Objective Probiotics participate in regulating oral microbiota and reducing the prevalence of oral diseases; however, clinical research on probiotics is insufficient. Therefore, in this study, we performed in vitro screening of potential oral protective probiotic strains and then evaluated the clinical efficacy of the selected strains on maintaining oral health. Materials and methods Fifty healthy individuals were recruited and randomly assigned into the placebo group and probiotics group, which included three strains of probiotics, Lactobacillus salivarius subs. salicinius AP‐32, Lactobacillus paracasei ET‐66, and Lactobacillus plantarum LPL28. Each group was blindly administered placebo or probiotics for four weeks. Results Next‐generation sequencing results showed that the oral microbiota of Lactobacillus salivarius in the oral cavity were significantly increased in subjects supplemented with mixed probiotic lozenges. The anti‐bacterial activities of viable probiotics were observed within two weeks. Both IgA levels and Lactobacillus and Bifidobacterium abundances in the oral cavity were significantly increased in the experimental groups, along with a reduced formation of plaque. Most participants reported that their oral health conditions and intestinal symptoms had improved. Conclusions Overall, our clinical study suggests that oral probiotic lozenges may enhance oral immunity, modulate oral microbiota, and improve oral health.