In a multi-layer AM packaging system, an AM agent has to migrate across different phases, from the AM layer into the controlling layer, before it is released from the film system onto the food medium. The AM layer must be comprised of a material that has high capability to retain the AM agent in the matrix while the controlling layer is usually comprised of different materials in order to restrict the migration of an AM agent. In this research, a coating material comprised of EA-MMA is shown to perform satisfactorily as an AM layer. Furthermore, the results show that LDPE is capable of controlling the release of an AM agent into a food simulant. Without the LDPE controlling layer, direct contact with a food simulant at the surface of the AM layer results in a high rate of release of AM agent analogous to the release by a monolithic system. For a multi-layer AM packaging system, the retention of the AM agent behind the controlling or protecting layer can promote not only the slower release but also a longer period of AM activity.
5.1.1 Activity of natural AM agent and coating formulation
The AM agents carvacrol and linalool exhibited positive antimicrobial activity against E. coli at a density of 105 cfu mL-1 whereas methylchavicol showed a weaker activity in solid media. These AM agents were successfully incorporated in a MC-HPMC coating solution at a maximum level of 4% (w/w). A dried coating containing the maximum concentration of carvacrol showed a stronger AM activity against 105 cfu mL-1E. coli when compared with the activity of the coating that contained linalool. In order to obtain higher stability in aqueous food media, the hydrophobic EA-MMA coating was selected to replace the hydrophilic MC-HPMC coating. Following the evaporation of the solvent from the EA-MMA coating solution during the drying process, ca. 95% (w/w) of the carvacrol was retained in the film product. The absolute concentration of carvacrol was ca. 30% (w/w) in the dry coating equating to ca. 0.2 g carvacrol per 1 cm3 of dry coating. Multi-layer AM films comprised of nylon/LDPE/EA-MMA/LDPE incorporated with carvacrol were successfully produced. The release of AM agent was studied in order to investigate the effects of the thickness of controlling and AM layers as well as solubility. In this study, a
coating system that contains natural AM agents applied to an existing, commercially available packaging film was successfully formulated and the produced films can adequately retain the incorporated AM substances.
5.1.2 Effect of controlling layer thickness on AM agent release
The slow release of AM agent from the films occurred before the release reached equilibrium therefore the diffusion of AM agent is controlled by the thickness of the LDPE layer. In a food simulant such as ethanol that can effectively dissolve many active agents, equilibrium was achieved when the concentration of carvacrol in the LDPE controlling layer reached the solubility limit. Consequently, the diffusion of carvacrol through the controlling layer was restricted, which maintained the level of the AM agent in the EA-MMA layer. As expected, the release rate was found to be a function of temperature. After observing the release for 1000 h at 4°C, the multi-layer films continued releasing the AM agent without approaching equilibrium. At 20°C, however, the release was approaching equilibrium and at 37°C the system had reached equilibrium. The equilibrium concentration of carvacrol in ethanol obtained from the AM film with a 40 µm thick AM layer and a 50 µm thick controlling layer was 7.4 × 10-4 g cm-3 while there was 2.4 × 10-2 g carvacrol left in the AM layer.
The release of AM agent to the atmosphere was studied by FT-IR spectroscopy. The IR absorbance band at wavenumber 3450 cm-1 was successfully used to determine the
retention of carvacrol in the AM film. The results of the IR study showed that the AM agent could migrate through the LDPE to the atmosphere, indicating the capability of the film to release the active agent to a packaging headspace. Furthermore, the diffusion rate was controlled by the thickness of the LDPE controlling layer. The effect of the thickness of the controlling layer on the release of the incorporated AM agents into selected food simulants was successfully investigated.
5.1.3 Effect of AM layer thickness on AM agent release
In an application where the AM agent is a volatile substance and there is a potential loss from evaporation, a higher amount of the agent must be incorporated in the AM layer to maintain the activity of the film. A thicker AM layer, however, could
achieved a higher equilibrium concentration of AM agent in the AM film. Under the same release conditions at 20°C, a 40 µm thick AM layer retained 1.4 × 10-2 g carvacrol left and provided an equilibrium concentration of 4.2 × 10-4 g cm-3 in ethanol whereas a 20 µm thick AM layer retained 3.6 × 10-3 g carvacrol with an equilibrium concentration of 2.2 × 10-4 g cm-3. The effect of AM layer thickness on the release of the incorporated AM agents into selected food simulants was successfully investigated.
5.1.4 Effect of solubility of AM agent on release
In a compatible food simulant that is a good receptor, the AM agent present in the controlling layer can be released almost entirely into the receptor phase. Therefore, the release of the agent will be dominated by the controlling layer. Conversely, in a poor receptor the release is restricted by the solubility limit of the AM agent in receptor phase. Under the same release conditions at 20°C, the AM film with a 40 µm thick AM layer yielded equilibrium concentrations of 4.2 × 10-4 g cm-3 in ethanol and 1.2 × 10-4 g cm-3 in water. In this case, ethanol is a more compatible food simulant for carvacrol than water.
5.1.5 Antimicrobial activity of multi-layer AM films
In food media, the AM films produced in this work demonstrated satisfactory inhibition of the growth of E. coli at 8°C and 12°C. However, since a higher temperature accelerates the microbial growth, the activity of the films was negligible at 20°C. On Cheddar cheese, the multi-layer AM films successfully inhibited the growth of E. coli with initial cell density of 4.3 log10 cfu g-1. The films reduced the
population to ca. 2.4 and 1.9 log10 cfu g-1 after storage at 8°C and 12°C respectively
for 360 h.