金龙胆草提取液体外抑菌活性分析
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摘要
金龙胆草是传统道地中药材,但目前受到材料来源不足的限制,其应用和研究均受到很大限制,为更好地开发和应用该道地中药材,本研究以实验室野生金龙胆草和组培金龙胆草出发,分别采用热水浴和索式提取法对金龙胆草有效成份进行了提取和体外抑菌活性研究。结果表明,热水浴浸提法的抑菌效果优于索式提取法,均表现出对金黄色葡萄球菌和枯草芽孢杆菌这类革兰氏阳性菌有很好的抑制效果,最小抑菌浓度分别达到12.5 mg/mL和50 mg/mL。野生型提取液和组培型的提取液体外抑菌比较实验显示,野生型金龙胆草的抑菌效果要显著优于组培型金龙胆草提取液。这将为金龙胆草作为中药抗菌药的合理利用提供一定的指导作用,同时为减少抗生素的滥用造成的环境污染作出一定的贡献。
Abstrac t Conyza blinii H. Lév is a traditional Chinese herbal medicine, but its application and research are limited by the shortage of materials. In order to better develop and apply this traditional Chinese medicine,we used wide type and tissue culture of Conyza blinii H. Lév as our experiment materials, respectively, using hot water bath and soxhlet extraction method to extract their active ingredients and carry out in vitro antibacterial activity experiment. The results showed that hot-bath extraction method was more superior of the antibacterial effect than that of soxhlet extraction method, both methods showed Staphylococcus aureus and Bacillus subtilis such Gram-positive bacteria had a good inhibitory effect, the minimum inhibitory concentration was 12.5 mg/mL and50 mg/mL. respectively, the antibacterial activity of the wild-type extract and the tissue culture extract in vitro showed that the antibacterial activity of the wild-type was significantly better than that of the tissue culture extract.The results of this study would provide a guideline for the rational utilization of Conyza blinii H. Lév as an antimicrobial medication for traditional Chinese herb and make some contribution to reducing environmental pollution caused by the abuse of antibiotics.
Abstrac t Conyza blinii H. Lév is a traditional Chinese herbal medicine, but its application and research are limited by the shortage of materials. In order to better develop and apply this traditional Chinese medicine,we used wide type and tissue culture of Conyza blinii H. Lév as our experiment materials, respectively, using hot water bath and soxhlet extraction method to extract their active ingredients and carry out in vitro antibacterial activity experiment. The results showed that hot-bath extraction method was more superior of the antibacterial effect than that of soxhlet extraction method, both methods showed Staphylococcus aureus and Bacillus subtilis such Gram-positive bacteria had a good inhibitory effect, the minimum inhibitory concentration was 12.5 mg/mL and50 mg/mL. respectively, the antibacterial activity of the wild-type extract and the tissue culture extract in vitro showed that the antibacterial activity of the wild-type was significantly better than that of the tissue culture extract.The results of this study would provide a guideline for the rational utilization of Conyza blinii H. Lév as an antimicrobial medication for traditional Chinese herb and make some contribution to reducing environmental pollution caused by the abuse of antibiotics.
