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The demand for natural antimicrobials as food preservatives has increased due to the growing interest of the population for a healthy lifestyle. The application of screening methods to identify the antimicrobial activity of natural compounds is of great importance. The in vitro determination of antimicrobial activity requires determining their minimum inhibitory concentrations to assess microbial susceptibility. This study aimed to evaluate the minimum inhibitory concentrations of three natural antimicrobial compounds-chitosan, ethanolic propolis extract, and nisin-against 37 microorganisms (different pathogens and spoilage microorganisms) by the methods of agar dilution and drop diffusion on agar. Culture media at different pH values were used for both methods to simulate different food products. Most of the microorganisms were inhibited by chitosan (0.5% w/v ) and propolis (10 mg/mL), and most of the Gram-positive bacteria by nisin (25 μg/mL). Different pH values and the in vitro method used influenced the inhibition of each compound. Generally, lower minimum inhibitory concentrations were observed at lower pH values and for the agar dilution method. Furthermore, some microorganisms inhibited by the compounds on the agar dilution method were not inhibited by the same compounds and at the same concentrations on the drop diffusion technique. This study reinforces the need for using defined standard methods for the in vitro determination of minimum inhibitory concentrations. Natural compounds with potential antimicrobial action are a bet on food preservation. The use of standard techniques such as those used for antimicrobials of clinical applications are crucial to compare results obtained in different studies and different matrices.

The Inhibitory Concentration of Natural Food Preservatives May Be Biased by the Determination Methods