Enhancing Antibacterial Efficacy: Synergistic Effects of Citrus aurantium Essential Oil Mixtures against Escherichia coli for Food Preservation

Ines Ellouze; Boutheina Ben Akacha; Ivana Generalić Mekinić; Rania Ben Saad; Miroslava Kačániová; Maciej Ireneusz Kluz; Wissem Mnif; Stefania Garzoli; Anis Ben Hsouna

doi:10.3390/foods13193093

Enhancing Antibacterial Efficacy: Synergistic Effects of Citrus aurantium Essential Oil Mixtures against Escherichia coli for Food Preservation

Foods. 2024 Sep 27;13(19):3093. doi: 10.3390/foods13193093.

Authors

Affiliations

¹ Department of Vegetal Biotechnology, Higher Institute of Biotechnology of Beja, Jendouba University, Beja 9000, Tunisia.
² Functional and Bio-Resources Valorization Laboratory, Higher Institute of Biotechnology of Beja, Jendouba University, Beja 9000, Tunisia.
³ Laboratory of Biotechnology and Plant Improvement, Center of Biotechnology of Sfax, B.P 1177, Sfax 3018, Tunisia.
⁴ Department of Food Technology and Biotechnology, Faculty of Chemistry and Technology, University of Split, R. Boškovića 35, 21000 Split, Croatia.
⁵ Institute of Horticulture, Faculty of Horticulture, Slovak University of Agriculture, Tr. A. Hlinku2, 94976 Nitra, Slovakia.
⁶ School of Medical and Health Sciences, University of Economics and Human Sciences in Warsaw, Okopowa 59, 01043 Warszawa, Poland.
⁷ Department of Chemistry, College of Sciences at Bisha, University of Bisha, P.O. Box 199, Bisha 61922, Saudi Arabia.
⁸ Department of Chemistry and Technologies of Drug, Sapienza University, P.le Aldo Moro 5, 00185 Rome, Italy.
⁹ Department of Environmental Sciences and Nutrition, Higher Institute of Applied Sciences and Technology of Mahdia, University of Monastir, Monastir 5000, Tunisia.

Abstract

Essential oils (EOs) from various medicinal and aromatic plants are known for their diverse biological activities, including their antimicrobial effects. Citrus aurantium EO is traditionally used for therapeutic benefits due to its high content of bioactive compounds. Therefore, this study focuses on its potential use as a food preservative by investigating the combined antibacterial properties of EOs from leaves (EO1), flowers (EO2), and small branches (EO3) of Citrus aurantium against six bacterial strains by the agar disk diffusion, minimum inhibitory concentration (MIC), and minimum bactericidal concentration (MBC) methods. The chemical compositions of the EOs were analysed by gas chromatography-mass spectrometry (GC-MS) and revealed the presence of numerous compounds responsible for their antimicrobial properties. The MIC values for the EOs were 3.12 mg/mL, 4.23 mg/mL, and 1.89 mg/mL, for EO1, EO2 and EO3, respectively, while the MBC values were 12.5 mg/mL, 6.25 mg/mL, and 6.25 mg/mL, respectively. A simplex centroid design was created to analyse the effect of the individual and combined EOs against E. coli. The combined EOs showed enhanced antibacterial activity compared to the individual oils, suggesting a synergistic effect (e.g., trial 9 with an MIC of 0.21 mg/mL), allowing the use of lower EO concentrations and reducing potential negative effects on food flavour and aroma. Additionally, the practical application of investigated EOs (at concentrations twice the MIC) was investigated in raw chicken meat stored at 4 °C for 21 days. The EOs, individually and in combination, effectively extended the shelf life of the meat by inhibiting bacterial growth (total bacterial count of less than 1 × 10⁴ CFU/g in the treated samples compared to 7 × 10⁷ CFU/g in the control on day 21 of storage). The study underlines the potential of C. aurantium EOs as natural preservatives that represent a sustainable and effective alternative to synthetic chemicals in food preservation.

Keywords: Escherichia coli; bitter orange; chemical composition; essential oils; statistical modelling; synergistic effects; ternary mixture.

Grants and funding

APVV-20-0058/This research was funded by the grant APVV-20-0058 "The potential of the essential oils from aromatic plants for medical use and food preservation".