Introduction: Plant extracts have antimicrobial properties and can be substituted as chemical flavorings in food. Preservatives are compounds that are used to inhibit the growth or to eliminate harmful microorganisms to increase the shelf life of foods. Nowadays, the use of natural preservatives derived from medicinal plants is increasing due to their greater compatibility and the presence of volatile compounds (Celiktas et al., 2007). The antimicrobial properties, antioxidant and anticancer effects of these natural additives have been proven (Tajkarimi et al., 2010). In this study, the antimicrobial activity enhancement effect of the combination of hydroalcoholic extracts of basil, cinnamon and salvia on Escherichia coli, Staphylococcus aureus and Saccharomyces cerevisiae yeast in Iranian yoghurt drink (doogh) was investigated for 4 weeks at ambient temperature.
Materials and Methods: The milk was boiled in hygienic conditions at 95 ° C for 5 min. After cooling to 45 ° C, they were inoculated. The aerial parts of the studied plants were milled and powdered. Cinnamon, basil and salvia extracts were then soaked. The food strains of Staphylococcus aureus ATCC 25923, Escherichia coli ATCC 700728 and Saccharomyces cerevisiae PTCC 5193 were prepared as lyophilized ampoules. The microdilution broth method was used to determine the minimum inhibitory concentration. The minimum bactericidal concentration (MBC) was determined in accordance with the minimum inhibitory concentration (MIC) results. To determine the antimicrobial effect of the extracts on the yoghurt drink, 9 ml of the prepared yoghurt drink was poured into each test tube. Then 1 cc of each extract and 1 cc of microbial suspension (1.5 × 108 CFU / ml) were added. One tube was considered as positive control (bacteria + yoghurt drink), one negative control tube (extract + yoghurt drink) and one control tube (yoghurt drink only) and the tubes were placed at ambient temperature. Then, on day 7, 14, 21 and 30 during storage, 20 µl of each tube, were cultured on Muller Hinton agar medium and the same amount of yeast was cultured on blood agar medium. After incubation for 24 h at 37 ° C, the colonies were counted. The interaction of antimicrobial activity of the combination of basil, cinnamon and sage extracts was evaluated on the basis of differential inhibitory concentration index (FIC).
Results and discussion: The results showed that the minimum inhibitory concentration of the combined basil, cinnamon and sage extracts against Staphylococcus aureus and Saccharomyces cerevisiae and Escherichia coli were 0.4, 0.4 and 3.12 mg / ml, respectively. Also, Staphylococcus aureus was more susceptible to the extracts, which were used alone, and was inhibited by a lower concentration of used extract (Ahmadi et al., 2018). However, due to the maximum minimum inhibitory concentration (MIC), Escherichia coli was less sensitive to the extracts which was used alone. The combined usage of all three extracts significantly reduced the maximum minimum inhibitory concentration (MIC) and the minimum bactericidal concentration (MBC), indicating that their antimicrobial activity was more effective against the studied microorganisms than each of the extracts which was used alone. The results of this study showed that synergistic effect against Staphylococcus aureus and Saccharomyces cerevisiae was more effective with the combination of the aforementioned extracts (p <0.05). The results of the Escherichia coli population counting during storage indicated that most of the extracts had a good effect on preventing the growth of Escherichia coli in the doogh environment compared to the control sample (p <0.05). Addition of the extract significantly prevented pH reduction over time (p <0.05), so that the control sample had the highest and the sample containing all three extracts showed the lowest pH changes over time (p <0.05). Therefore, it can be concluded that acid production by Lactobacillus bacteria in samples containing extract during storage period was higher than control. The results of this study were in agreement with the results of Mortazavian et al., (2007) and Voosogh et al., (2009). The acidity of the doogh samples increased significantly over time (P<0.05), while the addition of the extract significantly prevented the acidity increase over time (P<0.05). Sensory properties are the main factors of acceptance and satisfaction of many products. In this study, the mean viewpoint of doogh samples containing the combination of the three above mentioned extracts in terms of taste, odor, color and general acceptance are acceptable. Additionally, it was found that about 90% of participants were willing to use it. Although studies of the overlapping or synergistic effects of medicinal plant extracts in real food environments are not extensive, their results have shown that, the addition of different medicinal plants compounds can increase antibacterial effects. Therefore, the simultaneous use of these extracts can be used in effective control of food pathogens as a complementary method. The results of this study were in agreement with the results of Karimi (2007).