一种高效率筛选抗菌中药的微量点菌药敏实验新方法  

An efficient new method of antimicrobial susceptibility testing for screening Chinese herbal medicine

作　　者：邹文政[1] 李忠琴[1] 赖晓健[1] 林茂[1] 江兴龙[1] 

机构地区：[1]集美大学水产学院鳗鲡现代产业技术教育部工程研究中心厦门市渔用药物工程技术研究中心,福建厦门361021

出　　处：《饲料工业》2016年第24期57-61,共5页Feed Industry

基　　金：福建省教育厅项目[JA15260];厦门市重点项目[3502Z20110014];鳗鲡现代产业技术教育部工程研究中心开放基金[RE201606;RE201607];大学生创新创业训练计划项目[201510390053]

摘　　要：通常检测药物抗菌活性的方法有试管二倍稀释法、牛津杯法、滤纸片法、平板二倍稀释法等,在筛选具有抗菌活性的中药时,均需预先粗提药材的有效部位,并去除萃取溶剂,操作繁琐、耗时长、效率低。自然界中药用植物种类繁多、数量庞大,为了能够快速高通量筛选其中的抗菌药材,本研究建立了一种简便易行的药敏实验方法。实验利用超微粉碎机将15种中药药材粉碎成粒径为5~10μm微粉后,将药粉按一定比例添入M-H琼脂培养基内,经过121℃高压灭菌20 min,制成不同浓度梯度的固体药敏培养基,再在同一个药敏琼脂平板上同时接种9株致病菌;培养24 h后,没有供试菌菌落形成的最低浓度为该中药的最低抑菌浓度（MIC）,此方法称为超微药粉琼脂稀释法。采用该方法测得中药抑菌浓度与同种药材粗提液的滤纸片法和平板二倍稀释法相比,超微药粉琼脂稀释法测得中药的抑菌效果更显著,三种方法检测获得致病菌对各种中药敏感程度的级别基本一致。三种方法的比较结果说明,超微药粉琼脂稀释法的药敏实验结果准确、灵敏度高,而且操作简便、快捷、耗时短,易于判断;并且一个药敏琼脂平板可同时测试多株致病菌,处理量大,效率高。本法可作为一种从天然植物药材中广泛初步筛选具有抗菌活性中药的高效方法,在实验研究中具有很好的实用价值,为抗菌中药开发应用提供基础的参考数据。Methods of antimicrobial activity test, conmonly used for detecting Chinese medicine in vitro commonly include tube double dilution method, oxford cup method, filter paper and plate double dilution method. In the above mentioned methods, however, effective ingredients of Chinese herbs have to be extracted at first in a rough way, and then the extraction solvent must be removed as well, so the operation process is cumbersome, time-consuming and low efficiency. There are a huge numbers of various pharmaceutical plants in nature, and a simple and convenient method of drug susceptibility testing has been developed in this study so that antibacterial herbs can be screened out fast and high-throughput. In the study, 15 kinds of Chinese herbs were first ground into ultrafine powder of 5~10 μm particles, which then were added to Mueller-Hinton Broth to prepare the medium drug nutrient agar plate with different concentration gradient.After that, the plates were sterilized at 121 ℃ for 20 min and then nine different strains of pathogens were inoculated in the same drug agar plate for 24 h. The minimum inhibitory concentration（MIC）, under which the colonies of bacteria could not been formed, was recorded to evaluate antimicrobial activity of different herbs. The method is named as superfine powder agar dilution method. Compared with filter paper method and plate double dilution method of crude extracts from herbal botanicals, the superfine powder agar dilution method possessed smaller MIC under the same kinds of botanicals. So it is obvious that the method proposed in this paper is more sensitive than other two else, though the sensitivity level of pathogens on Chinese medicine obtained by our method is still consistent with the results of other two methods. The experiments results indicate that ultrafine powder agar dilution method is simple, less time-consuming, high efficiency and high-throughput. And susceptibility test results are accurate, sensitive and easy to observe. Moreover the drug sensitivity test of mor

关 键 词：中药药材 超微药粉琼脂稀释法 抗菌活性 最低抑菌浓度 

分 类 号：S816.17[农业科学—饲料科学]