Groundwater is the main water source for many areas in Saudi Arabia and the only source of water in some areas. Many local studies have reported that high nitrate concentrations in some wells of groundwater. To provide safe drinking water, the excess amounts of nitrate have to be removed by bio-denitrification process. This study aims to develop a denitrifying biological filter using denitrifying bacteria immobilized on microbial cellulose for the removal of nitrates from water contaminated with nitrate. Denitrifying bacteria that can form biofilter on microbial cellulose were isolated from different regions in Saudi Arabia and were characterized by molecular techniques. They were evaluated for their ability to analyze nitrates and to develop biofilter to remove nitrates from contaminated water. In the results of this project, an optimal microbial cellulose production was achieved by Gluconacetobacter xylinus ATCC 23768 in the lab, which had facilitated the use of biofilter with the immobilized nitrate-reducing bacteria Pseudomonas aeruginosa. The reduction rate of nitrate was reached 1.9mg/L from the starting concentration of 100 mg/L after 18h. Promising results of nitrate removal rate on MC immobilized with Pseudomonas aeruginosa on biofilter at optimized lab conditions of pH, and proper carbon source were achieved. The results suggest that water contaminated with nitrate can be removed by the bio-denitrification process effectively.