Berberine–INF55 hybrids are a promising class of antibacterials that combine berberine and the NorA multidrug resistance pump inhibitor INF55 (5-nitro-2-phenylindole) together in one molecule via a chemically stable linkage. Previous studies demonstrated the potential of these compounds for countering efflux-mediated antibacterial drug resistance but they didn’t establish whether the compounds function as originally intended, i.e. with the berberine moiety providing antibacterial activity and the attached INF55 component independently blocking multidrug resistance pumps, thereby enhancing the activity of berberine by reducing its efflux. We hypothesised that if the proposed mechanism is correct, then hybrids carrying more potent INF55 pump inhibitor structures should show enhanced antibacterial effects relative to those bearing weaker inhibitors. Two INF55 analogues showing graded reductions in NorA inhibitory activity compared with INF55 were identified and their corresponding berberine–INF55 hybrids carrying equivalent INF55 inhibitor structures synthesised. Multiple assays comparing the antibacterial effects of the hybrids and their corresponding berberine–INF55 analogue combinations showed that the three hybrids all show very similar activities, leading us to conclude that the antibacterial mechanism(s) of berberine–INF55 hybrids is different from berberine–INF55 combinations.

小ber碱-INF55杂化物是一类很有前途的抗菌剂，通过化学稳定的键合将小ber碱和NorA多药耐药泵抑制剂INF55（5-硝基-2-苯基吲哚）结合在一起。先前的研究表明，这些化合物具有抵抗外排介导的抗菌药物耐药性的潜力，但它们并未确定化合物是否如最初预期的那样发挥功能，即黄连素部分提供了抗菌活性，并且所附着的INF55成分独立地阻断了多药耐药泵，从而通过降低小ber碱的流出量来增强小ber碱的活性。我们假设，如果所提出的机制是正确的，则与具有较弱抑制剂的那些相比，带有更有效的INF55泵抑制剂结构的杂种应该显示出增强的抗菌作用。鉴定出两个INF55类似物，与INF55相比，它们显示NorA抑制活性逐渐降低，并合成了带有相应INF55抑制剂结构的相应小碱-INF55杂种。多次分析比较了杂种及其相应的小ber碱-INF55类似物组合的抗菌作用，结果表明这三种杂种均显示出非常相似的活性，从而使我们得出结论，小ber碱-INF55杂种的抗菌机理与小ber碱-INF55不同组合。