Small molecule disruption of the bacterial membrane is both a challenge and interest for drug development. While some avoid membrane activity due to toxicity issues, others are interested in leveraging the effects for new treatments. Existing assays are available for measuring disruption of membrane potential or membrane permeability, two key characteristics of the bacterial membrane, however they are limited in their ability to distinguish between these properties. Here, we demonstrate a high throughput assay for detection and characterization of membrane active compounds. The assay distinguishes the effect of small molecules on either the membrane potential or membrane permeability using the fluorescent dyes TO-PRO-3 iodide and DiOC₂(3) without the need for secondary assays. We then applied this assay to a library of 3520 synthetic molecules previously shown to inhibit growth of B. subtilis in order to determine the frequency of membrane activity within such a biologically active library. From the library, we found 249 compounds that demonstrated significant membrane activity, suggesting that synthetic libraries of this kind do not contain a plurality of membrane active molecules.

中文翻译：

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小分子枯草芽孢杆菌生长抑制剂的膜活性谱分析

细菌膜的小分子破坏是药物开发的挑战和兴趣。尽管有些药物由于毒性问题而避免了膜的活性，但另一些药物则有兴趣利用这种作用进行新的治疗。现有的测定法可用于测量膜电位或膜通透性的破坏，这是细菌膜的两个关键特性，但是它们区分这些特性的能力受到限制。在这里，我们展示了用于膜活性化合物检测和表征的高通量检测方法。使用荧光染料TO-PRO-3碘化物和DiOC ₂可以区分小分子对膜电位或膜通透性的影响（3）无需二次测定。然后，我们将这种测定方法应用于3520个先前显示可抑制枯草芽孢杆菌生长的合成分子的文库，以确定这种生物活性文库中膜活性的频率。从该文库中，我们发现了249种具有明显膜活性的化合物，这表明此类合成文库不包含多个膜活性分子。