This study investigated the relationship of arginine deiminase (ADS) and urease activities with dental caries through a case-control study. ADS and urease activities were measured in dental smooth-surface supragingival plaque and whole saliva samples from 93 subjects, who were in three different groups: caries-free (n = 31), caries-active (n = 30), and caries-experienced (n = 32). ADS activity was measured by quantification of the ammonia generated from the incubation of plaque and saliva samples in a mixture containing 50 mM arginine-HCl and 50 mM Tris-maleate buffer, pH 6.0. ADS-specific activity was defined as nanomoles of ammonia generated per minute per milligram of protein. Urease activity was determined by quantification of ammonia produced from 50 mM urea. For bacterial identification and enumeration real-time qPCR analysis was used. Groups were compared using Kruskal-Wallis tests. Spearman correlations were used to analyze plaque metabolic activity and bacterial relationships. The results revealed significantly higher ammonia production from arginine in saliva (1.06 vs. 0.18; p < 0.0001) and plaque samples (1.74 vs. 0.58; p < 0.0001) from caries-free subjects compared to caries-active subjects. Urease levels were about 3-fold higher in the plaque of caries-free subjects (p < 0.0001). Although higher urease activity in saliva of caries-experienced and caries-free subjects was evident, no significant difference was found between the groups.