Tretyakova N, Lin Y, Sangaiah R, Upton P B, Swenberg J A
Department of Environmental Sciences and Engineering, University of North Carolina, Chapel Hill 27599-7400, USA.
Carcinogenesis. 1997 Jan;18(1):137-47. doi: 10.1093/carcin/18.1.137.
3,4-Epoxy-1-butene (EB) is the major mutagenic metabolite of butadiene (BD), an important industrial chemical classified as a probable human carcinogen. Although the mechanism of carcinogenicity of EB is not known, its reactions with nucleophilic sites of DNA giving pro-mutagenic lesions are likely to constitute the early crucial step in multistage carcinogenesis. This study was conducted to characterize the adducts formed from reactions of EB with the most nucleophilic DNA nucleobases, adenine (Ade) and guanine (Gua), as free nucleobases, 2'-deoxyribonucleosides and constituents of calf thymus DNA (CT DNA) in order to provide insight into the nature of DNA modification by EB. The adducts were isolated using HPLC separation coupled with diode array detection (DAD) and structurally characterized from their electronic, mass- and nuclear magnetic resonance spectra. Four EB-adenine products were identified as N-1-(2-hydroxy-3-buten-1-yl) adenine (EB-Ade I), N-1-(1-hydroxy-3-buten-2-yl) adenine (EB-Ade II), N-3-(2-hydroxy-3-buten-1-yl) adenine (EB-Ade III) and N-3-(1-hydroxy-3-buten-2-yl) adenine (EB-Ade IV). Two previously reported guanine adducts: N-7-(2-hydroxy-3-buten-1-yl) guanine (EB-Gua I) and N-7-(1-hydroxy-3-buten-2-yl) guanine (EB-Gua II) were also collected. The purified adducts were used as reference compounds to detect and quantitate the corresponding adduct species formed in calf thymus DNA incubated with EB. All six adducts were detected in treated DNA. The N-7 position of guanine was the most reactive in DNA followed by N-3 of adenine and N-1 of adenine. The formation of N-1 and N-3-adenine adducts (EB-Ade I, 1.2 +/- 0.36; EB-Ade II, 0.8 +/- 0.27; EB-Ade III, 2.7 +/- 0.38; EB-Ade IV, 5.9 +/- 0.68 nmol/micromol Ade) in CT DNA was approximately one-tenth that of EB-guanine adducts (50.7 +/- 2.37 and 47.9 +/- 3.6 nmol/micromol Gua, respectively). The N-1-EB-Ade adducts detected in this study are likely to be the precursors of previously reported N6-EB-adenine adducts (Koivisto et al., 1995) through Dimroth rearrangement. Since BD and EB induce significant numbers of point mutations at A:T base pairs, the EB-adenine adducts may represent important lesions involved in BD-induced mutagenesis and carcinogenesis.
3,4-环氧-1-丁烯（EB）是丁二烯（BD）的主要诱变代谢产物，丁二烯是一种重要的工业化学品，被归类为可能的人类致癌物。尽管EB的致癌机制尚不清楚，但其与DNA亲核位点反应产生促诱变损伤的过程可能是多阶段致癌作用早期的关键步骤。本研究旨在表征EB与最具亲核性的DNA碱基（腺嘌呤（Ade）和鸟嘌呤（Gua））作为游离碱基、2'-脱氧核糖核苷以及小牛胸腺DNA（CT DNA）成分反应形成的加合物，以便深入了解EB对DNA修饰的性质。使用高效液相色谱分离结合二极管阵列检测（DAD）分离加合物，并通过其电子、质谱和核磁共振光谱对其结构进行表征。鉴定出四种EB-腺嘌呤产物，分别为N-1-（2-羟基-3-丁烯-1-基）腺嘌呤（EB-Ade I）、N-1-（1-羟基-3-丁烯-2-基）腺嘌呤（EB-Ade II）、N-3-（2-羟基-3-丁烯-1-基）腺嘌呤（EB-Ade III）和N-3-（1-羟基-3-丁烯-2-基）腺嘌呤（EB-Ade IV）。还收集了两种先前报道的鸟嘌呤加合物：N-7-（2-羟基-3-丁烯-1-基）鸟嘌呤（EB-Gua I）和N-7-（1-羟基-3-丁烯-2-基）鸟嘌呤（EB-Gua II）。将纯化的加合物用作参考化合物，以检测和定量与EB孵育的小牛胸腺DNA中形成的相应加合物种类。在处理过的DNA中检测到了所有六种加合物。鸟嘌呤的N-7位在DNA中反应性最强，其次是腺嘌呤的N-3位和腺嘌呤的N-1位。CT DNA中N-1和N-3-腺嘌呤加合物（EB-Ade I，1.2±0.36；EB-Ade II，0.8±0.27；EB-Ade III，2.7±0.38；EB-Ade IV，5.9±0.68 nmol/μmol Ade）的形成量约为EB-鸟嘌呤加合物（分别为50.7±2.37和47.9±3.6 nmol/μmol Gua）的十分之一。本研究中检测到的N-1-EB-Ade加合物可能是先前报道的N6-EB-腺嘌呤加合物（Koivisto等人，1995年）通过迪姆罗特重排的前体。由于BD和EB在A:T碱基对处诱导大量点突变，因此EB-腺嘌呤加合物可能代表参与BD诱导的诱变和致癌作用的重要损伤。
