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Journal of ZheJiang University(Medical Science)  2015, Vol. 44 Issue (2): 145-153    DOI: 10.3785/j.issn.1008-9292.2015.03.005
    
Screening of active compounds with myocardial protective effects from Tongmai Yangxin pill
TAO Shan1, LIANG Xin-ying1, WANG Yi2, WANG Yi1
1. College of Pharmacy, Zhejiang University, Hangzhou 310058, China;
2. Tianjin Chinese Medical University, Tianjin National Key Laboratory of Modern Chinese Medicine, Tianjin 300193, China
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Abstract  

Objective: Based on cell model and HPLC-MS technology, to screen myocardial protection active compounds from traditional patent medicine Tongmai Yangxin pill (TMYXP). Methods: Fractions of TMYXP were prepared by high performance liquid preparation technology. The cardioprotective effects of prepared fractions were tested on H2O2 oxidation-damaged H9c2 myocardiocytes. The active components were analyzed by high performance liquid chromatography (HPLC) coupled with high resolution mass spectrometry. The possible active compounds were putatively identified by comparison of their MS ions and molecular weight with literatures. Results: Ten TMYXP components presented significant myocardial protective activities, 5 of which were investigated and presented good dose-effect relationships. Their median effective concentrations (EC50) were respectively 11.66, 17.44, 13.10, 7.332, 15.15 μg/mL. Totally, 11 potential active compounds were analyzed and identified, including Glycyrrhizic acid, Glycycoumarin, Licoisoflavone, Ophiopogonin D', Licoricon, Gancaonin L, Neoglycyrol, Emodin, Angeloylgomisin H, Angeloylgomisin Q and Glyasperin A.Conclusion: The myocardial protection active compounds of TMYXP were screened successfully.



Key wordsMyocytes, cardiac      Anoxia      Chromatography, high pressure liquid      Glycyrrhiza uralensis/drug effects      Flavones/pharmacology      Glycyrrhizic acid/pharmacology      Schisandra chinensis      Ophiopogon japonicus      Disease models, animal     
Received: 06 November 2014      Published: 25 March 2015
CLC:  R965.1  
Cite this article:

TAO Shan, LIANG Xin-ying, WANG Yi, WANG Yi. Screening of active compounds with myocardial protective effects from Tongmai Yangxin pill. Journal of ZheJiang University(Medical Science), 2015, 44(2): 145-153.

URL:

http://www.zjujournals.com/med/10.3785/j.issn.1008-9292.2015.03.005     OR     http://www.zjujournals.com/med/Y2015/V44/I2/145


通脉养心丸中心肌保护活性物质的筛选

目的:基于细胞模型以及高效液相色谱—质谱联用技术对通脉养心丸中心肌保护活性物质进行筛选研究。方法:利用高效液相制备技术对通脉养心丸进行制备分离,然后采用过氧化氢氧化损伤H9c2大鼠心肌细胞模型对通脉养心丸活性组分进行筛选。利用高效液相色谱—质谱联用技术对活性组分进行分析,通过查阅文献和根据化合物裂解规律,鉴定推断出可能的活性化合物。结果:发现通脉养心丸中10个组分具有明显的心肌保护活性,考察了其中5个组分的半数有效浓度并发现其呈现良好的量效关系。从这5个组分中推断出11个可能的活性化合物,包含甘草酸、甘草香豆素、甘草利酮、Glyasperin A等。结论:筛选出通脉养心丸中心肌保护活性物质,有望为中药复方制剂活性物质筛选研究提供借鉴。


