Optimization of a natural antimicrobial formulation against potential meat spoilage bacteria and food-borne pathogens: Mixture design methodology and predictive modeling

This study is about the combined antimicrobial effect of essential oils (EOs), namely Mediterranean (MN) EO, German thyme (GT) EO, Cinnamon (CN) EO, Indian (IN) EO, Asian (AN) EO, and citrus extract (CE) against spoilage bacteria (Lactobacillus sakei, Lactobacillus curvatus, Leuconostoc mesenteroides, Carnobacterium divergens, Brochothrix thermosphacta, and Pseudomonas aeruginosa) and selected pathogenic bacteria (E. coli O157:H7, Salmonella Typhimurium and Listeria monocytogenes). Firstly, each EO and CE were screened for antibacterial activity by microdilution assay, and the most efficient antimicrobial extracts were selected based on the lowest MIC values to perform the combination assays. Afterward, a simplex-centroid mixture design was used to develop optimal antimicrobial mixtures capable of protecting meat from spoilage and pathogenic bacteria. The optimization tool allowed us to postulate models and validate them statistically as well as to create a prediction profile of the experiment. Thus, the optimal mixtures named active formulation 1 (AF1) containing MN EO/GT EO/VC EO/CE with a ratio of 1:2:2:1 and active formulation 2 (AF2) containing IN EO/AN EO/CE/VC EO with a ratio of 2:2:1:2, were developed based on the demonstration of their synergistic effect against tested bacteria. The obtained formulations at organoleptically acceptable concentrations could be applied in the preservation of meat and meat products.