Satoh H, Gillette J R, Davies H W, Schulick R D, Pohl L R
Mol Pharmacol. 1985 Nov;28(5):468-74.
Four hours after the administration of halothane to phenobarbital-pretreated rats, subcellular fractions of liver were isolated and the proteins in the fractions were separated by sodium dodecyl sulfate polyacrylamide gel electrophoresis, transferred to nitrocellulose sheets, and immunochemically stained with anti-trifluoroacetylated antibodies. The microsomal fraction contained the highest level of trifluoroacetylated adducts. Its major trifluoroacetylated component was immunochemically identified as a phenobarbital-inducible form of cytochrome P-450 (54 kDa), whereas the other observed trifluoroacetylated protein fraction (59 kDa) was not identified. The plasma membrane fraction also contained a 54-kDa trifluoroacetylated adduct, which was immunochemically related to the 54-kDa cytochrome P-450. Microsomes from untreated rats that were administered halothane contained only the 59-kDa trifluoroacetylated protein fraction. The specificity of the immunochemical staining for the bound oxidative metabolite of halothane was confirmed by the finding that rats treated with deuterated halothane had considerably less stained liver proteins than did those treated with halothane. These results suggest that the CF3COX oxidative metabolite of halothane is so reactive that it binds predominantly to the cytochrome P-450 that produced it.
给预先用苯巴比妥处理的大鼠施用氟烷4小时后，分离肝脏的亚细胞组分，通过十二烷基硫酸钠聚丙烯酰胺凝胶电泳分离各组分中的蛋白质，转移至硝酸纤维素膜上，并用抗三氟乙酰化抗体进行免疫化学染色。微粒体组分中三氟乙酰化加合物的含量最高。其主要的三氟乙酰化成分经免疫化学鉴定为细胞色素P-450的苯巴比妥诱导型（54 kDa），而观察到的其他三氟乙酰化蛋白组分（59 kDa）未得到鉴定。质膜组分也含有一种54 kDa的三氟乙酰化加合物，其与54 kDa的细胞色素P-450存在免疫化学相关性。给予氟烷的未处理大鼠的微粒体仅含有59 kDa的三氟乙酰化蛋白组分。用氘代氟烷处理的大鼠肝脏蛋白染色明显少于用氟烷处理的大鼠，这一发现证实了对氟烷结合氧化代谢物的免疫化学染色的特异性。这些结果表明，氟烷的CF3COX氧化代谢物具有很高的反应活性，以至于它主要与产生它的细胞色素P-450结合。
