School of Pharmacy, Nanjing University of Chinese Medicine, Nanjing 210023, China; Jiangsu Key Laboratory of Pediatric Respiratory Disease, Institute of Pediatrics, Nanjing University of Chinese Medicine, Nanjing 210023, China; Medical Metabolomics Center, Nanjing University of Chinese Medicine, No.138 Xianlin Avenue, Nanjing 210023, China.
School of Pharmacy, Nanjing University of Chinese Medicine, Nanjing 210023, China.
J Chromatogr B Analyt Technol Biomed Life Sci. 2021 Jul 15;1178:122857. doi: 10.1016/j.jchromb.2021.122857. Epub 2021 Jul 10.
The complexity of ingredients in traditional Chinese medicine (TCM) makes it challenging to clarify its efficacy in an acceptable and scientific approach. The present study was aimed to use quantification results from targeted cellular metabolomics to evaluate anti-aging efficacy of a famous Chinese medicine formula, Erzhi Wan (EZW), and screen possible effective extracts, depending on the developed strategy integrating multivariate receiver operating characteristic (ROC) curve and analytic hierarchy process (AHP). In this study, senescent NRK cells induced by D-galactose were treated with drug-containing serum of EZW and four kinds of extracts (petroleum ether, ethyl acetate, butanol and water). Intermediates of two major metabolic pathways for energy synthesis, tricarboxylic acid (TCA) cycle and glycolysis, were accurately quantified by GC-MS/MS to identify discriminate metabolites for clarifying therapeutic mechanism of EZW based on multivariate statistical analysis. Senescent and non-senescent cells were successfully distinguished using these metabolites by ROC curve analysis. Next, these metabolites were used as evaluation indexes to quantitatively reflect different effect of EZW and its extracts, according to the role of them in distinguishing groups and in conjunction with AHP. In vitro detection of senescence-associated β-galactosidase (SA-β-gal) activity was used to verify the reliability of evaluation results. The reversal after treatment of drug-containing serum of EZW and extracts was observed, and the petroleum ether extract might be the potential active extract responsible for the major anti-aging effect of EZW, which was in agreement with in vitro experiments. Altogether, metabolomics was a powerful approach for evaluation efficacy and elucidation action mechanisms of TCM. The integrated evaluation strategy in this paper with properties of high practicality, feasibility and effectivity was expected to provide a new insight into comprehensive and quantitative efficacy evaluation.
中药（TCM）成分的复杂性使得用可接受和科学的方法来阐明其疗效具有挑战性。本研究旨在使用靶向细胞代谢组学的定量结果来评估一种著名的中药方剂二至丸（EZW）的抗衰老功效，并根据整合多元接收器操作特征（ROC）曲线和层次分析法（AHP）的开发策略，筛选可能的有效提取物。在这项研究中，用 D-半乳糖诱导的衰老 NRK 细胞用 EZW 和四种提取物（石油醚、乙酸乙酯、正丁醇和水）的含药血清处理。通过 GC-MS/MS 准确定量了能量合成的两个主要代谢途径，三羧酸（TCA）循环和糖酵解的中间产物，以基于多变量统计分析来识别区分 EZW 治疗机制的代谢物。通过 ROC 曲线分析，使用这些代谢物成功区分了衰老和非衰老细胞。接下来，根据它们在区分组中的作用以及与 AHP 结合，将这些代谢物用作评估指标，定量反映 EZW 及其提取物的不同作用。体外检测衰老相关β-半乳糖苷酶（SA-β-gal）活性，以验证评估结果的可靠性。观察 EZW 含药血清和提取物的处理后逆转情况，石油醚提取物可能是负责 EZW 主要抗衰老作用的潜在有效提取物，这与体外实验结果一致。总之，代谢组学是一种评估 TCM 功效和阐明作用机制的有力方法。本文中具有实用性、可行性和有效性的综合评价策略有望为综合定量功效评价提供新的视角。
