目的 基于代谢组学和网络药理学分析相结合的方法探究蕲艾Artemisia argyi的安全性与毒理学机制。方法 采用气相色谱-质谱（gas chromatography-mass spectrometry，GC-MS）和超高效液相色谱串联四极杆飞行时间质谱（ultra performance liquid chromatography coupled to tandem quadrupole time-of-flight mass spectrometry，UPLC-Q-TOF-MS）技术分析蕲艾挥发油组分（essential oil from A. argyi，AAEO）、石油醚组分、醋酸乙酯组分、正丁醇组分和水组分的化学组成；利用KM小鼠连续7 d给药评价AAEO和4个非挥发性组分的安全性；代谢组学技术检测AAEO干预前后肝脏内源性代谢物的变化；网络药理学分析AAEO肝毒性的潜在成分和作用靶点；整合代谢组学和网络药理学并构建“代谢物-反应-酶-基因”相互作用网络揭示AAEO肝毒性的毒理学机制。结果 共鉴定出AAEO中39个成分，占挥发油含量的89.30%；从4个非挥发性组分中共鉴定出34个成分。与对照组比较，AAEO显著增加小鼠肝脏指数（P＜0.05、0.01），上调肝脏丙氨酸氨基转移酶（alanine aminotransferase，ALT）和天冬氨酸氨基转移酶（aspartate aminotransferase，AST）的活性（P＜0.01），并损伤肝脏组织病理结构呈现出明显肝毒性。代谢组学结果显示给予AAEO后肝脏的17个代谢物水平发生显著变化，包含9种氨基酸，主要涉及苯丙氨酸、酪氨酸和色氨酸生物合成，丙氨酸、天冬氨酸和谷氨酸代谢等途径；网络药理学分析提示樟脑、侧柏酮、石竹素、(−)-宁酮等10个成分是AAEO中潜在的肝毒性物质；整合代谢组学和网络药理学分析筛选出苯丙氨酸羟化酶（phenylalanine hydroxylase，PAH）为AAEO肝毒性的关键靶点，且PAH mRNA和蛋白表达在AAEO高、低剂量组均显著降低（P＜0.05、0.01），揭示AAEO通过下调PAH表达抑制酪氨酸生物合成产生肝毒性。分子对接结果显示10个潜在毒性物质与PAH结合能均小于−20.00 kJ/mol。结论 结合代谢组学和网络药理学阐明了蕲艾的毒性组分、可能的毒性物质及毒理学机制，为蕲艾产业的发展和临床应用提供了依据。

Objective To investigate the safety and toxicological mechanism of Artemisia argyi based on combination of metabolomics and network pharmacological analysis. Method The chemical compositions of essential oil from A. argyi (AAEO), petroleum ether fraction, ethyl acetate fraction, n-butanol fraction and water fraction of A. argyi were analyzed by gas chromatography-mass spectrometry (GC-MS) and ultra performance liquid chromatography coupled to tandem quadrupole time-of-flight mass spectrometry (UPLC-Q-TOF-MS). The safety of AAEO and four non-volatile fractions of A. argyi was evaluated in KM mice for 7 d. Metabonomics technique was used to detect the changes of endogenous metabolites in liver before and after AAEO intervention. The potential components and targets of hepatotoxicity of AAEO were analyzed by network pharmacology. The toxicological mechanism of AAEO hepatotoxicity was revealed by integrating metabolomics and network pharmacology and constructing a “metabolite-reaction-enzyme-gene” interaction network. Results A total of 39 components were identified in AAEO, accounting for 89.30% of the essential oil content, and 34 components were identified from four non-volatile fractions. Compared with control group, AAEO significantly increased the liver coefficient (P < 0.05, 0.01), up-regulated the activities of alanine aminotransferase (ALT) and aspartate aminotransferase (AST) in liver (P < 0.01), and damaged the pathological structure of the liver showing obvious hepatotoxicity. Metabonomics revealed significant changes in the levels of 17 metabolites in the liver after AAEO intervention, including nine amino acids, mainly involved in phenylalanine, tyrosine and tryptophan biosynthesis, alanine, aspartate and glutamate metabolism. The network pharmacological analysis suggested that 10 components of AAEO, including (+)-camphor, thujone, caryophyllene oxide and (+)-3-thujone were potential hepatotoxic substances. Integrated metabonomics and network pharmacology analysis identified phenylalanine hydroxylase (PAH) as the key target of AAEO hepatotoxicity, and PAH mRNA and protein expressions were significantly decreased in both high and low dose groups of AAEO (P < 0.05, 0.01), revealing that AAEO produced hepatotoxicity by down-regulating PAH expression and inhibiting tyrosine biosynthesis. The results of molecular docking showed that the binding energies of 10 potentially toxic substances to PAH were all less than −20.00 kJ/mol. Conclusion The toxic fraction, possible toxic substances and toxicological mechanism of A. argyi were clarified by combining with metabonomics and network pharmacology, which provides a basis for the development and clinical application of A. argyi industry.

R285.5

国家自然科学基金资助项目（32270391）；湖北省自然科学基金资助项目（2023AFA032，2022CFB391）；湖北省中医药管理局重点项目（ZY2023Z023）