Uji Aktivitas Antibakteri Ekstrak Etanol Kulit Citrus sp dan Kombinasinya dengan Vancomycin terhadap Bakteri Resisten Escherichia coli

Abstract

The effectiveness of antibiotics is decreasing due to the threat of resistance. Combining natural antibacterial compounds with antibiotics has been proven effective in combating many bacterial strains. Escherichia coli has received special attention due to the high prevalence of resistance in this bacterium. Natural compounds with antibacterial activity are considered a potential option to address resistance issues and improve the efficacy of antibiotics. This research aims to determine the antibacterial activity of ethanol extract from the skin of Citrus fruits and its combination with vancomycin against resistant E. coli bacteria. Extracts were obtained by maceration. Antibacterial activity was determined by microdilution method. The combination effect with vancomycin and extracts were evaluated using checkerboard-assay. Inhibition of biofilm formation, the activity on membrane permeability, and EtBr accumulation assay also investigated. The best Minimum Inhibitory Concentration (MIC) values were obtained at concentrations of 125 μg/mL and 250 μg/mL for the Citrus hystrix peel ethanol extract and Citrus aurantifolia peel ethanol extract, with Minimum Bactericidal Concentration (MBC) values of 250 μg/mL and 500 μg/mL. The Citrus aurantifolia peel ethanol extract and Citrus hystrix peel ethanol extract also showed synergistic effects when combined with vancomycin (FIC Index 0.3 and 0.2). Biofilm inhibition tests showed the best results for the combination of Citrus hystrix peel ethanol extract and vancomycin, with a percentage of 82%. Meanwhile, in the efflux pump inhibition test, the best results were shown by the single concentration of Citrus aurantifolia peel ethanol extract (250 μg/mL). In the membrane permeability test at OD 260 nm and OD 590 nm, the combination of Citrus hystrix peel ethanol extract and vancomycin also showed the best results with the highest absorbance value of 0.8201 and a crystal violet uptake of 58.92%. To support the results of the membrane permeability test, bacterial morphology analysis was performed using Scanning Electron Spectroscopy (SEM), and the leakage of magnesium and calcium ions from inside the test bacterial cell membrane was measured using ICP-OES. The results of both tests showed the best outcome with the combination of kaffir lime ethanol extract and vancomycin. The result of this study was revealed that both extracts increased the antibacterial activity of vancomycin and may be used as an herbal extracts combined with antibiotics will be a promising strategy to overcome antibiotic resistant Escherichia coli