FORMATION AND REACTIVITY OF TOXIC QUINONE METHIDES

羟基化芳烃和多羟基芳烃广泛存在于 环境环境，尤指作为可食用植物的成分。其中许多 酚类化合物可在酶作用下氧化成亲电性的苯二酚 但是，除少数例外情况外，参与甲基苯醌的 在调节取代苯酚的不利影响方面尚未得到 考虑过了。细胞色素P450和过氧化物酶活性均可催化 所以细胞成分的烷基化反应是由 这一途径可能发生在许多不同的组织中。初步数据 在对苯二酚的形成和反应活性方面表现出很大的差异 甲基是由于酚类前体的结构所致。现在 应用解决了苯酚结构和P450之间的关系- 催化的对苯二酚的形成、反应性和细胞毒性。至 为实现这些目标，提出了以下具体目标。(1) 测定化学结构对酚类化合物氧化的影响 从化合物到对苯二酚。(A)特定原氧的影响- 含有取代基和不饱和对烷基取代基的将是 检查，因为这些通常在自然产生的酚中发现，以及 预计会影响对苯二酚的形成和反应活性。这些 研究范围将扩大到调查从 致突变和非致突变的黄酮类化合物。(B)基质的影响 将研究细胞色素P450同工酶的结构和选择性，如 先前的研究结果表明，在催化作用中存在同工酶的差异。 形成了对苯二酚的甲烷。关于这种氧化作用的机制信息 途径将获得与氚标记的类似物和自由基 食腐动物。为了与P450活性进行比较，苯醌甲醚的形成 通过一个模型还将研究过氧化物酶系统。(2)调查 苯二酚甲醚结构对亲电活性的影响。 对苯二酚与水、谷胱甘肽、 将测定亲核氨基酸和嘌呤脱氧核苷。 这一数据将与苯二酚甲醚的结构相关联，从而推断 其反应性背后的一般原则，并深入了解 细胞内结合选择性。对苯二酚的选择性反应 含有纯蛋白质和DNA的甲烷也将被研究以产生 关于特定残留物的定性和定量数据 受影响。(3)研究甲基苯二酚在调节细胞生长发育中的作用。 酚类化合物的毒性。分离的大鼠肝细胞将被孵育 用取代的苯酚产生不同种类的苯醌甲基 与亲核试剂的反应。甲基苯二酚的形成效应 对细胞活性和与毒性相关的生化参数的影响 要下定决心。蛋白质和蛋白质与甲基苯醌的共价结合 将对DNA进行研究，以确定细胞内的烷基化位点。 由此产生的数据将为我们提供对 酚类化合物细胞毒性中的甲基苯醌。

Hydroxylated and polyhydroxylated aromatics are widespread in the environment, especially as constituents of edible plants. Many of these phenolic compounds can be oxidized enzymatically to electrophilic quinone methides, but, with a few exceptions, the involvement of quinone methides in mediating the adverse effects of substituted phenols has not been considered. Both cytochrome P450 and peroxidase activity can catalyze quinone methide formation, so the alkylation of cellular components by this pathway may occur in many different tissues. Preliminary data demonstrate wide variation both in the formation and reactivity of quinone methides due to the structure of the phenolic precursor. The present application addresses relationships between phenol structure and P450- catalyzed quinone methide formation, reactivity, and cytotoxicity. To accomplish these goals, the following specific aims are proposed. (1) Determine the influence of chemical structure on the oxidation of phenolic compounds to quinone methides. (a) The effects of specific ortho oxygen- containing substituents, and unsaturated para alkyl substituents will be examined, as these are commonly found in naturally-occurring phenols, and are expected to affect quinone methide formation and reactivity. These studies will be extended to investigate quinone methide formation from mutagenic and non-mutagenic flavonoids. (b) Influences of substrate structure on cytochrome P450 isozyme selectivities will be studied, as previous results have demonstrated isozymic differences in the catalysis of quinone methide formation. Mechanistic information on this oxidative pathway will be obtained with deuterium-labeled analogs and free radical scavengers. For comparison with P450 activity, quinone methide formation by a model peroxidase system also will be investigated. (2) Investigate the effects of quinone methide structure on electrophilic reactivity. Rates of reactions of quinone methides with water, glutathione, nucleophilic amino acids, and purine deoxynucleosides will be measured. This data will be correlated with quinone methide structures to deduce general principles underlying their reactivity, and gain insight into intracellular binding selectivities. Reactions of selected quinone methides with pure proteins and DNA will also be investigated to generate qualitative and quantitative data concerning the particular residues affected. (3) Investigate the roles of quinone methides in mediating the toxicity of phenolic compounds. Isolated rat hepatocytes will be incubated with substituted phenols which produce quinone methides of varying reactivities with nucleophiles. The effects of quinone methide formation on cell viability, and on biochemical parameters related to toxicity will be determined. The covalent binding of quinone methides to proteins and DNA will be investigated to determine intracellular sites of alkylation. The resulting data will provide substantial insight into the roles of quinone methides in the cytotoxicity of phenolic compounds.