薛东芳;邹宗尧;陈彪;王燕枝;吴昊;叶小利;李学刚;
XUE Dong-fang;ZOU Zong-yao;CHEN Biao;WANG Yan-zhi;WU Hao;YE Xiao-li;LI Xue-gang;School of Life Science,Southwest University;Chongqing Pharmaceutical Process and Quality Control Engineer Technology Research Center;College of Pharmaceutical Sciences,Southwest University;

• 1:西南大学生命科学学院
• 2:重庆药物过程与质量控制工程技术研究中心
• 3:西南大学药学院

摘要(Abstract)：

为探索黄连总生物碱及小檗碱对嗜水气单胞菌的抑菌活性及作用机制,测定了黄连总生物碱和小檗碱对嗜水气单胞菌的最小抑菌浓度、膜通透性、膜流动性、膜蛋白构象及毒力因子的影响。结果显示,黄连总生物碱和小檗碱对嗜水气单胞菌均有抑菌活性,其最小抑菌浓度分别为62.5,125 mg·L-1。黄连总生物碱和小檗碱能增加膜脂流动性,改变膜蛋白构象,并使菌体膜通透性分别增加24.52%,19.66%。此外,黄连总生物碱和小檗碱显著性地抑制菌体外毒素的溶血活性及aer A和hly A mRNA的表达(P<0.05,P<0.01),同时降低嗜水气单胞菌内毒素的分泌量和Lpx C mRNA的表达(P<0.05,P<0.01)。结果表明,黄连总生物碱和小檗碱对嗜水气单胞菌的抑菌作用可能与细胞膜损伤有关,通过改变膜脂流动性和膜蛋白构象使菌体膜通透性增加从而抑制了细菌生长;同时降低了细菌的毒力因子的分泌,使得菌体的致病性降低。
To explore the antibacterial activity and mechanism of total alkaloids and berberine from Coptidis Rhizoma on Aeromonas hydrophila,and determine the effect of total alkaloids and berberine from Coptidis Rhizoma on minimum inhibitory concentrations,permeability and fluidity of cell membrane,conformation of membrane proteins and virulence factors of A. hydrophila. The results showed that both total alkaloids and berberine from Coptidis Rhizoma had antibacterial activities on A. hydrophila,with minimum inhibitory concentrations of 62. 5 and 125 mg·L- 1,respectively. Total alkaloids and berberine from Coptidis Rhizoma could increase the fluidity of membrane,change the conformation of membrane proteins and increase the permeability of bacteria membrane by 24. 52% and 19. 66%,respectively. Besides,total alkaloids and berberine from Coptidis Rhizoma significantly decreased the hemolysis of exotoxin and the mRNA expressions of aer A and hly A( P < 0. 05,P < 0. 01),the secretion of endotoxin and the mRNA expression of Lpx C( P < 0. 05,P <0. 01). The results suggested that the antibacterial activity of total alkaloids and berberine from Coptidis Rhizoma on A. hydrophila may be related to the bacteria membrane injury. They inhibited the bacterial growth by increasing membrane lipid fluidity and changing conformation of membrane proteins,and reduced the secretion of virulence factors of A. hydrophila to weaken the pathogenicity.

关键词(KeyWords)： 黄连生物碱;嗜水气单胞菌;抑菌机制
alkaloids from Coptidis Rhizoma;Aeromonas hydrophila;antibacterial mechanism

Abstract:

Keywords:

基金项目(Foundation): 国家“十二五”科技支撑计划项目(2011BAI13B02-1);; 重庆市教委重点项目(KJZH14203);; 教育部博士点基金(20130182110023);; 重庆市百名高端工程技术人才培养计划(2013-2016)

作者(Author): 薛东芳;邹宗尧;陈彪;王燕枝;吴昊;叶小利;李学刚;
XUE Dong-fang;ZOU Zong-yao;CHEN Biao;WANG Yan-zhi;WU Hao;YE Xiao-li;LI Xue-gang;School of Life Science,Southwest University;Chongqing Pharmaceutical Process and Quality Control Engineer Technology Research Center;College of Pharmaceutical Sciences,Southwest University;

Email:

