Fluoride has been used as a remineralization agent for many years. In recent years, the possible side effects of fluoride have led researchers to explore new remineralization agents. Theobromine, derived from cocoa beans, is one of the current remineralization agents. The aim of this in vitro study was to assess the remineralization efficacy of different concentrations of theobromine and fluoride in the treatment of initial caries lesions. For this purpose, 5 experimental groups were created with each consisting of 20 prepared bovine enamel samples: 200 mg/L theobromine group (Group T1), 500 mg/L theobromine group (Group T2), 500 ppm fluoride group (Group F1), 1,450 ppm fluoride group (Group F2), and a control group (Group C). All samples were incubated with demineralization solution for 32 hours to create artificial caries lesions. Surface microhardness values (SMH) (n=10 per group) calculated with a Knoop microhardness device and quantities of calcium (Ca), phosphorus (P), and Ca/P ratios (n=10 per group) were analyzed with scanning electron microscopy and energy dispersive X-ray spectrometry (SEM-EDS). Remineralization agents were applied to the groups in an experimental pH cycle for 8 days. Microhardness and mineral deposition measurements of the samples were repeated after treatment. According to the results of our study, the values of SMH, Ca, and P deposition after treatment in all groups except group C showed a statistically significant increase compared with post-demineralization values. There was a statistically significant increase in the F2 and T2 groups in terms of SMH and Ca values after treatment compared to all other groups. The results show that 500 mg/L theobromine increased the surface hardness and Ca and P deposition at a level close to the levels obtained with 1,450 ppm fluoride.