Abstract The objective of this study was to evaluate the influence of salt concentration on action of natamycin on important microorganisms in food manufacturing as this preservative has been used in immersion baths in several dairy products in the country. Strains of Candida albicans, Escherichia coli and Staphylococcus aureus were inoculated at different saline and peptone water concentrations and received natamycin treatments. These solutions were maintained at 12 °C and the behavior of the microorganisms evaluated at 0, 24 and 48 hours (T0, T1 and T2). Each microorganism was assessed in isolation as well as the association of C. albicans and E. coli. Under the conditions proposed by the research, it was possible to conclude that 0.025% natamycin has no efficacy on C. albicans inoculated at saline concentrations below 5%. The results obtained in E. coli counts suggest that natamycin may interfere with its development even at concentrations that may be considered low (0.1%) and at salinity conditions of 7.5% to 10%. The association of natamycin with sodium chloride potentiates its antimicrobial action, which can represent an economy and its use is amplified by the industries.