Kozumbo W J, Rubin R J
Department of Medicine, University of North Carolina, Chapel Hill 27599.
J Toxicol Environ Health. 1991 May;33(1):29-46. doi: 10.1080/15287399109531503.
As an active ingredient in insect repellents, dimethyl phthalate (DMP) had previously been shown to produce chromosomal aberrations in the livers of rats following subchronic application of the phthalate to skin. When we tested DMP in the Ames mutagenesis assay, it produced in bacterial tester strain TA100 (but not TA98) a dose-related mutagenic response that was abolished by NAD- and NADP-independent metabolism associated with rat liver microsomal preparations (S9). In a host-mediated mutagenesis assay, rats were injected ip with DMP (2 g/kg body weight); urine was collected for 24 h, extracted, and analyzed for mutagenic activity and phthalic acid-containing derivatives. The extracted urine was not mutagenic to TA100 and contained an equivalent of 1.96 mg phthalate/ml urine. More than 97% of the phthalic acid-containing derivatives present in the extracted urine consisted of the nonmutagenic metabolite of DMP, monomethyl phthalate (MMP). In vitro experiments showed that rat liver homogenates hydrolyzed 93% of carbonyl-labeled 14C-DMP (7.7 mM) to MMP in 2 h and bound 0.07 nmol of [14C]phthalate/mg liver macromolecules. By contrast, rat epidermal homogenates metabolized only 5% and bound 38-fold higher levels of carbonyl-labeled 14C-DMP (2.66 nmol/mg of macromolecules), with no detectable binding to nucleic acids. Compared to epidermis and plasma, liver had a fivefold higher rate of DMP monoesterase activity (1240 nmol/h/mg protein), which, when inhibited by 67%, resulted in a 4.4-fold increase in phthalate-bound hepatic macromolecules (0.31 vs. 0.07 nmol of carbonyl-labeled 14C-DMP/mg macromolecules). In addition to MMP, formaldehyde was produced during the metabolism of DMP by liver. When ethanol was used to inhibit the oxidation of DMP-derived methanol by hepatic homogenates, there resulted a 74% reduction in the accumulation of formaldehyde and similar reductions of 71 and 73% in the binding of methyl-labeled 14C-DMP to nucleic acids and macromolecules. (Methyl-labeled, unlike carbonyl-labeled, 14C-DMP yields a 14C-labeled methanol when hydrolyzed.) These results indicate that the DMP diester is a weak bacterial mutagen, which binds to epidermal and hepatic macromolecules other than nucleic acids, and that although the rapid hepatic metabolism of DMP to its monoester (MMP) and methanol affords protection against higher levels of phthalate binding as well as against DMP-induced bacterial mutagenesis, it also oxidizes DMP-derived methanol to formaldehyde, a metabolite that binds macromolecules, including nucleic acids.
邻苯二甲酸二甲酯（DMP）作为驱虫剂中的一种活性成分，先前的研究表明，将该邻苯二甲酸盐经皮肤亚慢性给药后，可在大鼠肝脏中产生染色体畸变。当我们在艾姆斯诱变试验中测试DMP时，它在细菌测试菌株TA100（而非TA98）中产生了剂量相关的诱变反应，而与大鼠肝脏微粒体制剂（S9）相关的不依赖NAD和NADP的代谢可消除这种反应。在宿主介导的诱变试验中，给大鼠腹腔注射DMP（2 g/kg体重）；收集24小时尿液，进行提取，并分析其诱变活性和含邻苯二甲酸的衍生物。提取的尿液对TA100无诱变作用，且每毫升尿液中含有相当于1.96 mg的邻苯二甲酸盐。提取尿液中存在的超过97%的含邻苯二甲酸的衍生物由DMP的非诱变代谢产物邻苯二甲酸单甲酯（MMP）组成。体外实验表明，大鼠肝脏匀浆在2小时内将93%的羰基标记的14C-DMP（7.7 mM）水解为MMP，并使每毫克肝脏大分子结合0.07 nmol的[14C]邻苯二甲酸盐。相比之下，大鼠表皮匀浆仅代谢5%，并使羰基标记的14C-DMP（2.66 nmol/mg大分子）的结合水平高38倍，且未检测到与核酸的结合。与表皮和血浆相比，肝脏的DMP单酯酶活性高五倍（1240 nmol/h/mg蛋白质），当该活性被抑制67%时，邻苯二甲酸盐结合的肝脏大分子增加4.4倍（0.31对0.07 nmol羰基标记的14C-DMP/mg大分子）。除MMP外，肝脏在DMP代谢过程中还产生甲醛。当使用乙醇抑制肝脏匀浆对DMP衍生甲醇的氧化时，甲醛的积累减少74%，甲基标记的14C-DMP与核酸和大分子的结合也分别减少71%和73%。（与羰基标记不同，甲基标记的14C-DMP水解时会产生14C标记的甲醇。）这些结果表明，DMP二酯是一种弱细菌诱变剂，它与除核酸外的表皮和肝脏大分子结合，并且尽管DMP在肝脏中快速代谢为其单酯（MMP）和甲醇可防止较高水平的邻苯二甲酸盐结合以及DMP诱导的细菌诱变，但它也将DMP衍生的甲醇氧化为甲醛，甲醛是一种可与包括核酸在内的大分子结合的代谢产物。
