丹参中丹参酮提取工艺优化及活性研究
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摘要
采用超临界CO2提取法对丹参中丹参酮的提取工艺进行优化。通过液质联用法(LC-MS)对丹参粗提物进行成分分析,鉴定出丹参提取物中的4种主要成分分别为丹参酮IIA、丹参酮I、隐丹参酮和二氢丹参酮;采用了AKTA纯化系统对丹参粗提取物进行进一步的分离,利用高效液相色谱(HPLC)鉴定4种丹参酮纯度分别为:丹参酮IIA96.685%,丹参酮I93.083%,隐丹参酮94.968%和二氢丹参酮99.621%。CFU实验和MIC试验结果表明,4种丹参酮均表现出不同程度的抑菌效果,其中隐丹参酮的抑菌能力最强。选用丹参中含量最高的活性成分丹参酮IIA作为抗癌活性研究对象,抗癌活性实验中经过流式细胞仪检测,证明丹参酮IIA-P188具有抑制癌细胞生长的作用,且抗癌活性随药物浓度上升而提高。
In this paper, supercritical CO2 extraction method was used to optimize the extraction of tanshinone from Salvia miltiorrhiza. Liquid chromatography-mass spectrometry(LC-MS) was used to analyze the crude extracts of Salvia miltiorrhiza, and four main components in Salvia miltiorrhiza extract were identified as tanshinone IIA, tanshinone I, cryptotanshinone and dihydrotanshinone. The AKTA purification system was used to further separate the crude Salvia miltiorrhiza extract. High performance liquid chromatography(HPLC) results showed that the four main tanshinone components were well separated, and the purity of the four tanshinones determined by HPLC was: tanshinone IIA, 96.685%; tanshinone I, 93.083%; cryptotanshinone, 94.968%; dihydrotanshinone, 99.621%. The results of CFU and MIC experiments showed that the four tanshinones had different degrees of bacteriostatic effect, of which cryptotanshinone had the strongest antibacterial ability. The highest content of tanshinone IIA in Salvia miltiorrhiza was used for analysis of anticancer activity tested by flow cytometry, and it was confirmed that tanshinone IIA-P188 had an inhibiting effect on the growth of cancer cells, and the anticancer activity increased with the increase of the drug concentration.
In this paper, supercritical CO2 extraction method was used to optimize the extraction of tanshinone from Salvia miltiorrhiza. Liquid chromatography-mass spectrometry(LC-MS) was used to analyze the crude extracts of Salvia miltiorrhiza, and four main components in Salvia miltiorrhiza extract were identified as tanshinone IIA, tanshinone I, cryptotanshinone and dihydrotanshinone. The AKTA purification system was used to further separate the crude Salvia miltiorrhiza extract. High performance liquid chromatography(HPLC) results showed that the four main tanshinone components were well separated, and the purity of the four tanshinones determined by HPLC was: tanshinone IIA, 96.685%; tanshinone I, 93.083%; cryptotanshinone, 94.968%; dihydrotanshinone, 99.621%. The results of CFU and MIC experiments showed that the four tanshinones had different degrees of bacteriostatic effect, of which cryptotanshinone had the strongest antibacterial ability. The highest content of tanshinone IIA in Salvia miltiorrhiza was used for analysis of anticancer activity tested by flow cytometry, and it was confirmed that tanshinone IIA-P188 had an inhibiting effect on the growth of cancer cells, and the anticancer activity increased with the increase of the drug concentration.
引文
[1]王炜辰,吴学辉,郑芳.丹参药理学研究进展[J].海峡药学, 2013, 25(10):24-25.
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[9]LI L N. Water soluble components of Danshen[M]//LI L N.DanShen(Salviamiltiorrhiza)inmedicine.Dordrecht:Springer Netherlands, 2014:97-117
[10]姜雪,史磊.丹参活性成分及药理作用研究进展[J].药学研究, 2017, 36(3):166-169.
[11]梁勇,羊裔明,袁淑兰.丹参酮药理作用及临床应用研究进展[J].中草药, 2000, 31(4):304-306.
[12]FRANZONIF,QUI?ONES-GALVANA,REGOLIF,et al. A comparative study of the in vitro antioxidant activity of statins[J]. Int J Cardiol, 2003, 90(2/3):317-321.
[13]李宣怡.丹参的化学成份分离及其抗氧化活性分析[J].科技资讯, 2017, 15(31):213-214.
