Preservatives are utilized to inhibit the growth of spoilage and pathogenic microorganisms in food, thus improving shelf life and reducing food waste. Despite the effectiveness of current preservatives, consumer health concerns have spurred interest in natural alternatives. This study explored the effects of traditional preservatives, nisin (N), potassium sorbate (Ps) and protective cultures, Lactiplantibacillus plantarum (Lp), Lacticaseibacillus rhamnosus (Lr) both individually and in various combinations on the physicochemical properties and microbial inhibition of Turkish white cheese over 28 days of storage. To this end, thirteen cheese samples were produced and analyzed: one without preservatives (C), four with a single preservative (N, Ps, Lp, Lr), and eight with combinations (NLr, NLp, NPs, PsLr, PsLp, LpLr, LpLrN, LpLrPs). The use of preservatives did not alter compositional characteristics, but acidity increased rapidly in samples with protective cultures. The combination of N and Ps with protective cultures produced a stronger antimicrobial effect compared to N and Ps alone, while Lp and Lr combinations showed promising results. Inhibition of coliforms and Escherichia coli was achieved using either N or protective cultures alone. The combined use of Lp and Lr was more effective against all analyzed microorganisms than either alone. Sensory analysis indicated no significant difference in overall acceptability between preservative-treated and control samples. These findings highlight the potential of protective cultures as a natural alternative to traditional preservatives and suggest their use in combination can enhance food safety by providing more effective inhibition.