引文
Chen H.,Liu S.,Sun R.,Tang Z.Z.,Gao J.L.,Li C.L.,Wu Q.,and Huang H.Y.,2015,SCAR(sequence characterized amplified region)primer and method for identifying felwort and adulterants thereof by using SCAR technology,China Patant,ZL201310373522.1(陈惠,刘姗,孙蓉,唐自钟,高静雷,李成磊,韩学易,吴琦,黄晓燕,2015,一种利用SCAR技术鉴别金龙胆草及其混淆品的SCAR引物及其方法,中国专利,ZL201310373522.1)
Chen L.,2007,Study on chemical composition of low polarity of conyza blinii H.Lév,Thesis for M.S.,Tianjin University,Supervisor:Su Y.F,pp.21-26(陈磊,2007,金龙胆草低极性部分化学成分研究,硕士学位论文,天津大学,导师:苏艳芳,pp.21-26)
Deavours B.E.,and Dixon R.A.,2005,Metabolic engineering of isoflavonoid biosynthesis in alfalfa,Plant Physiol.,138(4):2245-2259
Duan S.Z.,2015,Study on antifungal activity of tobacco extracts,Thesis for M.S.,Biotechnology Research Institute CAAS,Supervisor:Zhang Z.F.,pp.8-13(段苏珍,2015,烟草提取物抑菌活性研究,硕士学位论文,中国农业科学院,导师:张忠锋,pp.8-13)
Fei W.B.,Liang X.X.,and He M.,2016,The changes of polyphenolic contents in two kinds of Terminalia chebula retz,traditional tibetian processing methods,Journal of Chinese Pharmaceutical Sciences,25(2):150-153
Hu J.R.,Jiang H.,and Li X.X.,2013,Study on three non-water-soluble solvents influencing of inhibitory activity in antibacterial test,Xibu Zhongyiyao(Western Journal of Traditional Chinese Medicine),26(11):30-31(胡君茹,姜华,李喜香,2013,三种非水溶性供试品溶剂对抑菌试验效果的影响,西部中医药,26(11):30-31)
Hua G.Q.,2005,Preliminary study on the antimicrobial characteristics and antimicrobial mechanism of berberine,Thesis for M.S.,Shandong University,Supervisor:Jin J.L.,pp.22-27(华国强,2005,小檗碱抑菌特点及抑菌机制的初步研究,硕士学位论文,山东大学,导师:金建玲,pp.31-35)
Liu J.P.,2009,The study on parching process and quality assessment of semen cassiae,Thesis for M.S.,Shandong University,Supervisor:Wang S.Y.,pp.20-31(刘建平,2009,决明子炮制工艺及质量评价方法研究,硕士学位论文,山东大学,导师:王少云,pp.20-31)
Liu H.,2010,Study on antimicrobial effects of Vitex negundo Linn.stem,Thesis for M.S.,Zhejiang University,Supervisor:Shen S.R.,pp.22-24(刘虹,2010,黄荆茎的抑菌作用研究,硕士学位论文,浙江大学,导师:沈生荣,pp.22-24)
Liu P.,Zhou L.J.,Su Y.F.,and Yan S.L.,2011,Effects of total saponins from Conyza blinii on the apoptosis of hela cells and SPC-A1 cells,Zhongguo Yaofang(China Pharmacy),22(35):3288-3291(刘培,周立军,苏艳芳,颜世伦,2011,金龙胆草总皂苷诱导Hela细胞和SPC-A1细胞凋亡的研究,中国药房,22(35):3288-3291)
Ma L.,and Liu J.,2014,The protective activity of Conyza blinii saponin against acute gastric ulcer induced by ethanol,J.Ethnopharmacol.,158:358-363
Ma L.,Liu H.,Qin P.,Hu C.,Man S.,Li Y.,Liu Z.,and Diao A.,2017,Saponin fraction isolated from Conyza blinii H.Lév demonstrates strong anti-cancer activity that is due to its NF-κB inhibition,Biochem.Bioph.Res.Co.,483(1):779-785
Peng J.Y.,Sun R.,Tang Z.Z.,Chen H.,Wang T.,Zheng T.R.Zhang R.M.,Wang Y.X.,and Zhang X.L.,2017,Optimization of extraction process of polysaccharides from Amaranthus hybridus by response surface methodology,Jiyin zuxue Yu Yingyong Shengwuxue(Genomics and Applied Biology)36(2):791-797(彭继燕,孙蓉,唐自钟,陈惠,王涛,郑天润张润敏,王一行,王晓丽,2017,响应面优化绿穗苋多糖的提取工艺,基因组学和应用生物学,36(2):791-797)
