Antimicrobial activity of chitosan, organic acids and nano-sized solubilisates for potential use in smart antimicrobially-active packaging for potential food applications

Highlights

• The molecular weight (MW) of the chitosan effected antimicrobial activity (AA).

• Low & medium MW chitosan had the highest AA against all bacteria tested.

• Nano-sized solubilisates of organic acids had higher AA than their non-nano equivalents.

Abstract

Antimicrobial activity of low- and medium-molecular weight chitosan and organic acids (Benzoic acid and Sorbic acid and commercially-available nano-sized benzoic- and sorbic-acid solubilisate equivalents) was investigated and compared against commercial mixtures of organic acids used as meat coatings (Articoat DLP-02^® and Sulac-01^®). From the antimicrobials tested, both low molecular weight (LMW) and medium molecular weight (MMW) chitosan exhibited the highest antimicrobial activity against all bacterial cultures tested, with mean MIC values of 0.010 and 0.015% w/v, respectively. The results suggested that the MW of the chitosan used effected antimicrobial activity of the chitosan. Nano-sized solubilisates of benzoic acid and sorbic acid had significantly (P < 0.05) higher antimicrobial properties than their non-nano equivalents. The results found in this study open opportunities for the nano-sized solubilisates, derived from food compatible sources, to be used in smart and active antimicrobial packaging applications, as less of the antimicrobial substances in question is required to deliver the same antimicrobial effect.

Introduction

There continues to be a demand from both retailers and consumers for further improvement in food quality and safety while still offering acceptably prolonged product shelf-lives (Theron & Lues, 2007). Consequently, there is always a great interest in new smart or active food packaging systems that can meet these demands in an acceptable and cost-effective manner (Cruz-Romero & Kerry, 2011). Developments in the area of smart antimicrobial packaging systems and materials are guaranteed to attract attention due to the potential impact that such materials could have on product safety and shelf-life, if commercially adopted. The food grade antimicrobial agents most commonly utilized at present, particularly in edible food or packaging coatings are: sorbic acid, propionic acid, potassium sorbate, benzoic acid, sodium benzoate and citric acid, bacteriocins, such as nisin and pediocin; enzymes, such as peroxidase and lysozyme; and polysaccharides displaying natural antimicrobial properties, such as chitosan (Chien, Sheu & Yang, 2007; Devlieghere, Vermeulen, & Debevere, 2004; Durango, Soares & Andrade 2006; Kim et al., 2011; Quintavalla & Vicini, 2002) Chitosan offers real potential for applications in the food industry due to its particular physicochemical properties, short time biodegradability, biofunctional, biocompatible, antimicrobial and antifungal activities, and non-toxicity (Aider, 2010; Dutta, Tripathi, Mehrotra, & Dutta, 2009). In recent years much attention has turned to the field of nanotechnology in the quest to improve antimicrobial packaging (Cushen, Kerry, Morris, Cruz-Romero, & Cummins, 2012). Active packaging systems with incorporated antimicrobial and antioxidant agents offers new opportunities to develop novel packaging materials with the potential to maintain food quality and improve food safety and product shelf-lives.

Nanoparticles with bactericidal activity have been immobilized, coated on to surfaces and controlled in terms of their active release (Ruparelia, Chatterjee, Duttagupta, & Mukherji, 2008). The potential application for a given nanoparticle depends on many factors, including; material type (Ren et al., 2009), particle shape (Wang, Xin & Tao, 2005) and usage concentration (Kim, Kim, Cho, & Cho, 2003). The intrinsic properties of a nanoparticle are determined predominantly by its shape, size, composition, crystallinity and morphology (Vigneshwaran, Kumar, Kathe, Varadarajan, & Prasad, 2006).

Chitosan is a linear polysaccharide consisting of (1,4)-linked 2-amino-deoxy-β-d-glucan, is a deacetylated derivative of chitin, which is the second most abundant polysaccharide found in nature after cellulose. Edible film coatings consisting solely of chitosan or consisting of chitosan used in combination with another biopolymer such as sodium caseinate have been applied to carrot, cheese and salami samples (Moreira, Pereda, Marcovich, & Roura, 2011). These authors found that chitosan and sodium caseinate/chitosan films were found to exert significant bactericidal action on mesophilicand psychrotrophic bacteria, as well as causing a reduction in yeast and mould counts in all samples. Greater bactericidal properties were observed in the caseinate/chitosan than in the chitosan alone. This antimicrobial activity has been displayed on a wide selection of foods, including; meat, dairy products and vegetables (Moreira et al., 2011; Suman et al., 2010).

The organic acids sorbic acid, p-aminobenzoic acid, lactic acid, and acetic acid have a long history as generally recognized as safe (GRAS) food preservatives and there is a growing demand for such natural preservatives in the food industry today (Burt, 2004). When used in combination with lactic and/or acetic acid, sorbic acid can inhibit the growth of Listeria monocytogenes, Salmonella typhimurium, and Escherichia coli O157:H7 in many low acid foods, including; cold-pack cheese, bologna and beaker sausage (Cagri, Ustunol, & Ryser, 2001), while p-aminobenzoic acid has been reported to exhibit significant inhibitory activity against L. monocytogenes, E. coli, and Salmonella enteritidis (Richards, 1995). Edible whey protein isolate films incorporating sorbic acid and p-aminobenzoic acid have shown inhibitory action towards of S. typhimurium, L. monocytogenes, and E. coli O157:H7 when placed in direct contact with inoculated culture (Cagri et al., 2001). Ouattara, Simard, Piette, Bégin and Holley (2000) evaluate antimicrobial films prepared by incorporating acetic or propionic acid into a chitosan matrix, with or without the addition of lauric acid or the essential oil, cinnamaldehyde. These films were directly applied to bologna, regular cooked ham or pastrami. Propionic acid was released from the chitosan matrix at a faster rate than acetic acid and the addition of lauric acid, but not cinnamaldehyde, to the chitosan matrix reduced the release of acetic acid. While lactic acid bacteria were not affected by the antimicrobial films under study, the growth of Enterobacteriaceae and Serratia liquefaciens was delayed or completely inhibited as a result of film application. One novel study combined MAP packaging and organic acid incorporation on the preservation of fresh salmon (Schirmer et al., 2009). The salmon was packed with a small amount of 100% CO₂ (gas/product ratio 0.2/1.0 v/v) and a brine solution containing various combinations of citric acid (3% w/w, pH 5), acetic acid (1% w/w, pH 5) and cinnamaldehyde (200 μg ml⁻¹). CO₂, acetic acid and citric acid alone each inhibited the growth of total bacterial counts, lactic acid bacteria, sulphur reducing bacteria and Enterobacteriaceae, but effects were enhanced in combination. It was found that the combination of CO₂ and organic acids completely inhibited bacterial growth during 14 days of storage at 4 °C, both in inoculation experiments and in experiments on salmon possessing a natural background flora.

Where a limited number of studies have examined the effect of chitin/chitosan and organic acids usage in films for the control of microorganisms in food packaging application, to date, little or no evaluation of nanoparticled forms of these substances have been carried out for the same purposes, therefore the objectives of this study were to assess the antimicrobial activity of low- and medium-molecular weight chitosan, and several acids widely used as preservatives (Benzoic acid and Sorbic acid) and compared to their commercially available nano-sized solubilisates against commonly food spoilage microoganisms. A mixture of organic acids used as meat coatings (Articoat DLP-02 and Sulac) were compared as commercial controls.