，活性TCB代谢产物与细胞色素P-450、NADPH-细胞色素P-450还原酶以及高分子量聚集体结合，而在反应混合物中加入细胞则导致血红蛋白发生明显烷基化，并减少了与电子传递蛋白的结合。通过薄层色谱法检测到了除与蛋白质结合的代谢产物之外的几种代谢产物；发现其中一些被掺入红细胞，而另一些则没有。这些结果表明，细胞色素P-450形成的几种TCB代谢产物在体外可与大鼠红细胞相互作用，并且这些相互作用可能是体内观察到的TCB代谢产物在血液中持续存在的原因。
