Background: White spot lesion is considered as one of the main problems in the orthodontic treatment. Brackets used in fixed orthodontic treatment create an environment that provides enamel demineralization. Objectives: The objective of the current study was to perform an in vitro study to compare different applications of fluoride supplements on enamel demineralization adjacent to orthodontic brackets and finally to understand the best supplement to recommend the patients. Methods: One hundred and twenty extracted caries-free human premolar teeth were randomly assigned into six groups: group 1: Control group, group 2: Fluoride toothpaste, group 3: Fluoride toothpaste/mouth rinse, group 4: Fluoride toothpaste/vanish, group 5: Fluoride toothpaste/gel and group 6: Fluoride toothpaste/foam. After bonding the brackets to the teeth, the fluoride supplements were applied based on each group above, except the control group. Then all the specimens were cycled for 30 days in demineralization solution for 8 hours a day, rinsed, placed in artificial saliva for 4 hours a day, brushed (except the control group), and put back to artificial saliva for 12 hours. DIAGNOdent laser fluorescence was used to quantify the demineralization changes. Results: Significant differences existed between all fluoride-containing groups and control group. Analyses of the results showed a significant difference between control group and the rest 5 treatment groups (P < 0.001). Other significant differences were between groups 2/5, 3/5, 2/4 and 5/6 (P < 0.05). There was no significant difference among the other groups (P > 0.05). Conclusions: According to the results, all fluoride supplements could be used during orthodontic treatment to decrease the enamel demineralization. It has been illustrated that fluoride-containing toothpaste and mouth rinse is better than no fluoride treatment but is not effective as well as fluoride gel and varnish.