关键词: 肌细胞,心脏,  缺氧,  色谱法,高压液相,  甘草/药物作用,黄酮类/药理学,  甘草酸/药理学,  五味子、麦冬,  疾病模型,动物 

[1] 王怡, 张玲, 肖扬, 等.通脉养心丸对缺氧诱导心肌细胞损伤炎症因子及氧化应激的影响[J].中医杂志, 2011,52(4):326-328. WANG Yi, ZHANG Ling, XIAO Yang, et al. Effect of Tongmai Yangxin Pills on inflammatory factors and oxidative stress of cardiomyocyte injury induced by hypoxia[J].Journal of Traditional Chinese Medicine, 2011,52(4):326-328.(in Chinese)
[2] 蔡小军, 王怡, 胡利民, 等.通脉养心丸抗肾上腺素急性心律失常作用[J].天津中医药大学学报, 2009,28(3):133-135. CAI Xiao-jun, WANG Yi, HU Li-min, et al. Effect of Tongmai Yangxin pill in treating adrenaline-induced acute arrhythmia[J].Journal of Tianjin University of Traditional Chinese Medicine, 2009,28(3):133-135.(in Chinese)
[3] 肖扬, 张家福, 张玲, 等.从钙超载角度探讨通脉养心丸心肌保护作用的机制[J].中西医结合心脑血管病杂志, 2011,9(5):562-563. XIAO Yang, ZHANG Jia-fu, ZHANG Ling, et al. Investigation of protection mechanism of Tongmai Yangxin Pill on myocardium from the perspective of calcium overload [J]. Chinese Journal of Integrative Medicine on Cardio-/Cerebro Vascular Disease, 2011,9(5):562-563. (in Chinese)
[4] SUN B, SUN G B, XIAO J, et al. Isorhamnetin inhibits H2O2-induced activation of the intrinsic apoptotic pathway in H9c2 cardiomyocytes through scavenging reactive oxygen species and ERK inactivation[J]. J Cell Biochem, 2012,113(2):473-485.
[5] 祝波, 李宏霞, 张凌云, 等. 槲皮素对H2O2所致乳鼠培养心肌细胞损伤的保护作用[J].哈尔滨医科大学学报, 2006,40(6):457. ZHU Bo, LI Hong-xia, ZHANG Ling-yun, et al. Protective effect of quercetin on hydroperoxide-induced neonate rat myocardial injury[J].Journal of Harbin Medical University, 2006,40(6):457.(in Chinese)
[6] 高丽, 张哲, 许树钦, 等. 赤芍总苷对过氧化氢所致H9C2心肌细胞损伤的保护作用[J].时珍国医国药, 2011,22(5):1194-1195. GAO Li, ZHANG Zhe, XU Shu-qin, et al. The protective effect of total paeony glycoside on H9C2 myocardial cells injury induced by hydrogen peroxide[J]. Lishizhen Medicine and Materia Medica Research, 2011, 22(5):1194-1195. (in Chinese)
[7] 谈志龙, 邢国胜, 王淑云, 等. MTT法观察补肾健骨汤对大鼠成骨细胞增殖的影响[J]. 中国骨质疏松杂志, 2004, 10(1):94-95. TAN Zhi-long, XING Guo-sheng, WANG Shu-yun, et al. Effects of Bushen Jiangu decoction on proliferation of rat osteoblast assessed by MTT method[J]. Chinese Journal of Osteoporosis, 2004,10(1):94-95. (in Chinese)
[8] 陈旭光, 唐俊明, 张蕾, 等. 过氧化氢预处理对H9C2心肌细胞氧化应激损伤的保护作用[J].现代生物医学进展, 2013, 13(34):6615-6618. CHEN Xu-guang, TANG Jun-ming, ZHANG Lei, et al. Protective effects of H2O2 preconditioning against oxidative stress injury in H9C2 cells[J].Progress in Modern Biomedicine, 2013,13(34):6615-6618.(in Chinese)
[9] 李月婷, 刘影, 王静娴, 等. 过氧化氢对原代心肌细胞及H9C2细胞的损伤作用[J].贵阳医学院学报, 2012, 37(3):225-227. LI Yue-ting, LIU Ying, WANG Jing-xian, et al. Damage effect of hydrogen peroxide on primary cardiac myocytes and H9C2 cells [J]. Journal of Guiyang Medical College, 2012,37(3):225-227.(in Chinese)
[10] 葛苗苗. 蜂胶活性成分抗心肌细胞氧化性损伤的作用及机制研究[D]. 福州:福建农林大学,2014. GE Miao-miao. Protective effects and mechanisms of bioactive compounds in propolis on the oxidative injury of cardiomyocytes [D]. Fuzhou:Fujian Agriculture and Forestry University, 2014. (in Chinese)
[11] TAN G, ZHU Z, JING J, et al. Characterization of constituents in Sini decoction and rat plasma by high-performance liquid chromatography with diode array detection coupled to time-of-flight mass spectrometry[J]. Biomed Chromatogr, 2011, 25(8):913-924.
[12] TAN G, ZHU Z, ZHANG H, et al. Analysis of phenolic and triterpenoid compounds in licorice and rat plasma by high-performance liquid chromatography diode-array detection, time-of-flight mass spectrometry and quadrupole ion trap mass spectrometry[J]. Rapid Commun Mass Spectrom, 2010, 24(2):209-218.
