Antibacterial activity of leaf essential oils of Citrus crenatifolia and Citrus reticulata: a comparative study

Abstract

Plant essential oils (EOs) are smart alternatives to synthetic antibiotics. Citrus crenatifolia (CC, Heen naran), a native plant to Sri Lanka and Citrus reticulata (CR, Mandarin) are two species of genus Citrus in the family Rutaceae. Both species are widely used in traditional medicine in Sri Lanka. This study aimed to determine the antibacterial activity of pure and combined formulae of EOs of CC and CR (1:1 ratio) against Staphylococcus aureus, Escherichia coli and Bacillus cereus bacterial species. Hydrodistillation was used to collect CC and CR leaf EOs. Kirby-Bauer disc diffusion method was used to assess antimicrobial susceptibility to EOs. Amoxicillin (0.01mg/mL for E. coli and S. aureus, 0.1mg/mL for B. cereus) was used as a positive control in the experimental setup. Diameters of inhibition zones were measured after 24-hour incubation at 37ºC. Descriptive and inferential statistical analyzes were performed using R studio version 4.1.1. Least significance test determined the differences at p<0.05. The CC EO showed significantly strong antibacterial activity against B. cereus compared to amoxicillin, CR and CC+CR and against S. aureus compared to amoxicillin. This is suggestive that a synergistic effect drawn by the presence of high concentrations of antibacterial compounds in CC EO. In contrast, CR EO showed significant and moderately strong growth inhibition of S. aureus and E. coli compared to that of B. cereus. Low CR effectiveness in B. cereus could be due to weaker CR penetration, cell wall thickening and reduced hydrophobicity. The CC+CR combination also showed significant, moderate growth inhibition of all three bacterial strains. B. cereus and E. coli showed significantly higher inhibition on CC+CR, however, S. aureus showed least susceptibility. In conclusion, both CC and CR EOs demonstrated potent antibacterial effects against the three tested bacterial species whereas CC+CR combination showed additive effect.