[14]LUQ,ZhangF R,ZHANGX,et al.Experimentalstudy oftheanti-cancermechanismoftanshinoneIIAagainst humanbreastcancer[J].IntJMolMed,2009,24(6):773-780
[15]SU C C, LIN Y H. Tanshinone IIA inhibits human breast cancer cells through increased Bax to Bcl-xL ratios[J]. Int J Mol Med, 1998, 22:357-361
[16]HE J, ZHOU Q, YUAN S, et al. Apoptosis-inducing effect oftanshinoneanditsmolecularmechanismonhuman lung cancer cells. Zhongguo Feiai Zazhi 2002, 5(4):257-259.
[17]LIU C, LI J Q, WANG L J, et al. Analysis of tanshinone IIAinducedcellularapoptosisinleukemiacellsbygenome-wideexpressionprofiling[J].BMCComplement Altern Med, 2012, 12:491.
[18]何金涛,周清华,袁淑兰,等.丹参酮对人肺癌细胞株的增殖抑制作用及其分子机理[J].中国肺癌杂志,2002,5(2):123-125.
[19]ZHANG Z C, GAO J, WANG Y Y, et al. Tanshinone IIA triggers p53 responses and apoptosis by RNA polymerase II upon DNA minor groove binding[J]. Biochem Pharmacol, 2009, 78(10):1316-1322.
[2]SUGIYAMA A, ZHU B M, TAKAHARA A, et al. Cardiac effectsofsalviamiltiorrhiza/dalbergiaodoriferamixture,an intravenously applicable Chinese medicine widely used for patients with ischemic heart disease in China[J]. Circ J,2002, 66(2):182-182.
[3]杨瑞峰.复方丹参滴丸治疗冠状动脉粥样硬化性心脏病心绞痛临床疗效观察[J].河南中医,2013,33(1):128-129.
[4]李朝霞,王地.丹参水溶性成分的研究进展[J].北京中医, 2004, 23(3):176-178.
[5]LIU W, ZHANG L, ZHOU J, et al. Comparison of active constituentsatdifferentgrowthstagesinSalviamiltiorrhizaBge.andSalviamiltiorrhizaBge.f.alba[J].Plant Sci J, 2013, 31(6):596-602.
[6]宋凯凯,翟莲.丹参水溶性成分的研究进展[J].医药前沿, 2014(7):126-127.
[7]LI W, ZHOU S P, JIN Y P, et al. Salvianolic acids T and U:Apairofatropisomerictrimericcaffeicacidsderivatives fromrootofSalviamiltiorrhiza[J].Fitoterapia,2014,98:248-253.
[8]季庆亮.高效液相色谱法分析丹参水溶性成分研究概况[J].中国新技术新产品, 2011(4):194.
[9]LI L N. Water soluble components of Danshen[M]//LI L N.DanShen(Salviamiltiorrhiza)inmedicine.Dordrecht:Springer Netherlands, 2014:97-117
[10]姜雪,史磊.丹参活性成分及药理作用研究进展[J].药学研究, 2017, 36(3):166-169.
[11]梁勇,羊裔明,袁淑兰.丹参酮药理作用及临床应用研究进展[J].中草药, 2000, 31(4):304-306.
[12]FRANZONIF,QUI?ONES-GALVANA,REGOLIF,et al. A comparative study of the in vitro antioxidant activity of statins[J]. Int J Cardiol, 2003, 90(2/3):317-321.
[13]李宣怡.丹参的化学成份分离及其抗氧化活性分析[J].科技资讯, 2017, 15(31):213-214.
[14]LUQ,ZhangF R,ZHANGX,et al.Experimentalstudy oftheanti-cancermechanismoftanshinoneIIAagainst humanbreastcancer[J].IntJMolMed,2009,24(6):773-780
[15]SU C C, LIN Y H. Tanshinone IIA inhibits human breast cancer cells through increased Bax to Bcl-xL ratios[J]. Int J Mol Med, 1998, 22:357-361
[16]HE J, ZHOU Q, YUAN S, et al. Apoptosis-inducing effect oftanshinoneanditsmolecularmechanismonhuman lung cancer cells. Zhongguo Feiai Zazhi 2002, 5(4):257-259.
[17]LIU C, LI J Q, WANG L J, et al. Analysis of tanshinone IIAinducedcellularapoptosisinleukemiacellsbygenome-wideexpressionprofiling[J].BMCComplement Altern Med, 2012, 12:491.
[18]何金涛,周清华,袁淑兰,等.丹参酮对人肺癌细胞株的增殖抑制作用及其分子机理[J].中国肺癌杂志,2002,5(2):123-125.
[19]ZHANG Z C, GAO J, WANG Y Y, et al. Tanshinone IIA triggers p53 responses and apoptosis by RNA polymerase II upon DNA minor groove binding[J]. Biochem Pharmacol, 2009, 78(10):1316-1322.
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