Thin-layer chromatographic (TLC) separations and bioassays of plant extracts to identify antimicrobial compounds.
Thin-layer chromatographic (TLC) separations and bioassays of plant extracts to identify antimicrobial compounds.
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DOI：
10.3791/51411
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发表时间：
2014-03-27
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影响因子：
--
通讯作者：
Flythe, Michael D
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文献类型：
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作者：
Kagan, Isabelle A;Flythe, Michael D
植物抗微生物化合物的常见筛选包括通过纸或薄层色谱（PC或TLC）分离植物提取物，将色谱图暴露于微生物悬浮液（例如肉汤或琼脂中的真菌或细菌），为微生物留出时间在潮湿的环境中生长，并观察没有微生物生长的区域。这种被称为生物自显影的筛选方法的有效性取决于色谱分离的质量和对微生物培养条件的关注。本文介绍了标准的协议，薄层色谱和接触生物自显影与新的应用氨基酸发酵细菌。在柔性（铝背衬）硅胶TLC板上分离提取物，并在紫外（UV）光下观察条带。切出区域并面朝下在接种有测试微生物的琼脂上孵育。通过用四唑红对琼脂平板染色来观察抑制带。将该方法应用于红三叶草（Trifolium pratense cv. Kenland）的酚类化合物及其抗斯氏梭菌（Clostridium sticklandii）活性的筛选，斯氏梭菌是一种产氨细菌（HAB），其是牛瘤胃的原生菌。TLC方法适用于许多类型的植物提取物和其他细菌物种（需氧或厌氧），以及真菌，如果培养条件进行了修改，以适应物种的生长要求，可以用作测试生物。
A common screen for plant antimicrobial compounds consists of separating plant extracts by paper or thin-layer chromatography (PC or TLC), exposing the chromatograms to microbial suspensions (e.g. fungi or bacteria in broth or agar), allowing time for the microbes to grow in a humid environment, and visualizing zones with no microbial growth. The effectiveness of this screening method, known as bioautography, depends on both the quality of the chromatographic separation and the care taken with microbial culture conditions. This paper describes standard protocols for TLC and contact bioautography with a novel application to amino acid-fermenting bacteria. The extract is separated on flexible (aluminum-backed) silica TLC plates, and bands are visualized under ultraviolet (UV) light. Zones are cut out and incubated face down onto agar inoculated with the test microorganism. Inhibitory bands are visualized by staining the agar plates with tetrazolium red. The method is applied to the separation of red clover (Trifolium pratense cv. Kenland) phenolic compounds and their screening for activity against Clostridium sticklandii, a hyper ammonia-producing bacterium (HAB) that is native to the bovine rumen. The TLC methods apply to many types of plant extracts and other bacterial species (aerobic or anaerobic), as well as fungi, can be used as test organisms if culture conditions are modified to fit the growth requirements of the species.