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The objective of this study was to determine whether chitosan (poly-␤-1,4-glucosamine) and hydrolysates of chitosan can be used as novel preservatives in foods. Chitosan was hydrolyzed by using oxidative-reductive degradation, crude papaya latex, and lysozyme. Mild hydrolysis of chitosan resulted in improved microbial inactivation in saline and greater inhibition of growth of several spoilage yeasts in laboratory media, but highly degraded products of chitosan exhibited no antimicrobial activity. In pasteurized apple-elderflower juice stored at 7°C, addition of 0.3 g of chitosan per liter eliminated yeasts entirely for the duration of the experiment (13 days), while the total counts and the lactic acid bacterial counts increased at a slower rate than they increased in the control. Addition of 0.3 or 1.0 g of chitosan per kg had no effect on the microbial flora of houmous, a chickpea dip; in the presence of 5.0 g of chitosan per kg, bacterial growth but not yeast growth was substantially reduced compared with growth in control dip stored at 7°C for 6 days. Improved antimicrobial potency of chitosan hydrolysates like that observed in the saline and laboratory medium experiments was not observed in juice and dip experiments. We concluded that native chitosan has potential for use as a preservative in certain types of food but that the increase in antimicrobial activity that occurs following partial hydrolysis is too small to justify the extra processing involved.Chitosan (poly-␤-1,4-glucosamine) is prepared commercially by alkaline deacetylation of chitin obtained from the exoskeletons of marine crustaceans (6, 17). Chitosan has a pK a value of approximately 6.3 (18); at lower pH values, the molecule is cationic due to protonation of the amino groups. Previous reports have indicated that when chitosan is dissolved in saline, distilled water, or laboratory media, it exhibits antimicrobial activity against some strains of filamentous fungi (2, 9, 15), yeasts (15, 18), and bacteria (21). Considerable variation in the MICs and/or minimum bactericidal concentrations of chitosan, both between and within genera, has been described. Nevertheless, on the basis of the available evidence, bacteria appear to be generally less sensitive to the antimicrobial action of chitosan than fungi are. The antifungal activity of chitosan is greater at lower pH values (15).Chitosan has a mean molecular mass of up to 1 MDa, which corresponds to a chain length of approximately 5,000 U, but there is considerable variation between commercial batches. There is limited in vitro evidence which suggests that partial depolymerization may enhance the antimicrobial activity of chitosan against phytopathogenic fungi and some bacteria of medical significance. For example, Uchida et al. (20) produced chitosan hydrolysates with total reducing sugar (TRS) contents of 50, 508, and 590 mg/g by using Bacillus chitosanase. The antifungal activities of the hydrolysates were tested on solid laboratory media at pH 6 by using three Fusarium spp. isolates. The MI...

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The effects of carvacrol, a natural biocide, on dual-species biofilms formed by Staphylococcus aureus and Salmonella enterica serovar Typhimurium were investigated with a constant-depth film fermentor. Biofilm development reached a quasi-steady state in 12 days at 25°C with S. aureus predominance (Ϸ99%). Cryosectional analysis detected viable S. aureus and S. enterica serovar Typhimurium at depths of 320 and 180 m from the film surface, respectively. Carvacrol pulses (1.0 mmol/h) inhibited S. aureus by 2.5 log CFU/biofilm during the early stages of film formation, ultimately causing a significant reduction (P < 0.001) of the staphylococcal population at quasi-steady state. Initial carvacrol pulsing elicited a 3 log CFU/biofilm reduction in viable S. enterica serovar Typhimurium, and additional periodic carvacrol pulses instigated significant inhibition of salmonellae (1 to 2 log CFU/biofilm) during biofilm development. Carvacrol pulsing reduced protein levels fivefold (P < 0.001) during initial biofilm development. Comparative studies with a peroxide-based commercial sanitizer (Spor-Klenz RTU) revealed that this commercial sanitizer was more biocidal than carvacrol during early biofilm development. When the biofilm reached quasi-steady state, however, periodic pulses with 1 mmol of carvacrol per h (P ‫؍‬ 0.021) elicited a significantly higher inhibition than Spor-Klenz RTU (P ‫؍‬ 0.772). Dual-species microcolonies formed under the influence of continuously fed low carvacrol concentrations (1.0 mmol/h) but failed to develop into a mature quasi-steady-state biofilm and did not reach any stage of film formation in the presence of high concentrations (5.0 mmol/h). These data show that carvacrol is an effective natural intervention to control dual-species biofilm formation.