UPLC-MS/MS法测定蛇菰中7种成分含量及其活性成分没食子酸的降尿酸作用研究  

Determination of Seven Components in Balanophora laxiflora Hemsl.and B.henryi Hemsl.by UPLC-MS/MS and the Uric Acid-Lowering Effect of the Component Gallic Acid

作　　者：刘丽 桂利利 伍超奇 曹文洁 钱永帅 吴正坤 余惠凡 李飞 LIU Li;GUI Lili;WU Chaoqi;CAO Wenjie;QIAN Yongshuai;WU Zhengkun;YU Huifan;LI Fei(School of Pharmaceutical Sciences,Hubei University of Medicine,Shiyan 442000;Hubei Key Laboratory of Wudang Local Chinese Medicine Research,Shiyan 442000;Hubei University of Medicine,Institute of Biomedicine,Shiyan 442000)

机构地区：[1]湖北医药学院药学院,十堰442000 [2]武当特色中药研究湖北省重点实验室,十堰442000 [3]湖北医药学院生物医药研究院,十堰442000

出　　处：《中药药理与临床》2023年第9期49-55,共7页Pharmacology and Clinics of Chinese Materia Medica

基　　金：湖北医药学院PI项目(编号:HBMUPI201805);湖北医药学院研究生科技创新项目(编号:YC2021038、YC2021039、YC2022023);湖北医药学院药学院本科生科研训练计划项目(编号:YSRTP202102);湖北省卫健委青年人才项目(编号:WJ2021Q010);湖北省教育厅重点项目(编号:D20192101);大学生创新创业计划项目(编号:S202110929013、X202110929009)。

摘　　要：目的:建立疏花蛇菰和宜昌蛇菰中7种成分的含量测定方法,并探索其活性成分没食子酸的降尿酸作用。方法:采用超高效液相色谱-三重四极杆串联质谱法(UPLC-MS/MS)建立蛇菰中没食子酸、没食子酸甲酯、短叶苏木酚酸、咖啡酸、对香豆酸等5种酚酸类成分及圣草酚、柚皮素2种黄酮类成分的含量测定方法;建立腺苷诱导的肾小管上皮细胞NRK-52E高尿酸细胞模型,给予没食子酸4、20、100μmol/L,利用高效液相色谱(HPLC)法检测细胞上清液中尿酸含量;建立高尿酸血症(HUA)小鼠模型,给予100、300 mg/kg没食子酸后检测小鼠血清中尿酸(UA)、肌酐(Cr)、尿素氮(BUN)含量及肝脏和血清中黄嘌呤氧化酶(XO)活力。结果:7种成分在各自的浓度范围内线性关系良好(R~2>0.9982),精密度、重复性和稳定性均良好,平均加样回收率为87.35%~105.47%,RSD为1.14%~2.55%;没食子酸可降低腺苷诱导的细胞尿酸含量,且可降低HUA小鼠血清中UA、Cr、BUN含量,抑制肝脏及血清中XO活力。结论:本研究首次建立了UPLC-MS/MS法测定蛇菰成分,简单快速,重复性和稳定性良好,灵敏度高,且可用于同时检测蛇菰中多个成分的含量;蛇菰中活性成分没食子酸在体内外均有较好降尿酸作用,可能是通过减少尿酸生成发挥降尿酸作用,且可改善HUA所致尿酸性肾病。Objective:To develop a method for determining seven components in Balanophora laxiflora Hemsl.and B.henryi Hemsl.and explore the uric acid(UA)-lowering effect of the component gallic acid(GA).Methods:Ultra performance liquid chromatography-tandem mass spectrometry(UPLC-MS/MS)was used to establish a method for detecting the contents of five phenolic acids(GA,methyl gallate,brevifolincarboxylic acid,caffeic acid,p-coumaric acid)and two flavonoids(eriodictyol and naringenin).Adenosine was employed to induce high UA in NRK-52E cells and the cells were then treated with 4,20,and 100μmol/L GA.The concentration of UA in the supernatant was detected by HPLC.Hyperuricemia(HUA)was elicited in mice and then mice were treated with 100 and 300 mg/kg GA.The concentration of UA,creatinine(Cr),and blood urea nitrogen(BUN)in serum and the activity of xanthine oxidase(XO)in liver and serum were detected.Results:Seven components showed good linearity(R2>0.9982)in their respective concentration ranges,and the method had high precision,reproducibility,and stability,with average recoveries of 87.35%-105.47%and RSDs of 1.14%-2.55%.GA obviously reduced the level of UA induced by adenosine and the serum levels of UA,Cr,and BUN,and inhibited the activity of XO in liver tissue and serum.Conclusion:The UPLCMS/MS method was established for the first time for the determination of the components of B.laxiflora and B.henryi.The method features ease of implementation and high repeatability,stability,and sensitivity,which can be applied to simultaneously detect the content of multiple components.GA,the active component of B.laxiflora and B.henryi,lowers the level of UA in vitro and in vivo by suppressing the production of UA and ameliorates UA nephropathy caused by HUA.

关 键 词：疏花蛇菰 宜昌蛇菰 超高效液相色谱-三重四极杆串联质谱法 含量测定 没食子酸 降尿酸 尿酸性肾病 

分 类 号：R284.1[医药卫生—中药学]