Holm K A, Engell R J, Kupfer D
Arch Biochem Biophys. 1985 Mar;237(2):477-89. doi: 10.1016/0003-9861(85)90301-7.
The effects of methylcholanthrene (MC) treatment of male rats on the regioselectivity of hydroxylation of prostaglandins E1 and E2 (PGE1 and PGE2) by liver microsomes, supplemented with NADPH or H2O2, was examined. In the presence of NADPH, control microsomes catalyzed the hydroxylation at omega-1 (C19) and at omega-(C20) sites with minimal formation of novel monohydroxy metabolites of PGE1 and PGE2, referred to as compounds X1 and X2, respectively. Similarly, H2O2 supported the 19-hydroxylation and the formation of compounds X1 and X2, but yielded only minimal amounts of 20-hydroxy products. With NADPH, MC-treated microsomal incubations demonstrated only minor quantitative change in the 19- and 20-hydroxylation as compared with controls, but showed a 7- to 11-fold increase in formation of compound X1 and a 10-fold increase in formation of X2. By contrast with H2O2, MC-treatment increased by about 3-fold the 19- and 20-hydroxylation of PGE1 and by 35- to 46-fold the formation of X1; similarly, there was an approximate 2-fold increase in 19- and 20-hydroxylation of PGE2 and a 10-fold increase in formation of X2. These findings suggest that several monooxygenases are involved in catalyzing the hydroxylation at the various sites of the PGE molecule. Inhibitors of monooxygenases (SKF 525A, alpha-naphthoflavone, and imidazole derivatives) provided further evidence that the hydroxylation at the three sites of PGEs is catalyzed by different P-450 monooxygenases. It is striking that the inhibitors had a much lesser effect on the 20-hydroxylation of PGE1 as compared with other sites of hydroxylation. Structural identification of compounds X1 and X2 was elucidated as follows. Resistance of the PGB derivative of X1 to periodate oxidation and mass fragmentation analysis of the t-butyldimethylsilyl ether methyl ester, placed the hydroxylation at C17 or C18. Finally, mass fragmentation of trimethylsilyl ether methyl ester PGB derivatives of X1 and X2 provided conclusive evidence that X1 and X2 are 18-hydroxy-PGE1 and 18-hydroxy-PGE2, respectively. The above findings indicate that the high regioselectivity of hydroxylation of PGE1 and PGE2, resulting in the formation of 18-hydroxy-PGE1 and 18-hydroxy-PGE2, respectively, is catalyzed by P-450 isozyme(s) which are induced by MC, possibly by P-450c.
研究了用甲基胆蒽（MC）处理雄性大鼠后，其肝脏微粒体在补充烟酰胺腺嘌呤二核苷酸磷酸（NADPH）或过氧化氢（H2O2）的情况下，对前列腺素E1和E2（PGE1和PGE2）羟基化区域选择性的影响。在NADPH存在的情况下，对照微粒体催化在ω-1（C19）和ω-（C20）位点的羟基化，PGE1和PGE2新的单羟基代谢产物（分别称为化合物X1和X2）的生成量极少。同样，H2O2支持19-羟基化以及化合物X1和X2的生成，但仅产生极少量的20-羟基产物。对于NADPH，与对照相比，经MC处理的微粒体孵育在19-和20-羟基化方面仅显示出微小的定量变化，但化合物X1的生成量增加了7至11倍，X2的生成量增加了10倍。与H2O2相比，MC处理使PGE1的19-和20-羟基化增加了约3倍，X1的生成量增加了35至46倍；同样，PGE2的19-和20-羟基化增加了约2倍，X2的生成量增加了10倍。这些发现表明，几种单加氧酶参与催化PGE分子不同位点的羟基化。单加氧酶抑制剂（SKF 525A、α-萘黄酮和咪唑衍生物）提供了进一步的证据，表明PGEs三个位点的羟基化是由不同的P-450单加氧酶催化的。令人惊讶的是，与其他羟基化位点相比，抑制剂对PGE1的20-羟基化影响要小得多。化合物X1和X2的结构鉴定如下。X1的PGB衍生物对高碘酸盐氧化具有抗性，以及对叔丁基二甲基甲硅烷基醚甲酯的质谱碎裂分析，确定羟基化位于C17或C18。最后，X1和X2的三甲基甲硅烷基醚甲酯PGB衍生物的质谱碎裂提供了确凿证据，表明X1和X2分别是18-羟基-PGE1和18-羟基-PGE2。上述发现表明，PGE1和PGE2羟基化的高区域选择性分别导致18-羟基-PGE1和18-羟基-PGE2的形成，是由MC诱导的P-450同工酶催化的，可能是由P-450c催化的。
