Institute for Environmental Research, RWTH Aachen University, Worringerweg 1, 52074 Aachen, Germany.
Institute for Environmental Research, RWTH Aachen University, Worringerweg 1, 52074 Aachen, Germany; Department of Evolutionary Ecology and Environmental Toxicology, Goethe University Frankfurt, Max-von-Laue-Str. 13, 60438 Frankfurt am Main, Germany.
Sci Total Environ. 2021 Mar 10;759:143522. doi: 10.1016/j.scitotenv.2020.143522. Epub 2020 Nov 14.
Metabolism has to be considered during the toxicological assessment of chemical and environmental samples because it is an important process in the mammalian liver. It can be assessed in vitro via liver homogenates called S9-fractions, an external metabolic activation system. However, the external metabolic activation systems can vary greatly in their composition due to biological variations among individual animals and animal strains that the S9-fraction are derived as well as the differences in the production treatment. To gain more insight into these variances, three different but commonly used rat-derived S9-fractions were compared in the present study for their variance and performance with a reference compound in the Ames fluctuation assay with Salmonella typhimurium strains TA 98 and TA 100 according to ISO 11350. Severe shortcomings of conventional rat-derived S9-fractions were observed in the present study, such that S9-fractions differed significantly within the same rat strain and for different types of induction procedures in regards to the metabolic capability. An intrinsic mutagenic potential of the three rat-derived S9-fractions were identified in the Ames fluctuation assay with varying S9-fraction concentrations. To address some of the shortcomings of the animal-derived S9-fraction, the present study investigated the use and performance of a biotechnological, animal-free alternative, ewoS9R, in comparison to one of the rat-derived S9-fraction as the others showed a mutagenic potential themselves. Specifically, 12 different chemicals were used as a reference to determine if ewoS9R could serve as an adequate and more consistent replacement of traditional rat-derived metabolic activation systems: 8 pro-mutagenic compounds (i.e., require metabolic activation to show a mutagenic potential), one pro-mutagenic compound but not in the tested strains, one mutagenic compound without metabolic activation and two compounds that are equivocal in the literature. EwoS9R was evaluated as a promising approach in the Ames fluctuation assay with 5 compounds observed to have similar results with both rat-derived S9-fraction and ewoS9R (41%), for 3 compounds ewoS9R was a better metabolization system than the rat-derived S9-fraction (16%). Further research is necessary to determine the full potential of ewoS9R in comparison to rat-derived S9-fractions.
在对化学和环境样品进行毒理学评估时，必须考虑代谢，因为它是哺乳动物肝脏中的一个重要过程。可以通过称为 S9 级分的肝匀浆在体外评估，这是一种外部代谢激活系统。然而，由于个体动物和 S9 级分来源的动物品系之间的生物学差异以及生产处理的差异，外部代谢激活系统在组成上可能有很大差异。为了更深入地了解这些差异，本研究比较了三种不同但常用的源自大鼠的 S9 级分，以参考化合物在沙门氏菌 typhimurium 菌株 TA 98 和 TA 100 中的波动试验中的性能，根据 ISO 11350。本研究观察到常规大鼠衍生的 S9 级分存在严重缺陷，即 S9 级分在同一大鼠品系内以及不同诱导程序之间在代谢能力方面存在显著差异。在波动试验中，三种源自大鼠的 S9 级分均具有内在的致突变潜力，S9 级分浓度不同。为了弥补动物源性 S9 级分的一些缺陷，本研究研究了使用和性能生物技术，无动物替代物，ewoS9R，与大鼠衍生的 S9 级分之一进行比较，因为其他两种 S9 级分本身具有致突变潜力。具体而言，使用 12 种不同的化学品作为参考，以确定 ewoS9R 是否可以作为传统大鼠衍生代谢激活系统的充分和更一致的替代品：8 种促突变化合物（即需要代谢激活才能显示出致突变潜力），一种促突变化合物，但不在测试菌株中，一种无代谢激活的致突变化合物和两种在文献中模棱两可的化合物。ewoS9R 在波动试验中被评估为一种很有前途的方法，其中 5 种化合物与大鼠衍生的 S9 级分和 ewoS9R 具有相似的结果（41%），对于 3 种化合物，ewoS9R 是一种比大鼠衍生的 S9 级分更好的代谢系统（16%）。需要进一步研究以确定 ewoS9R 与大鼠衍生的 S9 级分相比的全部潜力。
