Purpose: To determine if use of an arginine-containing dentifrice increases the population of arginolytic bacteria in orthodontic patients with white spot lesions. Methods: Forty-seven orthodontic patients, aged 12-21 years with white spot lesions were recruited from the UCSF orthodontic clinic and randomized to 1 of 4 groups: (1) arginine-containing dentifrice (AD)+fluoride rinse (FR) group, (2) AD+FR+antiseptic rinse (AR) group, (3) 1100 ppm fluoride dentifrice (FD)+AR group, or (4) FD+placebo rinse (PR) group. Saliva and plaque samples were collected at baseline, 2 weeks, 2, 4 and 6 months. Mutans streptococci (MS), lactobacilli (LB), and arginolytic bacterial colonies were cultured on MSSB, Rogosa, and BHI agar, respectively. MS and LB were enumerated by observing colony morphologies with a dissecting microscope. Arginolytic bacteria were enumerated using an arginine overlay technique and arginolytic detection reagent. Results: There were no baseline differences in the age, sex, ethnicity and bacterial levels among the study groups. MS levels ranged from non-detectable to 2.1x107 colony-forming units/ml saliva (cfu/ml), LB levels ranged from non-detectable to 1.6x107 cfu/ml, and arginolytic bacterial levels ranged from 0.1% to 76.9% of the total cfu. The ratio of arginolytic bacteria to cariogenic bacteria (MS+LB) ranged from <0.1 to ~1,400,000. No consistent changes in arginolytic bacterial populations were observed within individual participants or among the groups throughout the 6-month study period. Conclusion: Population levels of MS, LB, and arginolytic bacteria in orthodontic participants with white spot lesions were not consistently affected by the use of either fluoride-containing dentifrice or arginine-containing dentifrice.