基于UPLC-Q-Exactive Orbitrap MS的佛耳草多成分定量分析  

Quantitative Analysis of M ulti-components in Gnaphalium affine Based onUPLC-Q-Exactive Orbitrap MS

作　　者：曹春艳 张家祺[1] 赵振铎 沈沁 戚胜兰 刘伟[2] 张立超[1] CAO Chunyan;ZHANG Jiaqi;ZHAO Zhenduo;SHEN Qin;QI Shenglan;LIU Wei;ZHANG Lichao(Shanghai Municipal Hospital of Traditional Chinese Medicine(TCM),Shanghai 200071,China;Third Grade Laboratory of TCM Preparations of National Administration of TCM,Key Laboratory of Liver and Kidney Diseases,Ministry of Education,Shanghai Key Laboratory of Traditional Chinese Clinical Medicine,Shuguang Hospital Affiliated to Shanghai University of TCM,Shanghai 201203,China)

机构地区：[1]上海中医药大学附属市中医医院,上海200071 [2]上海中医药大学附属曙光医院,国家中医药管理局中药制剂三级实验室,肝肾疾病证教育部重点实验室,上海市中医临床重点实验室,上海201203

出　　处：《中国实验方剂学杂志》2024年第9期148-155,共8页Chinese Journal of Experimental Traditional Medical Formulae

基　　金：国家自然科学基金面上项目(81973551);上海市中医药三年行动计划项目[ZY(2021—2023)-0203-04];上海医学创新发展基金会未来计划项目(WL-YSXM-2022002K);上海市教育委员会和上海市教育发展基金会“晨光计划”项目(20CG50);中国科协青年托举工程项目(2020QNRC001);上海医学创新发展基金会中医药传承发展项目(WLJH20212Y-ZYY007)。

摘　　要：目的:建立一种快速稳定的液质联用方法同时分析佛耳草中17种化学成分的含量,为完善佛耳草质量标准提供实验依据。方法:采用超高效液相色谱-四极杆/静电场轨道阱质谱法(UPLC-Q-Exactive Orbitrap MS)对15批不同产地佛耳草中的17种成分进行定量分析,使用ACQUITY UPLC?BEH C18色谱柱(2.1 mm×100 mm,1.7μm),以甲醇(A)-0.1%甲酸水溶液(B)为流动相梯度洗脱(0~1.0min,8%A;1.0~4.0min,8%~26%A;4.0~9.0min,26%A;9.0~14.0min,26%~34%A;14.0~14.5min,34%~45%A;14.5~15.0min,45%~60%A;15.0~18.0min,60%~90%A;18.0~19.0min,90%A;19.0~19.01min,90%~8%A;19.01~20.0 min,8%A),流速0.3 mL·min^(-1),柱温40℃,进样量2μL;运用电喷雾离子源,正、负离子模式全扫描,扫描范围m/z 100~1000。结果:所建立的测定方法经方法学验证良好,能够用于17种成分的同时定量。15批佛耳草样品中奎宁酸、没食子酸、原儿茶酸、新绿原酸、绿原酸、隐绿原酸、咖啡酸、1,3-O-二咖啡酰奎宁酸、异绿原酸A、异槲皮苷、1,5-O-二咖啡酰奎宁酸、大波斯菊苷、紫云英苷、异绿原酸C、木犀草素、芹菜素、高车前素的质量范围依次为39.60~179.12、0.17~0.84、2.41~8.38、4.33~31.50、13.63~180.38、2.43~14.75、1.16~19.68、0.49~5.63、55.77~445.16、0.23~10.26、62.04~530.10、1.11~18.01、11.36~90.61、12.22~65.98、7.22~69.84、3.37~45.65、0.30~2.59μg·g^(-1),其中有机酸类成分在佛耳草中含量比黄酮类成分更高,以1,5-O-二咖啡酰奎宁酸、异绿原酸A、奎宁酸、绿原酸含量较高,同时,贵州产地佛耳草样品中黄酮类成分含量高于江苏产地样品,江苏产地佛耳草样品中有机酸类成分含量则高于贵州产地样品。结论:建立的检测方法可用于佛耳草中17种成分含量的快速、准确测定,明确了佛耳草主要成分的含量范围,可为佛耳草质量控制标志物的选择提供参考。Objective:To establish a rapid and stable liquid chromatography-mass spectrometry(LC-MS)for simultaneous analysis of 17 chemical components in Gnaphalium affine aboveground parts with flowers,so as to provide experimental basis for improving the quality standard of this herb.M ethod:Ultra performance liquid chromatography-quadrupole/electrostatic field orbitrap mass spectrometry(UPLC-Q-Exactive Orbitrap MS)was used for the quantitative analysis of 17 constituents in 15 batches of G.affine from different origins,the separation was performed on an ACQUITY UPLC®BEH C18 column(2.1 mm×100 mm,1.7μm)with the mobile phase of methanol(A)-0.1%formic acid aqueous solution(B)for gradient elution(0-1.0 min,8%A;1.0-4.0 min,8%-26%A;4.0-9.0 min,26%A;9.0-14.0 min,26%-34%A;14.0-14.5 min,34%-45%A;14.5-15.0 min,45%-60%A;15.0-18.0 min,60%-90%A;18.0-19.0 min,90%A;19.0-19.01 min,90%-8%A;19.01-20.0 min,8%A),the flow rate was 0.3 mL·min-1,the column temperature was 40°C and the injection volume was 2μL.And the electrospray ionization was used with full scanning in both positive and negative ion modes,and the scanning range was m/z100-1000.Result:The established method has been verified by the methodology and could be used for the simultaneous quantification of 17 components in G.affine.The content ranges of the 17 components(quinic acid,gallic acid,protocatechuic acid,neochlorogenic acid,chlorogenic acid,cryptochlorogenic acid,caffeic acid,1,3-O-dicaffeoylquinic acid,isochlorogenic acid A,isoquercitrin,1,5-O-dicaffeoylquinic acid,apigenin-7-O-glucoside,astragalin,isochlorogenic acid C,luteolin,apigenin and hispidulin)in 15 batches of G.affine samples was 39.60-179.12,0.17-0.84,2.41-8.38,4.33-31.50,13.63-180.38,2.43-14.75,1.16-19.68,0.49-5.63,55.77-445.16,0.23-10.26,62.04-530.10,1.11-18.01,11.36-90.61,12.22-65.98,7.22-69.84,3.37-45.65,0.30-2.59μg·g-1,respectively.The content of organic acids was higher than that of flavonoids in G.affine,and the contents of 1,5-O-dicaffeoylquinic acid,isochlorogenic acid A,quinic acid and chloroge

关 键 词：佛耳草 定量分析 黄酮类 有机酸类 超高效液相色谱-四极杆/静电场轨道阱质谱法(UPLC-Q-Exactive Orbitrap MS) 质量控制 质量标志物 

分 类 号：R22[医药卫生—中医基础理论] R28[医药卫生—中医学] R931[理学—分析化学] O657[理学—化学]