黄芩对肺炎克雷伯菌抑制作用及其机制研究  

Analysis of the bacteriostatic action of Scutellaria baicalensis on Klebsiella pneumoniae and the mechanisms of that action

作　　者：殷姿[1] 欧宜文 李蓓[1] 杨靖[1] 欧琴[1] 

机构地区：[1]湖北医药学院基础医学院,湖北十堰442000 [2]湖北医药学院第一临床学院

出　　处：《中国病原生物学杂志》2016年第5期388-392,共5页Journal of Pathogen Biology

基　　金：湖北医药学院硕士研究启动基金项目(No.2014QDJ18);湖北省2011协同创新中心基金项目(项目编号4)

摘　　要：目的观察黄芩对肺炎克雷伯菌的抑制作用,分析其抗菌机制,为寻找治疗肺炎克雷伯菌感染新途径提供实验依据。方法制备黄芩药液,以多重耐药肺炎克雷伯菌为试验菌株,采用试管法测定黄芩液对细菌的最低抑菌浓度(MIC)和最小杀菌浓度(MBC);通过细菌生长曲线和生物膜形成情况评价黄芩液对细菌生长的影响。不同浓度黄芩液作用后,取培养液上清进行电导率和碱性磷酸酶活性测定,分析其对细菌细胞膜通透性的影响;采用细菌沉淀荧光法进行核酸DNA和RNA含量及SDS-PAGE蛋白谱带定量分析;提取DNA拓扑异构酶,进行相关生物学活性测定,分析其对细菌合成的影响。结果黄芩液对肺炎克雷伯菌的MIC和MBC分别为(32±2.2)mg/ml和(64±3.6)mg/ml。不同浓度黄芩液均可抑制肺炎克雷伯菌生长和细胞生物膜形成,使细胞膜通透性增加。黄芩液作用24h后肺炎克雷伯菌菌体蛋白、DNA和RNA减少。黄芩能抑制DNA拓扑异构酶Ⅰ和Ⅱ活性,且呈浓度依赖。结论黄芩可通过抑制肺炎克雷伯菌生物膜形成抑制其生长,通过增加细胞膜通透性和抑制细菌DNA拓扑异构酶活性,进而抑制细菌核酸和蛋白合成,发挥抑制细菌繁殖和侵袭作用。Objectives To investigate the mechanisms by which Scutellaria baicalensis acts on Klebsiella pneumoniae and to analyze its antibacterial action. Methods A solution of S.baicalensis was prepared and tested on multi-drug-resistant K.pneumoniae.The tube dilution method was used to evaluate the minimal inhibitory concentration（MIC）and minimum bactericidal concentration（MBC）of S.baicalensis.The bacterial growth curve and a biofilm formation assay were used to observe the effects of S.baicalensis on bacterial growth.After cultured K.pneumoniae was treated with different concentrations of S.baicalensis,the supernatant was collected and its conductivity and alkaline phosphatase activity were determined.The effect of S.baicalensis on bacterial cell membrane permeability was analyzed.Bacterial sedimentation and fluorescence analysis was used to evaluate nucleic acid content（DNA and RNA）and protein bands in SDSPAGE were quantitatively analyzed.DNA topoisomerase was isolated and its biological activity on K.pneumoniae was studied.Results The MIC for S.baicalensis was 32±2.2mg.ml-1 and the MBC for S.baicalensis was 64±3.6 mg.ml-1.Different concentrations of S.baicalensis markedly inhibited K.pneumoniae growth and biofilm formation.Analysis of the mechanism of action of S.baicalensis indicated that S.baicalensis inhibited K.pneumoniae protein expression and DNA and RNA synthesis.S.baicalensisinhibited DNA topoisomerase I and II activity. Conclusions S.baicalensis markedly inhibited the growth of K.pneumoniae and increased its cell membrane permeability.S.baicalensis inhibited the DNA topoisomerase activity of the bacterium,thereby inhibiting the synthesis of bacterial nucleic acids and proteins.S.baicalensis plays an important role in inhibiting K.pneumoniae proliferation and invasion.

关 键 词：黄芩 肺炎克雷伯菌 抑菌机制 DNA拓扑异构酶 

分 类 号：R378.4[医药卫生—病原生物学]