Detection and quantification of specific-DNA adducts by LC-MS/MS in the livers of rats given estragole at the carcinogenic dose
Detection and quantification of specific-DNA adducts by LC-MS/MS in the livers of rats given estragole at the carcinogenic dose
复制标题
通过 LC-MS/MS 对给予致癌剂量草蒿素的大鼠肝脏中的特异性 DNA 加合物进行检测和定量
DOI：
10.1021/tx100410y
复制
发表时间：
2011
期刊：
影响因子：
--
通讯作者：
A. Nishikawa
中科院分区：
文献类型：
--
作者：
Y. Ishii;Y;Suzuki;D. Hibi;M. Jin;K. Fukuhara;T. Umemura;A. Nishikawa
雌酚(ES)是几种草药和香料的天然成分，在啮齿动物的肝脏中起致癌作用。由于ES的近端亲电形式是由细胞色素P450和磺基转移酶代谢途径产生的，因此与DNA碱基的共价加合物可能在癌变过程中发挥关键作用。已经报道了ES特异性脱氧鸟苷(DG)和脱氧腺苷(DA)加合物的存在，并证实了DG加合物的精确化学结构。在本研究中，我们用LC-ESI/MS检测了DA与模拟的磺基转移酶代谢途径产生的1‘-乙酰氧基-ES反应后，ES特异性的DA加合物的形成。虽然在LC-ESI/MS色谱图中观察到两个峰，但通过核磁共振分析确定ES-3‘-N6-da为可测峰。为了确定ES特异性DG和da加合物在体内的形成，建立了一种适用于ES-3‘-N6-da及其两个主要DG加合物，即ES-3’-C8-dG和ES-3‘-n2-dG体内样品的同位素稀释LC-ESI/MS/MS方法。Es-3‘-C8-dG和Es-3’-n2-dG的柱定量下限分别为0.2fmoL和0.06fmoL。利用发展中的分析方法，我们试图测量ES在可能致癌剂量(600 mg/kg bw)处理4周的大鼠肝脏中的加合物水平。ES组大鼠肝脏中ES-3‘-C8-dG、ES-N_2-dG和ES-3’-N_6-da的含量分别为3.5±0.4、4.8±0.8和20.5±1.6/106dG或da。这一定量数据和新开发的体内加合物观察技术可能有助于ES肝癌发生的研究。
Estragole (ES) is a natural constituent of several herbs and spices that acts as a carcinogen in the livers of rodents. Given that the proximal electrophilic form of ES with a reactive carbocation is generated by cytochrome P450 and a sulfotransferase metabolizing pathway, there is a possibility that the resultant covalent adducts with DNA bases may play a key role in carcinogenesis. The existence of ES-specific deoxyguanosine (dG) and deoxyadenosine (dA) adducts has already been reported with the precise chemical structures of the dG adducts being confirmed. In the present study, we examined ES-specific dA adduct formation using LC-ESI/MS after the reaction of dA with 1′-acetoxy-ES produced by a sulfotransferase metabolic pathway mimic. Although two peaks were observed in the LC-ESI/MS chromatogram, the identification of ES-3′-N6-dA as the measurable peak was determined by NMR analysis. To confirm ES-specific dG and dA adduct formation in vivo, an isotope dilution LC-ESI/MS/MS method applicable to in vivo samples for ES-3′-N6-dA together with the two major dG adducts, that is, ES-3′-C8-dG and ES-3′-N2-dG, was developed using selected ion recording. The limit of quantification was 0.2 fmol on column for ES-3′-C8-dG and ES-3′-N2-dG and 0.06 fmol on column for ES-3′-N6-dA, respectively. Using the developing analytical method, we attempted to measure adduct levels in the livers of rats treated with ES at a possible carcinogenic dose (600 mg/kg bw) for 4 weeks. ES-3′-C8-dG, ES-N2-dG, and ES-3′-N6-dA were detected at levels of 3.5 ± 0.4, 4.8 ± 0.8, and 20.5 ± 1.6/106dG or dA in the livers of ES-treated rats. This quantitative data and newly developed technique for adduct observation in vivo might be helpful for ES hepatocarcinogenesis investigations.