The instability of electrophilic reactive metabolites in in vitro metabolism studies makes their accurate analysis challenging. To stabilise the reactive compounds prior to their analysis, different trapping agents, such as thiols, amines and cob(I)alamin, have earlier been tested depending on the metabolites to be analysed and the type of study. In the present work, DNA is introduced as a trapping agent for measuring the formation of bulky electrophilic metabolites. Benzo[a]pyrene (B[a]P), a polycyclic aromatic hydrocarbon (PAH), was used as a model compound in a rat liver S9 metabolic system. Under physiological incubation conditions, B[a]P metabolises to diol epoxide (BPDE) metabolites which were trapped by DNA resulting in the formation of covalently bound DNA adducts. The methodology for analysis of these adducts included extraction of the DNA from the metabolic system, digestion of the DNA to yield nucleosides and analysis of the BPDE-adduct to deoxyguanosine (BPDE-dG) by liquid chromatography coupled to high resolution mass spectrometry (HRMS). The chromatographic conditions in combination with the high mass accuracy data (±3 ppm) was useful in resolving BPDE-dG in its protonated form from the complex set of ions present in the metabolic matrix. The method was validated in terms of sensitivity, specificity, accuracy, precision and recovery, and applied to provide a preliminary estimate of BPDE-dG levels from the metabolism of B[a]P in rat S9. The use of DNA as a trapping agent for in vitro metabolites has a potential to aid in cancer risk assessment procedure of PAHs, for instance, in inter-species comparison of metabolism to reactive metabolites and can be adapted for screening of genotoxic metabolites, e.g., from emerging environmental contaminants.

亲电子反应性代谢产物的体外不稳定性代谢研究使其准确分析具有挑战性。为了在分析之前稳定反应性化合物，较早地根据要分析的代谢物和研究类型对不同的捕集剂（例如硫醇，胺和钴（I）丙氨酸）进行了测试。在目前的工作中，引入DNA作为捕获剂，用于测量庞大的亲电子代谢物的形成。苯并[a] py（B [a] P），一种多环芳烃（PAH），被用作大鼠肝脏S9代谢系统中的模型化合物。在生理温育条件下，B [a] P代谢为二醇环氧化物（BPDE）代谢物，被DNA捕获，导致形成共价结合的DNA加合物。分析这些加合物的方法包括从代谢系统中提取DNA，消化DNA以产生核苷，并通过液相色谱与高分辨率质谱（HRMS）结合对BPDE加合物与脱氧鸟苷（BPDE-dG）进行分析。色谱条件与高质量准确度数据（±3 ppm）相结合可用于从代谢基质中存在的复杂离子组中分离出质子化形式的BPDE-dG。该方法在敏感性，特异性，准确性，精密度和回收率方面均得到验证，可用于根据大鼠S9中B [a] P的代谢提供BPDE-dG水平的初步估计。DNA作为诱捕剂的用途 色谱条件与高质量准确度数据（±3 ppm）相结合可用于从代谢基质中存在的复杂离子组中分离出质子化形式的BPDE-dG。该方法在敏感性，特异性，准确性，精密度和回收率方面均得到验证，可用于根据大鼠S9中B [a] P的代谢提供BPDE-dG水平的初步估计。DNA作为诱捕剂的用途 色谱条件与高质量准确度数据（±3 ppm）相结合可用于从代谢基质中存在的复杂离子组中分离出质子化形式的BPDE-dG。该方法在敏感性，特异性，准确性，精密度和回收率方面均得到验证，可用于根据大鼠S9中B [a] P的代谢提供BPDE-dG水平的初步估计。DNA作为诱捕剂的用途体外代谢物可能有助于PAHs的癌症风险评估程序，例如，在代谢与反应性代谢物的种间比较中，并且可以适用于从新出现的环境污染物中筛选遗传毒性代谢物。