Antimicrobial Activity of Chitosan-Based Films Enriched with Green Tea Extracts on Murine Norovirus, Escherichia coli, and Listeria innocua

Abstract

Edible films can be designed to serve as carriers of antimicrobial agents and be used to control pathogenic foodborne viruses and bacteria. This research tested this concept by dissolving green tea extract (GTE) in chitosan film-forming solutions (FFS) and using it to prepare dried chitosan edible films. As a control, the GTE was also dissolved in deionized water (DW). The FFS and the dried chitosan films with the GTE and the DW without chitosan were all evaluated against murine norovirus (MNV-1), Escherichia coli K12, and Listeria innocua. Both the FFS and the DW with GTE were incubated with ~10⁷ PFU/ml of the virus suspensions for 3 h. The chitosan films with GTE were incubated for 4 and 24 h at 23 ± 1°C. The results showed that the DW containing 1, 1.5, and 2.5% aqueous GTE, significantly (p < 0.05) reduced MNV-1 plaques by 1.7, 2.5, and 3.3 logs after 3 h exposure, respectively. Similarly, FFS containing 2.5 and 5.0% GTE reduced MNV-1 counts by 2.5 and 4.0 logs, respectively, after 3 h exposure. The dried chitosan films with 5, 10, and 15% GTE were also effective against MNV-1 infectivity. After 24 h incubation, the 5 and 10% chitosan GTE films produced significant (p < 0.05) titer reductions of 1.6 and 4.5 logs, respectively. Chitosan films containing 15% GTE reduced MNV-1 plaques to undetectable levels in 24 h. All chitosan GTE films reduced E. coli K12 and L. innocua populations to undetectable levels in tryptic soy broth after 24 h exposure. The results of this study showed that edible films enriched with GTE have potential to reduce both foodborne viruses and bacteria.

Figure 1

Flow chart of chitosan/GTE blend and film formation.

Figure 2

Infectivity of MNV-1 after treatment with different levels of aqueous solution of green tea extracts (GTE) and Dulbecco's Modified Eagle Medium (DMEM). Reduction in MNV-1 infectivity was detected by plaque assay after 3 h of incubation at 23 ± 1°C. Error bars indicate standard deviation.

Figure 3

Infectivity of MNV-1 after treatment with chitosan (CHI) film-forming solutions containing green tea extracts and Dulbecco's Modified Eagle Medium (DMEM). Reduction in MNV-1 infectivity was detected by plaque assay after 3 h of incubation at 23 ± 1°C. Error bars indicate standard deviation.

Figure 4

Infectivity of MNV-1 after treatment with chitosan (CHI) films with different levels of green tea extracts. Reduction in MNV-1 infectivity was detected by plaque assay after 4 and 24 h of incubation at 23 ± 1°C. Error bars indicate standard deviation.

Figure 5

The antibacterial activity of chitosan films incorporated with green tea extract. The effect of green tea extracts incorporated into chitosan films on the survival of L. innocua was determined in tryptic soy broth at 37°C. Error bars indicate standard deviation.

Figure 6

The antibacterial activity of chitosan films incorporated with green tea extract. The effect of green tea extracts (GTE) incorporated into chitosan (CHI) films on the survival of E. coli K12 was determined in tryptic soy broth at 37°C. Error bars indicate standard deviation.