Identification of the Antimicrobial Effect of Liquorice Extracts on Gram-Positive Bacteria: Determination of Minimum Inhibitory Concentration and Mechanism of Action Using a luxABCDE Reporter Strain

Natural preservatives have been used as alternatives to traditional chemical preservatives; however, a limited number have been commercially developed and many remain to be investigated as sources of safer and effective antimicrobials. In this study, we have been investigating the antimicrobial activity of an extract of Glycyrrhiza glabra (liquorice) that was provided as a waste material from the production of liquorice flavourings for the food industry, and to investigate if this retained the expected antimicrobial activity so it could be used as a natural preservative. Antibacterial activity of liquorice extract was screened for evidence of growth inhibition against eight species of Gram-negative and Gram-positive bacteria, including Listeria monocytogenes, Listeria innocua, Staphylococcus aureus, Enterococcus faecalis and Bacillus subtilis. The Gram-negative bacteria tested include Pseudomonas aeruginosa, Escherichia coli and Salmonella typhimurium but none of these were affected by the extract. In contrast, for all of the Gram-positive bacteria tested, growth was inhibited as monitored using optical density. However parallel studies using viable count indicated that the cells were not killed meaning that the extract was bacteriostatic rather than bacteriocidal. The Minimum Inhibitory Concentration [MIC] and Minimum Bactericidal Concentration [MBC] of the extract was also determined and a concentration of 50 µg ml-1 was found to have a strong bacteriostatic effect on Gram-positive bacteria. Microscopic analysis indicated that there were changes in cell shape suggesting the cell wall was affected. In addition, the use of a reporter strain of Listeria transformed with the bioluminescence genes luxABCDE indicated that cell energy levels were reduced when treated with either 12.5 or 50 µg ml-1 of the extract, with the reduction in light output being proportional to the concentration of the extract used. Together these results suggest that the extract is inhibiting the growth of Gram-positive bacteria only by damaging the cell wall and/or membrane.

Quality of Bali Beef and Broiler after Immersion in Liquid Smoke on Different Concentrations and Storage Times

The aim of this study was to improve the durability and quality of Bali beef (M. Longissimus dorsi) and broiler carcass through the addition of liquid smoke as a natural preservative. This study was using Longissimus dorsi muscle from male Bali beef aged 3 years, broiler breast and thigh aged 40 days. Three types of meat were marinated in liquid smoke with concentrations of 0, 5, and 10% for 30 minutes at the level of 20% of the sample weight (w/w). The samples were storage at 2-5°C for 1 month. This study designed as a factorial experiment 3 x 3 x 4 based on a completely randomized design with 5 replications; the first factor was meat type (beef, chicken breast and chicken thigh); the 2nd factor was liquid smoke concentrations (0, 5, and 10%), and the 3rd factor was storage duration (1, 2, 3, and 4 weeks). Parameters measured were TBA value, total bacterial colonies, water holding capacity (WHC), shear force value both before and after cooking (80°C – 15min.), and cooking loss. The results showed that the type of meat produced WHC, shear force value, cooking loss and TBA differed between the three types of meat. Higher concentration of liquid smoke, the WHC, shear force value, TBA, and total bacterial colonies were decreased; at a concentration of 10% of liquid smoke, the total bacterial colonies decreased by 57.3% from untreated with liquid smoke. Longer storage, the total bacterial colonies and WHC were increased, while the shear force value and cooking loss were decreased. It can be concluded that a 10% concentration of liquid smoke was able to maintain fat oxidation and bacterial growth in Bali beef and chicken breast and thigh.