DOI:

参考文献(References)：

• [1]Vivas J,Carracedo B,Riao J,et al.Behavior of an Aeromonas hydrophila aro A live vaccine in water microcosms[J].Appl Environ Microbiol,2004,70(5):2702.
• [2]Singh V,Chaudhary D K,Mani I,et al.Development of diagnostic and vaccine markers through cloning,expression,and regulation of putative virulence-protein-encoding genes of Aeromonas hydrophila[J].J Microbiol,2013,51(3):275.
• [3]Vaseeharan B,Thaya R.Medicinal plant derivatives as immunostimulants:an alternative to chemotherapeutics and antibiotics in aquaculture[J].Aquacul Int,2014,22:1079.
• [4]Su H C,Ying G G,Tao R,et al.Occurrence of antibiotic resistance and characterization of resistance genes and integrons in Enterobacteriaceae isolated from integrated fish farms in south China[J].J Environ Monit,2011,13(11):3229.
• [5]Su Z,Xiao H,Chen L.In vitro antibacterial effect of Chinese herbal medicine against Aeromonas hydrophila[J].Inf Techn Agricul Engin,2012,134(1):233.
• [6]方忻平,王天志,张浩.国产黄连5种生物碱的含量测定[J].中国中药杂志,1989,14(2):33.
• [7]Zhang D,Li A,Xie J,et al.In vitro antibacterial effect of berberine hydrochloride and enrofloxacin to fish pathogenic bacteria[J].Aquacul Res,2010,41(7):1095.
• [8]何爱华,张曦,陶琳丽,等.6种中草药对4种淡水鱼致病菌体外抑菌作用的研究[J].南方水产科学,2011,7(2):73.
• [9]Ye X,Li X,Yuan L,et al.Interaction of houttuyfonate homologues with the cell membrane of gram-positive and gram-negative bacteria[J].Colloids Surf A:Physicochem Engin Aspects,2007,301(1):412.
• [10]Ye X,Li X,Yuan L,et al.Effect of the surface activity on the antibacterial activity of octadecanoyl acetal sodium sulfite series[J].Colloids Surf A:Physicochem Engin Aspects,2005,268(1):85.
• [11]Ye X,Li X,Yuan L,et al.Interaction between houttuyfonate homologues and erythrocyte plasma membrane of rabbit in vitro[J].Colloids Surf A:Physicochem Engin Aspects,2006,279(3):218.
• [12]陶慰孙,李惟,姜涌明.蛋白质分子基础[M].2版.北京:高等教育出版社,1995:260.
• [13]李晔,戴小寒,田武林,等.中草药连翘提取物抑制金黄色葡萄球菌α-溶血素分泌的研究[J].中国兽医学报,2013,33(3):404.
• [14]刘雪锋,夏咏梅,方云,等.中药黄连有效成分盐酸小檗碱与牛血清白蛋白的相互作用[J].高等学校化学学报,2004,25(11):2099.
• [15]M T马迪根,J M马丁克.微生物生物学[M].11版.北京:科学出版社,2009:95.
• [16]刘梦茵,刘芳,周涛,等.乌梅提取物对蜡状芽孢杆菌的抑菌机制研究[J].食品科学,2012,33(1):103.
• [17]Chapkin R S,Hong M Y,Fan Y Y,et al.Dietary n-3 PUFA alter colonocyte mitochondrial membrane composition and function[J].Lipids,2002,37(2):193.
• [18]Stoll B A.N-3 fatty acids and lipid peroxidation in breast cancer inhibition[J].Br J Nutr,2002,87(3):193.
• [19]Nissen-Meyer J,Rogne P,Oppegard C,et al.Structure function relationships of the non-lanthionine-containing peptide(classⅡ)bacteriocins produced by gram-positive bacteria[J].Curr Pharm Biotechn,2009,10(1):19.
• [20]邱军强,杨先乐,程训佳.嗜水气单胞菌毒力因子特性及作用机理研究进展[J].中国病原生物学杂志,2009,4(8):616.
• [21]麻莉,刘友生.抗内毒素治疗研究进展[J].中华烧伤杂志,2004,19(3):190.