Howard P C, Reed K A, Koop D R
Department of Environmental Health Sciences, School of Medicine, Case Western Reserve University, Cleveland, Ohio 44106.
Cancer Res. 1988 Aug 1;48(15):4261-5.
Rabbit liver (male) microsomal metabolism of 10 microM [4,5,9,10-3H]-1-nitropyrene (1NP) was investigated. The total metabolism was not appreciably different with rates of 4.44 +/- 0.45, 3.98 +/- 0.19, 3.90 +/- 0.16, and 3.75 +/- 0.27 nmol/min/mg protein, respectively, for microsomes from phenobarbital, Aroclor-1254, ethanol-treated, and untreated rabbits. However, a more noticeable difference was found in the formation of specific metabolites. Phenobarbital treatment induced changes which favored 1-nitropyrene-3-ol formation, and Aroclor-1254 and ethanol-induced changes which favored 1-nitropyren-6-ol and 1-nitropyren-8-ol formation. 1NP was incubated with untreated microsomes and alpha-naphthoflavone, an inhibitor of rabbit cytochrome P-450 form 6 at low concentrations (less than 1 microM), and an activator of form 3c at high concentrations. The presence of alpha-naphthoflavone changed the profile of metabolites while not affecting the total metabolism. Using purified isozymes of rabbit P-450, we found the constitutive form 3b metabolized 1NP at the highest rate with a catalytic activity of 26.8 nmol/min/nmol P-450. Forms 2 and 6 exhibited rates of 2 and 2.2 nmol/min/nmol P-450. Forms 3a, 3c, and 4 had rates about 50- to 300-fold lower than form 3b. High performance liquid chromatography was used to identify the metabolites when the incubations were carried out in the presence of purified rabbit epoxide hydrolase. With form 6, 54% of the metabolites were accounted for as 1-nitropyren-3-ol, while with form 3b, 73% of the metabolites were 1-nitropyren-6-ol and 1-nitropyren-8-ol. The K-region dihydrodiols were formed by forms 2 and 3b, but not by forms 3c or 6. These results demonstrate that 1NP is a preferential substrate for form 3b, and that a preponderance of the metabolism with untreated rabbit liver microsomes can be attributed to this isozyme.
研究了10微摩尔[4,5,9,10-³H]-1-硝基芘（1NP）在雄性兔肝脏微粒体中的代谢情况。对于来自苯巴比妥、多氯联苯混合物1254、乙醇处理和未处理兔子的微粒体，总代谢率分别为4.44±0.45、3.98±0.19、3.90±0.16和3.75±0.27纳摩尔/分钟/毫克蛋白质，差异不明显。然而，在特定代谢产物的形成上发现了更显著的差异。苯巴比妥处理诱导的变化有利于1-硝基芘-3-醇的形成，多氯联苯混合物1254和乙醇诱导的变化有利于1-硝基芘-6-醇和1-硝基芘-8-醇的形成。1NP与未处理的微粒体以及α-萘黄酮一起孵育，α-萘黄酮在低浓度（小于1微摩尔）时是兔细胞色素P-450 6型的抑制剂，在高浓度时是3c型的激活剂。α-萘黄酮的存在改变了代谢产物的分布，同时不影响总代谢。使用纯化的兔P-450同工酶，我们发现组成型3b型以最高速率代谢1NP，催化活性为26.8纳摩尔/分钟/纳摩尔P-450。2型和6型的速率分别为2和2.2纳摩尔/分钟/纳摩尔P-450。3a、3c和4型的速率比3b型低约50至300倍。当在纯化的兔环氧化物水解酶存在下进行孵育时，使用高效液相色谱法鉴定代谢产物。对于6型，54%的代谢产物为1-硝基芘-3-醇，而对于3b型，73%的代谢产物为1-硝基芘-6-醇和1-硝基芘-8-醇。K-区域二氢二醇由2型和3b型形成，但不由3c型或6型形成。这些结果表明1NP是3b型的优先底物，并且未处理的兔肝脏微粒体的大部分代谢可归因于这种同工酶。