Su Y.F.,Liu J.S.,Guo D.A.,Li J.,Guo H.Z.,and Zheng J.H.2001,Flavonoids from aerial parts of Conyza blinii,Zhong caoyao(Chinese Traditional&Herbal Drugs),32(6):496-497(苏艳芳,刘建生,果德安,李菁,郭洪祝,郑俊华,2001,金龙胆草黄酮类成分的研究,中草药,32(6):496-497)
Su Y.F.,Guo Dean.,Guo H.Z.,Liu J.S.,Zheng J.H.,Koike K.and Nikaido T.,2001,Four new triterpenoid saponins from Conyza blinii,J.Nat.Prod.,64(1):32-36
Sun R.,Liu S.,Tang Z.Z.,Jin H.J.,Li C.L.,and Chen H.,2015Study on transcriptome characteristic of genuine traditiona chinese medicine Conyza blinii H.Lév Leaves of Sichuan Fenzi Zhiwu Yuzhong(Molecular Plant Breeding),13(12):2754-2760(孙蓉,刘姗,唐自钟,晋海军,李成磊,陈惠2015,四川道地中药金龙胆草叶片转录组特性研究,分子植物育种,13(12):2754-2760)
Wang H.T.,2009,Study on antimicrobial activity of soybean isoflavones and its mechanism,Thesis for M.S.,Liaoning Normal University,Supervisor:Xie M.J.,pp.11-17(王海涛,2009,大豆异黄酮的抑菌活性及其机制的研究,硕士学位论文,辽宁师范大学,导师:谢明杰,pp.11-17)
Wu Z.H.,Xu Y.,and Wu C.M.,2003,Study on the method of extracting natural preservative in aniseed and its bacteriostasis,Zhongguo Tiaoweipin(China Condiment),29(9):18-20(吴周和,徐燕,吴传茂,2003,八角中天然防腐剂的提取方法及其抑菌作用研究,中国调味品,29(9):18-20)
Yang F.Y.,2007,Studies on chemical constituents and anti-ulcer activity of Conyza blinii H.Lév,Thesis for M.S.,Tianjin University,Supervisor:Su Y.F.,pp.22-28(杨凤英,2007,金龙胆草皂苷化学成分及其抗溃疡活性研究,硕士学位论文,天津大学,导师:苏艳芳,pp.22-28)
Zhang W.,2008,Bacteriostatic activity and mechanism of lemon extract against muitiresistant Staphylococcus aureus,Thesis for M.S.,Tianjin Medical University,Supervisor:Zhong Q.P.,and Li X.X.,pp.21-32(张纬,2008,柠檬提取物对多重耐药金黄色葡萄球菌的抑菌作用及抑菌机制初探,硕士学位论文,天津医科大学,导师:钟启平,李晓霞,pp.21-32)
Chen L.,2007,Study on chemical composition of low polarity of conyza blinii H.Lév,Thesis for M.S.,Tianjin University,Supervisor:Su Y.F,pp.21-26(陈磊,2007,金龙胆草低极性部分化学成分研究,硕士学位论文,天津大学,导师:苏艳芳,pp.21-26)
Deavours B.E.,and Dixon R.A.,2005,Metabolic engineering of isoflavonoid biosynthesis in alfalfa,Plant Physiol.,138(4):2245-2259
Duan S.Z.,2015,Study on antifungal activity of tobacco extracts,Thesis for M.S.,Biotechnology Research Institute CAAS,Supervisor:Zhang Z.F.,pp.8-13(段苏珍,2015,烟草提取物抑菌活性研究,硕士学位论文,中国农业科学院,导师:张忠锋,pp.8-13)
Fei W.B.,Liang X.X.,and He M.,2016,The changes of polyphenolic contents in two kinds of Terminalia chebula retz,traditional tibetian processing methods,Journal of Chinese Pharmaceutical Sciences,25(2):150-153
Hu J.R.,Jiang H.,and Li X.X.,2013,Study on three non-water-soluble solvents influencing of inhibitory activity in antibacterial test,Xibu Zhongyiyao(Western Journal of Traditional Chinese Medicine),26(11):30-31(胡君茹,姜华,李喜香,2013,三种非水溶性供试品溶剂对抑菌试验效果的影响,西部中医药,26(11):30-31)
Hua G.Q.,2005,Preliminary study on the antimicrobial characteristics and antimicrobial mechanism of berberine,Thesis for M.S.,Shandong University,Supervisor:Jin J.L.,pp.22-27(华国强,2005,小檗碱抑菌特点及抑菌机制的初步研究,硕士学位论文,山东大学,导师:金建玲,pp.31-35)