[13] FARAG M A, PORZEL A, WESSJOHANN L A. Comparative metabolite profiling and fingerprinting of medicinal licorice roots using a multiplex approach of GC-MS, LC-MS and 1D NMR techniques [J]. Phytochemistry, 2012, 76:60-72.
[14] WANG Y, HE S, CHENG X, et al. UPLC-Q-TOF-MS/MS fingerprinting of Traditional Chinese Formula SiJunZiTang [J].J Pharm Biomed Anal, 2013, 80:24-33.
[15] YAN Y, CHAI C Z, WANG D W, et al. HPLC-DAD-Q-TOF-MS/MS analysis and HPLC quantitation of chemical constituents in traditional Chinese medicinal formula Ge-Gen Decoction [J]. J Pharm Biomed Anal, 2013, 80:192-202.
[16] MONTORO P, MALDINI M, RUSSO M, et al. Metabolic profiling of roots of liquorice (Glycyrrhiza glabra) from different geographical areas by ESI/MS/MS and determination of major metabolites by LC-ESI/MS and LC-ESI/MS/MS[J].J Pharm Biomed Anal, 2011, 54(3):535-544.
[17] 陈海明, 叶攀. 丹参酮ⅡA对血管内皮细胞氧化应激损伤的保护作用[J].中药材, 2008, 31(4):569-572. CHEN Hai-ming, YE Pan. Protective effects of Tanshinone II A on oxidative stress injury in vascular endothelial cells[J]. Journal of Chinese Medicinal Materials, 2008, 31(4):569-572.(in Chinese)
[18] TANG X Q, FENG J Q, CHEN J, et al. Protection of oxidative preconditioning against apoptosis induced by H2O2 in PC12 cells:mechanisms via MMP, ROS, and Bcl-2 [J]. Brain Res, 2005, 1057(1-2):57-64.
[19] PENNA C, MANCARDI D, RASTALDO R, et al. Cardio protection:a radical view free radicals in pre and post conditioning [J]. Biochim Biophys Acta, 2009, 1787(7):781-793.
[20] 张明发, 沈雅琴. 甘草酸类和甘草黄酮类化合物对心脏保护作用的研究进展[J]. 现代药物与临床, 2012, 27(4):429-434. ZHANG Ming-fa, SHEN Ya-qin. Advances in studies on cardio protection of glycyrrhizic acid compound and flavonoids [J]. Drugs & Clinic, 2012, 27(4):429-434. (in Chinese)
[21] 李珂, 孙兰军, 高克俭, 等. 通脉养心丸治疗冠心病室性早搏(气阴两虚证)多中心临床研究[J]. 中西医结合心脑血管病杂志, 2010, 8(4):401-403. LI Ke, SUN Lan-jun, GAO Ke-jian, et al. TongmaiYangxin Pill for treatment of ventricular premature in coronary heart disease with deficiency syndrome of both Qi and Yin:a multi-center clinical trial [J]. Chinese Journal of Integrative Medicine on Cardio-/Cerebrovascular Disease, 2010, 8(4):401-403. (in Chinese)
[22] 李美平, 何承敏, 胡继鹰, 等. 甘草汤对大鼠心肌缺血再灌注损伤的影响[J]. 湖北中医药大学学报, 2011, 13(1):9-11. LI Mei-ping, HE Cheng-min, HU Ji-ying, et al. The effects of Gancao decoction on myocardial ischemia reperfusion injury in rats[J].Journal of Hubei University of Chinese Medicine, 2011, 13(1):9-11.(in Chinese)
[23] 李作吉, 薛令辉, 杜晓红, 等. 五味子颗粒改善急性心肌梗死恢复期患者心功能的临床观察[J]. 中国医疗前沿, 2013, 8(4):22-23. LI Zuo-ji, XUE Ling-hui, DU Xiao-hong, et al. Clinical observation of schisandra chinensis particle improve acute myocardial infarction reconvalescent cardiac function [J].National Medical Frontiers of China, 2013, 8(4):22-23. (in Chinese)
[24] 李映红, 李湘楚, 罗德生, 等. 五味子提取液对动物缺氧及心肌缺血的保护作用[J]. 咸宁医学院学报, 1998, 12(2):79-82. LI Ying-hong, LI Xiang-chu, LUO De-sheng, et al. The protective effect of SCE on animal pneumatorexis and myocardial ischemia [J]. Journal of Xianning Medical College, 1998, 12(2):79-82. (in Chinese)
[25] 马艳春, 朱丹妮, 余伯阳. 麦冬水提物抗急性心肌缺血活性部位的初步筛选[J]. 时珍国医国药, 2013, 24(3):561-563. MA Yan-chun, ZHU Dan-ni, YU Bo-yang. Preliminary screening of the active components of the aqueous extract of Ophiopogon japonicus on actute myocardial ischemia[J]. Lishizhen Medicine and Materia Medica Research, 2013, 24(3):561-563. (in Chinese)

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