Quantification of the activity of detoxifying enzymes in terrestrial invertebrates: optimization, evaluation and use of in vitro and ex vivo methods

1. The negative effects of pesticides on beneficial terrestrial invertebrates have received much attention, and in this context quantification of the activity of metabolic enzymes provides an important tool. Unfortunately, current methodologies are limited in their use.
2. In this study, the grain beetle, Tenebrio molitor was used to illustrate why in vitro methods are ineffective to quantify metabolic enzymes in some insects. It was further used to optimize an ex vivo assay that can replace in vitro methods when these are ineffective. The ex vivo method can quantify metabolic enzymatic activities of cytochrome P450 mixed function oxidases (CYPs), general esterases (EST) and glutathione S-transferases (GST).
3. In detail we show, that T. molitor produces inhibitory and/or degrading agents, which are released during tissue homogenization. As these agents inhibit the in vitro CYP activity of rat liver microsomes (RLM) we suggest, that they are also responsible for the lack of in vitro C...

1. 农药对有益陆生无脊椎动物的负面影响已受到广泛关注，在此背景下，代谢酶活性定量分析成为一项重要研究手段。遗憾的是，现有方法的应用存在诸多局限。
2. 本研究以黄粉虫（Tenebrio molitor）为实验对象，阐明了体外检测方法为何无法对部分昆虫的代谢酶进行定量分析。同时，本研究进一步优化了一种离体检测体系，当体外方法失效时，该体系可替代体外检测方法。该离体方法可实现细胞色素P450混合功能氧化酶（CYPs）、总酯酶（EST）以及谷胱甘肽S-转移酶（GST）的代谢酶活性定量检测。
3. 具体而言，本研究证实黄粉虫会产生抑制性和/或降解性物质，此类物质在组织匀浆过程中会被释放出来。由于该类物质可抑制大鼠肝微粒体（RLM）的体外CYP酶活性，因此我们推测，此类物质亦是导致体外检测方法无法有效定量C...