CYP2E1 is one of the most crucial isozymes of CYP450. It is responsible for metabolizing and activating a large number of toxicants and carcinogens, but the correlation between its abundance and activity has not been widely studied. With the flourishing of modern mass spectrometry technology, quantifying complex biological proteins and studying the relationship between their abundance and activity have become practicable. In our study, an accurate, sensitive, and stable LC-MS/MS-based method was developed and validated. The method can accurately quantify the abundance of CYP2E1 in the rat liver microsome and S9 fraction. The quantitative linearity of the method is between 2 and 320 ng/mL, and the run time is 16.5 minutes. Meanwhile, we used the probe substrate method (with chlorzoxazone as the substrate) as a reference to analyze the correlation between its activity and abundance. The result illustrated that the abundance of CYP2E1 by LC-MS/MS has a strong positive correlation with its activity. This is a relationship worth studying, which has not been reported before. We also explored the correlation between quantitative results by traditional methods (western blot and RT-PCR) and activity, and the positive correlation was not obvious. Therefore, when testing the correlation between metabolic enzyme abundance and activity, the LC-MS/MS-based method is confirmed to be more accurate than conventional methods. It will provide a meaningful way of researching the metabolic enzymes in drug interactions. Furthermore, we found that the S9 fraction can also be used for mass spectrometry quantitative analysis, which is helpful for promoting the practical application of targeted protein technology.

CYP2E1是CYP450最关键的同工酶之一。它负责代谢和激活大量的毒物和致癌物，但其丰度与活性之间的关系尚未得到广泛研究。随着现代质谱技术的蓬勃发展，对复杂的生物蛋白进行定量并研究其丰度与活性之间的关系已变得可行。在我们的研究中，开发并验证了一种基于LC-MS / MS的准确，灵敏和稳定的方法。该方法可以准确定量大鼠肝微粒体中CYP2E1和S9组分的丰度。该方法的定量线性在2至320 ng / mL之间，运行时间为16.5分钟。与此同时，我们采用探针底物法（以氯唑沙宗为底物）作为分析其活性与丰度之间相关性的参考。结果表明，LC-MS / MS对CYP2E1的丰度与其活性呈强正相关。这是一个值得研究的关系，以前没有报道过。我们还探讨了传统方法（western blot和RT-PCR）定量结果与活性之间的相关性，但正相关性不明显。因此，当测试代谢酶丰度与活性之间的相关性时，证实了基于LC-MS / MS的方法比常规方法更准确。这将为研究药物相互作用中的代谢酶提供有意义的方法。此外，