This study was conducted to examine the effect of dietary n-3/n-6 fatty acid (FA) ratio on in vitro dry matter digestibility (IVDMD), fermentation indices and FA profile. Rice bran was mixed with oil sources (cotton seed oil and linseed oil) to make the diets at 0.02, 0.29 and 0.61 of dietary n-3/n-6 FA ratio. These diets (0.5g) were placed into the incubation bottles with 40 ml of anaerobic culture medium, which contained rumen fluid and Van Soest medium at 1:2 ratio. Five replicates of each diet and two blanks were incubated at 39℃ for 48 hours. After incubation, the incubated contents were centrifuged. The residues were freeze-dried for DMD and FA analyses. The supernatant was used for pH, NH3-N and volatile fatty acid analyses. The concentrations of lactate (p<0.001) and iso-valerate (p<0.001) decreased linearly with increasing dietary n-3/n-6 FA ratio, but acetate concentration (p=0.056) and the ratio of acetate to propionate (p=0.005) was increased linearly. The concentrations of n-3, n-6 FA and the ratio of n-3/n-6 FA in residues increased (p<0.001) linearly with increasing dietary n-3/n-6 FA ratio, but C18:1n-9 FA concentration was decreased (p<0.001) linearly. With these results, it could affect fermentation characteristics and FA profile of rumen content by dietary n-3/n-6 FA ratio.