Yang X, Kulkarni A P
Florida Toxicology Research Center, Department of Environmental and Occupational Health, College of Public Health, University of South Florida, Tampa 33612-3805, USA.
J Biochem Mol Toxicol. 1998;12(3):175-83. doi: 10.1002/(sici)1099-0461(1998)12:3<175::aid-jbt6>3.0.co;2-i.
A hypothesis that lipoxygenase may mediate N-dealkylation of xenobiotics was investigated using the prototype drug aminopyrine and soybean lipoxygenase as a model enzyme in the presence of hydrogen peroxide. Formaldehyde production as a result of N-demethylation of aminopyrine exhibited pH optimum of 6.5. The reaction was dependent on the incubation time, amount of enzyme, and concentration of aminopyrine and hydrogen peroxide. Under the experimental conditions employed, the specific activity for N-demethylation of aminopyrine was found to be 823 +/- 93 nmoles per min/mg protein or 89 +/- 10 nmoles per min/nmole of enzyme. The reaction was significantly inhibited by nordihydroguaiaretic acid and gossypol, the classical inhibitors of lipoxygenase. Spectrophotometric analyses indicated the generation of a nitrogen-centered free-radical cation as the initial oxidation product of aminopyrine. The rate of accumulation of this radical species was also dependent on pH, the amount of enzyme, and concentration of aminopyrine and hydrogen peroxide. The radical production was markedly suppressed by ascorbate, glutathione, and dithiothreitol in a concentration-dependent manner. Preliminary data gathered for the oxidation of other chemicals indicated that the lipoxygenase exhibits a unique substrate specificity. Collectively, the evidence presented suggests for the first time that lipoxygenase pathway may be involved in N-demethylation of aminopyrine and other chemicals.
使用原型药物氨基比林和大豆脂氧合酶作为模型酶，在过氧化氢存在的情况下，研究了脂氧合酶可能介导外源性物质N-脱烷基化的假说。氨基比林N-去甲基化产生甲醛的反应最适pH为6.5。该反应取决于孵育时间、酶量、氨基比林和过氧化氢的浓度。在所采用的实验条件下，发现氨基比林N-去甲基化的比活性为每分钟每毫克蛋白质823±93纳摩尔或每分钟每纳摩尔酶89±10纳摩尔。该反应被脂氧合酶的经典抑制剂去甲二氢愈创木酸和棉酚显著抑制。分光光度分析表明，作为氨基比林初始氧化产物产生了以氮为中心的自由基阳离子。这种自由基的积累速率也取决于pH、酶量、氨基比林和过氧化氢的浓度。抗坏血酸、谷胱甘肽和二硫苏糖醇以浓度依赖的方式显著抑制自由基的产生。对其他化学物质氧化收集的初步数据表明，脂氧合酶表现出独特的底物特异性。总体而言，所提供的证据首次表明脂氧合酶途径可能参与氨基比林和其他化学物质的N-去甲基化。
