Our previous studies have verified that Galla chinensis (GC), a widely used Chinese medicine, has significant cariostatic effects on bacterial growth, metabolism, and acid production in vitro. Objectives: To evaluate the effects of GC on biofilm and root caries formation in a dynamic culture biofilm model. Methods: A four-organism consortium dynamic root caries model was developed in vitro. Treatments were 4.0 mg/mL GC with 25 mmol/L sucrose, 25 mmol/L sucrose, and distilled water. After bacterial growth in the fermenter stablized, the solution was “pulsed” into the chemostat, with 1/10 of chemostat volume, every 12 hrs for 4 days. HA discs and root blocks were examined 5 times every 24 hrs. To evaluate anticaries efficacy, we counted the numbers of organisms in the biofilm, examined the biofilm image by fluorescence microscopy (FM), examined lesion formation in root tissue by laser confocal scanning microscopy (CLSM), and tracked continuous pH responses to pulses. Results: GC significantly inhibited the adherence of L. rhamnosus and S. mutans on HA discs, but not S. sanguis and Actinomyces naeslundii (p < 0.05, t test). The compositions of L. rhamnosus and S. mutans were different between the experimental and positive control groups (p < 0.05, t test).The extracellular polysaccharide was reduced with the pulsing of GC. An obvious inhibitory effect was observed on acid production. The CLSM image found that the bands of lesions from GC groups were discontinuous and thinner than in the positive control group. Conclusion: GC can reduce the cariogenic response of a sucrose solution in a dynamic root caries model, which can inhibit inoculation, growth, and metabolism of bacteria, and can interfere with the formation of biofilm and inhibit the demineralization of root tissue, all of which relationships appeared over the time-course of the study. Support for this study was provided by a Chinese Natural Science Grant (Grant No. 30430800).