In this study, a bio-electro reactor in which H2 gas produced by electrolysis of water and organic matter were used for the reduction of nitrate was investigated. A mathematical model describing the utilization of both substrates in a biofilm attached on the surface of the cathode was developed. The following three phases were discussed;(1) The relation between distribution of substance concentration and denitrification rate inside of biofilm by using only electrolysis of water as electron donor supply.(2) The relation between distribution of substance concentration and denitrification rate inside of biofilm by using only acetate as electron donor supply.(3) The relation between distribution of substance concentration and denitrification rate inside of biofilm by using both of electrolysis of water and acetate as electron donor supply. The calculation results showed that the denitrification rate was significantly increased when H2 gas and organic matter were used simultaneously. The model prediction and experimental results were in fairly good agreements.