As a part of a study developing a biological reactive barrier system to treat nitrate-contaminated groundwater, the effects of reactive media composition and co-contaminants on sulfur-oxidizing autotrophic denitrification were investigated. The size of sulfur granules affected the denitrification rates; kinetic constants of 2.883, 2.949, and 0.677 mg-N(1/2)/L(1/2)/day were obtained when the granule sizes were below 2 mm, between 2 and 5 mm, and over 5 mm, respectively. When the volume ratios of sulfur to limestone were 1:1, 2:1, 3:1, and 4:1, kinetic constants of 5.490, 3.903, 4.072, and 2.984 mg-N(1/2)/L(1/2)/day were obtained, respectively. The presence of TCE up to 20 mg/L didn't significantly affect nitrate removal efficiency. At the TCE concentration of 80 mg/L, however, nitrate removal was markedly inhibited. Also, Zn and Cu inhibited the denitrification activity at more than 0.5 mg/L of concentration whereas Cr (VI) did not significantly affect the nitrate removal efficiency at all levels tested.