BIOACTIVATION OF DIETARY PHENOLS BY HEMOPROTEINS

人们对自然产生的酚类非常感兴趣，因为 它们作为抗氧化剂和癌症的巨大潜力 预防。然而，不幸的是，这些物质也会导致 自由基的形成、突变、肿瘤和细胞毒性。 酚的各种生物效应的潜在机制必须是 在此之前阐明了影响有益成分混合的因素 行列式性质可以得到更充分的理解。新陈代谢 这些化合物被细胞色素P450激活涉及众所周知的 反应性代谢物，如环氧化物和苯二酚，但苯氧基 由P450催化的单电子氧化产生的分子 氧对羟基形成氢过氧环己二烯酮， 在化学文献中被称为“过氧喹诺酮”。这部小说 途径还没有被广泛认识，因为过氧喹酚不 积累，但在生物系统中迅速降解。在.期间 目前的资助期，已经得到了证明，使用了一系列 烷基取代苯酚，过氧喹酚的形成发生在 在肝微粒体和分离的肝细胞中均有明显的表达。 这一途径的相对贡献受底物的影响。 结构和P450同工酶。过氧喹酚可被快速降解 沿着复杂途径的各种血红蛋白，形成自由基和 亲电产物，选择性氧化蛋氨酸残基完整 蛋白质，并严重损害细胞。数据有力地支持了继续 过氧喹酚在生物反应中的作用研究 酚类化合物的性质。我们现在建议将这项工作从 相对简单的烷基酚到更复杂的化合物，含有 天然酚类共有的结构特征。具体的 目的如下：(1)研究过氧喹酚的贡献 形成对单个核细胞微粒体和细胞代谢的影响 含有邻甲氧基或间羟基取代基的酚类，以及 具有甲基取代基的多核酚。确定雌激素是否 通过过氧喹酚途径氧化。(2)确定形成的产品和 血红素蛋白降解过氧喹酚的途径。 用肝细胞色素P450和定点突变体获得的数据 P450cam和肌红蛋白的相互作用将阐明血红蛋白的作用机制。 提供有关过氧喹酚生物活性产品的信息。 (3)过氧喹酚对大鼠肝细胞和小鼠肝细胞的影响 角质形成细胞。确定过氧喹酚介导的后果 非血红素蛋白中蛋氨酸残基的氧化，并确定是否 DNA结合的金属离子将过氧喹酚还原为破坏 DNA实现这些具体目标将大大增强我们的 对这一新的氧化途径的认识及其潜在的调节作用 某些酚类化合物的生物效应。

There is a great deal of interest in naturally occurring phenols because of their enormous potential as agents for antioxidant therapy and cancer prevention. Unfortunately, however, these substances also lead to the formation of free radicals, mutations, tumors, and cytotoxicity. Mechanisms underlying the various biological effects of phenols must be elucidated before those factors which influence the blend of beneficial and determinant properties can be more fully understood. The metabolic activation of these compounds by cytochrome P450 involves well known reactive metabolites such as epoxides and quinones, but phenoxy radicals produced by P450-catalyzed one-electron oxidation can add molecular oxygen para to the hydroxy group to form hydroperoxy-cyclohexadienones, referred to in the chemical literature as 'peroxyquinols.' This novel pathway has not been widely recognized because peroxyquinols do not accumulate, but are rapidly degraded in biological systems. During the current funding period, it has been demonstrated, using a series of alkyl-substituted phenols, that peroxyquinol formation occurs to a significant extent both in liver microsomes and in isolated hepatocytes. The relative contribution of this pathway is influenced by the substrate structure and P450 isozyme. Peroxyquinols are rapidly degraded by various hemeproteins along complex pathways, form radicals and electrophilic products, selectively oxidize methionine residues of intact proteins, and severely damage cells. The data strongly support continued studies on the roles of peroxyquinols in mediating the biological properties of phenolic compounds. We now propose to extend the work from relatively simple alkylphenols to more complex compounds that contain structural features common to naturally occurring phenols. The specific aims are as follows: (1) Investigate the contribution of peroxyquinol formation to the microsomal and cellular metabolism of mononuclear phenols containing ortho-methoxy or meta-hydroxy substituents, and polynuclear phenols with methyl substituents. Determine if estrogens are oxidized via the peroxyquinol pathway. (2) Identify products formed and pathways involved in the degradation of peroxyquinols by hemeproteins. Data obtained with hepatic cytochromes P450, and site-specific mutants of P450cam and myoglobin will elucidate hemeprotein mechanisms, and provide information on biologically active products of peroxyquinols. (3) Study the effects of peroxyquinols on rat hepatocyte and murine keratinocytes. Determine the consequences of peroxyquinol-mediated oxidations of methionine residues in non-heme proteins, and determine if DNA-bound metal ions reduce peroxyquinols to radical species that damage DNA. Accomplishing these specific aims will significantly enhance our knowledge of this novel oxidative pathway and its potential to mediate the biological effects of certain phenolic compounds.