Goon D, Cheng X, Ruth J A, Petersen D R, Ross D
Molecular Toxicology and Environmental Health Sciences Program, School of Pharmacy, University of Colorado, Boulder 80309-0297.
Toxicol Appl Pharmacol. 1992 May;114(1):147-55. doi: 10.1016/0041-008x(92)90107-4.
The metabolism of trans,trans-muconaldehyde (MA), a highly reactive alpha,beta-unsaturated dialdehyde, was examined in vitro using purified yeast alcohol and aldehyde dehydrogenases (ADH and ALDH, respectively). In the presence of NAD(+)-fortified ALDH, the mono-oxidation product (acid/aldehyde) was the primary metabolite formed with trace amounts of the dioxidation product (trans,trans-muconic acid). In NADH-fortified reactions with ADH, both the mono- and direduction products (hydroxy/aldehyde and dihydroxy, respectively) were readily detected. Oxidation and reduction products of MA were formed in incubates containing both dehydrogenases together with either NAD+ or NADH. Unexpectedly, an additional metabolite was detected, which was a major product in both NAD(+)- and NADH-fortified systems containing ALDH and ADH in combination and whose formation could be inhibited by pyrazole (an ADH inhibitor). ALDH-mediated oxidation of a synthetic standard of the hydroxy/aldehyde derivative of MA resulted in formation of this new metabolite, which was also a major product formed by rat hepatocytes incubated with MA. Using HPLC/photodiode array detection, the new metabolite was found to cochromatograph and have a uv spectrum identical to that of a synthetic standard of the hydroxy/acid derivative of MA. The metabolite was confirmed as the hydroxy/acid derivative of MA after preparative HPLC, TMS derivatization, and GC/MS analysis. The hydroxy/acid metabolite was not formed during ADH-mediated reduction of the mono-oxidation product of MA, suggesting that this metabolite was formed by yeast dehydrogenases via a primary reduction of MA and subsequent oxidation of the hydroxy/aldehyde to the hydroxy/acid. These data show that the hydroxy/acid derivative is a novel metabolite of MA, which arises from the interaction of both oxidative and reductive routes of metabolism.
使用纯化的酵母醇脱氢酶和醛脱氢酶（分别为ADH和ALDH）在体外研究了反式、反式粘康醛（MA）的代谢，MA是一种高反应性的α,β-不饱和二醛。在添加了NAD⁺的ALDH存在下，单氧化产物（酸/醛）是形成的主要代谢产物，同时伴有痕量的二氧化产物（反式、反式粘康酸）。在添加了NADH的ADH反应中，单还原产物和双还原产物（分别为羟基/醛和二羟基）都很容易被检测到。MA的氧化和还原产物在同时含有两种脱氢酶以及NAD⁺或NADH的孵育体系中形成。出乎意料的是，检测到了一种额外的代谢产物，它是在同时含有ALDH和ADH且添加了NAD⁺和NADH的体系中的主要产物，其形成可被吡唑（一种ADH抑制剂）抑制。ALDH介导的MA羟基/醛衍生物合成标准品的氧化导致了这种新代谢产物的形成，它也是与MA孵育的大鼠肝细胞形成的主要产物。使用HPLC/光电二极管阵列检测，发现该新代谢产物与MA羟基/酸衍生物的合成标准品共色谱，且紫外光谱相同。经过制备型HPLC、TMS衍生化和GC/MS分析后，该代谢产物被确认为MA的羟基/酸衍生物。MA单氧化产物在ADH介导的还原过程中未形成羟基/酸代谢产物，这表明该代谢产物是由酵母脱氢酶通过MA的初次还原以及随后羟基/醛氧化为羟基/酸而形成的。这些数据表明，羟基/酸衍生物是MA的一种新型代谢产物，它源于代谢的氧化和还原途径的相互作用。
