Citrus limon Essential Oil: Chemical Composition and Selected Biological Properties Focusing on the Antimicrobial (In Vitro, In Situ), Antibiofilm, Insecticidal Activity and Preservative Effect against Salmonella enterica Inoculated in Carrot

Abstract

New goals for industry and science have led to increased awareness of food safety and healthier living in the modern era. Here, one of the challenges in food quality assurance is the presence of pathogenic microorganisms. As planktonic cells can form biofilms and go into a sessile state, microorganisms are now more resistant to broad-spectrum antibiotics. Due to their proven antibacterial properties, essential oils represent a potential option to prevent food spoilage in the search for effective natural preservatives. In this study, the chemical profile of Citrus limon essential oil (CLEO) was evaluated. GC-MS analysis revealed that limonene (60.7%), β-pinene (12.6%), and γ-terpinene (10.3%) are common constituents of CLEO, which prompted further research on antibacterial and antibiofilm properties. Minimum inhibitory concentration (MIC) values showed that CLEO generally exhibits acceptable antibacterial properties. In addition, in situ antimicrobial research revealed that vapour-phase CLEO can arrest the growth of Candida and Y. enterocolitica species on specific food models, indicating the potential of CLEO as a preservative. The antibiofilm properties of CLEO were evaluated by MIC assays, crystal violet assays, and MALDI-TOF MS analysis against S. enterica biofilm. The results of the MIC and crystal violet assays showed that CLEO has strong antibiofilm activity. In addition, the data obtained by MALDI-TOF MS investigation showed that CLEO altered the protein profiles of the bacteria studied on glass and stainless-steel surfaces. Our study also found a positive antimicrobial effect of CLEO against S. enterica. The anti-Salmonella activity of CLEO in vacuum-packed sous vide carrot samples was slightly stronger than in controls. These results highlight the advantages of the antibacterial and antibiofilm properties of CLEO, suggesting potential applications in food preservation.

Keywords: anti-Candida and Salmonella effect; antibacterial activity; antibiofilm activity; carrot; chemical analysis; insecticidal activity; microorganisms; sous vide.

Figure 1

Graphic elaboration of in situ antimicrobial activity (%) in apple, carrot, and kohlrabi.

Figure 2

Representative MALDI-TOF mass spectra of S. enterica: (A) 3rd day, (B) 5th day, (C) 7th day, (D) 9th day, (E) 12th day, and (F) 14th day. SE = S. enterica; G = glass; S = stainless steel; PC = planktonic cells.

Figure 2

Representative MALDI-TOF mass spectra of S. enterica: (A) 3rd day, (B) 5th day, (C) 7th day, (D) 9th day, (E) 12th day, and (F) 14th day. SE = S. enterica; G = glass; S = stainless steel; PC = planktonic cells.

Figure 3

Dendrogram of S. enterica generated using MSPs of the planktonic cells and the control. SE = S. enterica; G = glass; S = stainless steel; PC = planktonic cells.

Figure 4

The outcomes of the total bacterial count. Total bacterial count treated at temperatures ranging between 50 and 65 °C for durations of 5 to 20 min. (expressed in log CFU/g) on the first day. Data are the mean (±SD) of 3 samples. Control: Fresh carrot sample was treated at 50–65 °C for 5 to 25 min after being packed in polyethylene bags and kept at 4 °C. Control vacuum: Fresh carrot sample was treated at 50–65 °C for 5 to 25 min after being vacuum-packed in polyethylene bags and kept at 4 °C. EO: vacuum-packed fresh carrot treated with 1% lime EO was kept at 4 °C and treated for 5–25 min at 50–65 °C. Salmonella: vacuum-packed fresh carrot treated with S. enterica was kept at 4 °C and treated for 5–25 min at 50–65 °C. Salmonella + EO: vacuum-packed fresh carrot treated with S. enterica and 1% lime EO was kept at 4 °C and treated for 5–25 min at 50–65 °C.

Figure 5

The outcomes of the coliform bacteria. Coliform bacteria treated at temperatures ranging between 50 and 65 °C for durations of 5 to 20 min (expressed in log CFU/g) on the seven day. Data are the mean (±SD) of 3 samples. Control: Fresh carrot sample was treated at 50–65 °C for 5 to 25 min after being packed in polyethylene bags and kept at 4 °C. Control vacuum: Fresh carrot sample was treated at 50–65 °C for 5 to 25 min after being vacuum-packed in polyethylene bags and kept at 4 °C. EO: vacuum-packed fresh carrot treated with 1% lime EO was kept at 4 °C and treated for 5–25 min at 50–65 °C. Salmonella: vacuum-packed fresh carrot treated with S. enterica was kept at 4 °C and treated for 5–25 min at 50–65 °C. Salmonella + EO: vacuum-packed fresh carrot treated with S. enterica and 1% lime EO was kept at 4 °C and treated for 5–25 min at 50–65 °C.

Figure 6

Krona chart: Isolated species of carrot sous vide samples of bacteria in % at day 1.

Figure 7

Krona chart: Isolated species of carrots sous vide samples of bacteria in % at day 7.