苗药羊耳菊中7个抗炎活性成分在Raw264.7细胞中含量测定方法的建立  

Establishment of a method determining the contents of seven anti-inflammatory active components of the Miao medicine Inula cappa in Raw 264.7 cells

作　　者：周杰 薛寸 张青 陈思颖 陈艺 黄静 李月婷 黄勇 郑林 巩仔鹏 ZHOU Jie;XUE Cun;ZHANG Qing;CHEN Siying;CHEN Yi;HUANG Jing;LI Yueting;HUANG Yong;ZHENG Lin;GONG Zipeng(School of Pharmacy,Provincial Key Laboratory of Pharmaceutics in Guizhou,State Key Laboratory of Functions and Applications of Medicinal Plants,Guizhou Provincial Engineering Research Center for the Development and Application of Ethnic Medicine and TCM,Guizhou Medical University,Guiyang Guizhou 550004,China)

机构地区：[1]贵州医科大学药学院,贵州省药物制剂重点实验室,省部共建药用植物功效与利用国家重点实验室,民族药与中药开发应用教育部工程研究中心,贵州贵阳550004

出　　处：《江苏大学学报（医学版）》2022年第6期524-531,共8页Journal of Jiangsu University：Medicine Edition

基　　金：国家自然科学基金资助项目(81860734);贵州省大学生创新创业计划项目(S202110660046);国家大学生创新创业计划项目(20195200132);贵州医科大学优秀青年人才计划(2022-104)。

摘　　要：目的:建立同时测定Raw264.7细胞中羊耳菊提取物中7个抗炎活性成分木犀草苷(luteolin,LUT)、绿原酸(chlorogenic acid,CA)、新绿原酸(neochlorogenic acid,NCA)、隐绿原酸(cryptochlorogenic acid,CCA)、3,4-二咖啡酰基奎宁酸(3,4-dicaffeoylquinic acid,3,4-DCQA)、3,5-二咖啡酰基奎宁酸(3,5-dicaffeoylquinic acid,3,5-DCQA)、4,5-二咖啡酰基奎宁酸(4,5-dicaffeoylquinic acid,4,5-DCQA)含量的方法。方法:以小鼠巨噬细胞Raw 264.7为研究对象,采用超高液相色谱-三重四级杆质谱联用仪(UPLC-MS/MS)建立同时测定Raw 264.7细胞中LUT、CA、NCA、CCA、3,4-DCQA、3,5-DCQA、4,5-DCQA等7个成分含量的方法,包括专属性、线性、基质效应、回收率、准确度、精密度和稳定性等。结果:建立了一个快速稳定测量Raw264.7细胞中微量药物的分析方法,该方法的最低定量限为0.48~3.84 ng/mL,准确度为(90.78±5.30)%~(113.99±13.51)%,提取回收率为(87.19±5.33)%~(103.59±14.00)%,基质效应为(94.50±15.17)%~(112.77±7.02)%,稳定性为(91.71±11.70)%~(109.45±10.63)%,能够满足测量的要求。结论:建立了快速、准确的同时测定Raw264.7细胞羊耳菊提取物中7个活性成分(LUT、CA、NCA、CCA、3,4-DCQA、3,5-DCQA、4,5-DCQA)含量的UPLC-MS/MS方法,为开展羊耳菊提取物的细胞药代动力学研究奠定基础。Objective:To establish a method simultaneously determining the contents of seven anti-inflammatory active components including luteolin(LUT),chlorogenic acid(CA),and neochlorogenic acid(NCA),cryptochlorogenic acid(CCA),3,4-dicaffeoylquinic acid(3,4-DCQA),3,5-dicaffeoylquinic acid(3,5-DCQA),4,5-dicaffeoylquinic acid(4,5-DCQA)of the Miao medicine Inula cappa in Raw264.7 cells.Methods:Using mouse macrophage Raw264.7 as the research object,ultra-high liquid chromatography-triple quadrupole mass spectrometry(UPLC-MS/MS)was used to establish the simultaneous determination of LUT,CA,NCA,CCA,3,4-DCQA,3,5-DCQA,4,5-DCQA methods for the content of seven components of the Miao medicine Inula cappa,including specificity,linearity,matrix effect,recovery,accuracy,precision and stability.Results:A rapid and stable analytical method for the measurement of trace drugs in Raw264.7 cells was established.The lowest limit of quantification of the method was 0.48-3.84 ng/mL,and the accuracy was(90.78±5.30)%-(113.99±13.51)%.Its extraction recovery was(87.19±5.33)%-(103.59±14.00)%and the matrix effect was(94.50±15.17)%-(112.77±7.02)%.The stability was(91.71±11.70)%-(109.45±10.63)%,which could meet the measurement requirements.Conclusion:A rapid and accurate method for simultaneous determination content of seven active components LUT,CA,NCA,CCA,3,4-DCQA,3,5-DCQA,4,5-DCQA using UPLC-MS/MS was established,which could provide a reference for the study of cellular pharmacokinetics of Inula cappa extract.

关 键 词：羊耳菊提取物 RAW264.7细胞 抗炎成分 绿原酸 细胞药代动力学 UPLC-MS/MS 

分 类 号：R965[医药卫生—药理学]