ABSTRACT
Antimicrobial resistance (AMR) is a global public health threat caused by the misuse and overuse of antibiotics. It leads to infections becoming difficult to treat, causing serious illness, disability, and death. Current antibiotic development is slow, with only 25% of current antibiotics exhibiting novel mechanisms against critical pathogens. Traditional medicinal plants' secondary metabolites offer potential for developing novel antibacterial compounds. These compounds, often with strong antimicrobial activity, can be used to develop safe and effective antibacterial chemotherapies. This study investigated the antibacterial activity of Phyllanthus niruri Linn. extracts against a panel of bacterial pathogens using disc diffusion and microdilution assays and quantified by calculation of minimum inhibition concentration (MIC). Additionally, the effects of combinations of the extracts and selected conventional antibiotics were examined by sum of fractional inhibition concentration (Æ©FIC) calculation and isobologram analysis. Liquid chromatography-mass spectrometry (LC-MS) phytochemistry analysis was used to identify noteworthy compounds in the active extracts and the Artemia nauplii bioassay was used to evaluate toxicity. The aqueous and methanolic extracts exhibited notable antibacterial activity in the broth microdilution assay against Staphylococcus aureus and methicillin-resistant S. aureus (MRSA) (MIC = 669 µg/mL and 738 µg/mL, respectively). The methanolic extract also showed noteworthy antibacterial action in the broth assay against Klebsiella pneumoniae (MIC = 738 µg/mL). The aqueous extract had noteworthy growth inhibitory activity against Bacillus cereus (MIC = 669 µg/mL), whilst the methanolic extract demonstrated good antibacterial activity against that bacterium (MIC = 184 µg/mL). The aqueous and methanol extracts showed minimal antibacterial action against Shigella flexneri and Shigella sonnei. The extracts were subjected to LC-MS analysis, which revealed several interesting phytochemicals, including a variety of flavonoids and tannins. The antibacterial activity and lack of toxicity of the P. niruri extracts indicates that they may be worthwhile targets for antibiotic development and further mechanistic and phytochemistry studies are required.
ABSTRACT
ETHNOPHARMACOLOGICAL RELEVANCE: Triphala is used in Ayurveda to treat a wide variety of diseases, including numerous bacterial infections. Interestingly, the plant components of triphala (Terminalia bellirica, Terminalia chebula and Emblica officinalis) are also good inhibitors of bacterial growth when used individually, yet plant preparations are generally used in combination in traditional medicine. Surprisingly, no previous studies have addressed the reason why the combination is preferred over the individual components to treat bacterial infections. AIM OF THE STUDY: To test and compare the antibacterial efficacy of triphala and its component parts to quantify their relative efficacies. The individual plant components will also be tested as combinations, thereby determining whether combining the individual components potentiates the antibacterial activity of the components used alone. MATERIALS AND METHODS: Triphala and the three individual plant components were extracted using solvents of varying polarity (methanol, water, ethyl acetate) and the antibacterial activity of the aqueous resuspensions was quantified by disc diffusion and broth microdilution MIC assays. Combinations of extracts produced from the individual components were also tested against each bacterial species and the ΣFICs was calculated to determine the class of interaction. Where synergy was detected, isobologram analysis was used to determine the optimal component ratios. The Artemia nauplii bioassay was used to test for toxicity and GC-MS headspace profiling analysis was used to highlight terpenoid components that may contribute to the antibacterial activity of triphala. RESULTS: The aqueous and methanolic triphala, T. bellirica, T. chebula and E. officinalis extracts displayed good inhibitory activity against all bacterial strains, with MICs often in the 250-750 µg/mL range. The methanolic extracts were generally more potent than the aqueous extracts and T. chebula was the most potent of the individual plant components. Combining the extracts of the different plant species resulted in potentiation of the growth inhibitory activity of most combinations compared to that of the individual components. Indeed, with the exception of S. flexneri, all bacterial species were potentiated by at least one combination of methanolic plant extracts, with a substantial proportion of these displaying synergistic interactions. All extracts were found to be either non-toxic, or of low to moderate toxicity in Artemia nauplii assays. CONCLUSION: Whilst the individual plant components of triphala all inhibit the growth of multiple pathogenic bacteria, the activity is potentiated for multiple combinations. Therefore, the traditional usage of the combination of the three plant materials in triphala not only extends the activity profile of the mixture over that of the individual components, but it also substantially potentiates the inhibitory activity towards multiple bacteria, partially explaining the preference of triphala compared to the individual components.