Liu J.P.,2009,The study on parching process and quality assessment of semen cassiae,Thesis for M.S.,Shandong University,Supervisor:Wang S.Y.,pp.20-31(刘建平,2009,决明子炮制工艺及质量评价方法研究,硕士学位论文,山东大学,导师:王少云,pp.20-31)
Liu H.,2010,Study on antimicrobial effects of Vitex negundo Linn.stem,Thesis for M.S.,Zhejiang University,Supervisor:Shen S.R.,pp.22-24(刘虹,2010,黄荆茎的抑菌作用研究,硕士学位论文,浙江大学,导师:沈生荣,pp.22-24)
Liu P.,Zhou L.J.,Su Y.F.,and Yan S.L.,2011,Effects of total saponins from Conyza blinii on the apoptosis of hela cells and SPC-A1 cells,Zhongguo Yaofang(China Pharmacy),22(35):3288-3291(刘培,周立军,苏艳芳,颜世伦,2011,金龙胆草总皂苷诱导Hela细胞和SPC-A1细胞凋亡的研究,中国药房,22(35):3288-3291)
Ma L.,and Liu J.,2014,The protective activity of Conyza blinii saponin against acute gastric ulcer induced by ethanol,J.Ethnopharmacol.,158:358-363
Ma L.,Liu H.,Qin P.,Hu C.,Man S.,Li Y.,Liu Z.,and Diao A.,2017,Saponin fraction isolated from Conyza blinii H.Lév demonstrates strong anti-cancer activity that is due to its NF-κB inhibition,Biochem.Bioph.Res.Co.,483(1):779-785
Peng J.Y.,Sun R.,Tang Z.Z.,Chen H.,Wang T.,Zheng T.R.Zhang R.M.,Wang Y.X.,and Zhang X.L.,2017,Optimization of extraction process of polysaccharides from Amaranthus hybridus by response surface methodology,Jiyin zuxue Yu Yingyong Shengwuxue(Genomics and Applied Biology)36(2):791-797(彭继燕,孙蓉,唐自钟,陈惠,王涛,郑天润张润敏,王一行,王晓丽,2017,响应面优化绿穗苋多糖的提取工艺,基因组学和应用生物学,36(2):791-797)
Su Y.F.,Liu J.S.,Guo D.A.,Li J.,Guo H.Z.,and Zheng J.H.2001,Flavonoids from aerial parts of Conyza blinii,Zhong caoyao(Chinese Traditional&Herbal Drugs),32(6):496-497(苏艳芳,刘建生,果德安,李菁,郭洪祝,郑俊华,2001,金龙胆草黄酮类成分的研究,中草药,32(6):496-497)
Su Y.F.,Guo Dean.,Guo H.Z.,Liu J.S.,Zheng J.H.,Koike K.and Nikaido T.,2001,Four new triterpenoid saponins from Conyza blinii,J.Nat.Prod.,64(1):32-36
Sun R.,Liu S.,Tang Z.Z.,Jin H.J.,Li C.L.,and Chen H.,2015Study on transcriptome characteristic of genuine traditiona chinese medicine Conyza blinii H.Lév Leaves of Sichuan Fenzi Zhiwu Yuzhong(Molecular Plant Breeding),13(12):2754-2760(孙蓉,刘姗,唐自钟,晋海军,李成磊,陈惠2015,四川道地中药金龙胆草叶片转录组特性研究,分子植物育种,13(12):2754-2760)
Wang H.T.,2009,Study on antimicrobial activity of soybean isoflavones and its mechanism,Thesis for M.S.,Liaoning Normal University,Supervisor:Xie M.J.,pp.11-17(王海涛,2009,大豆异黄酮的抑菌活性及其机制的研究,硕士学位论文,辽宁师范大学,导师:谢明杰,pp.11-17)
Wu Z.H.,Xu Y.,and Wu C.M.,2003,Study on the method of extracting natural preservative in aniseed and its bacteriostasis,Zhongguo Tiaoweipin(China Condiment),29(9):18-20(吴周和,徐燕,吴传茂,2003,八角中天然防腐剂的提取方法及其抑菌作用研究,中国调味品,29(9):18-20)
Yang F.Y.,2007,Studies on chemical constituents and anti-ulcer activity of Conyza blinii H.Lév,Thesis for M.S.,Tianjin University,Supervisor:Su Y.F.,pp.22-28(杨凤英,2007,金龙胆草皂苷化学成分及其抗溃疡活性研究,硕士学位论文,天津大学,导师:苏艳芳,pp.22-28)
Zhang W.,2008,Bacteriostatic activity and mechanism of lemon extract against muitiresistant Staphylococcus aureus,Thesis for M.S.,Tianjin Medical University,Supervisor:Zhong Q.P.,and Li X.X.,pp.21-32(张纬,2008,柠檬提取物对多重耐药金黄色葡萄球菌的抑菌作用及抑菌机制初探,硕士学位论文,天津医科大学,导师:钟启平,李晓霞,pp.21-32)