Sulfur utilizing autotrophic denitrification synthetic wastewater in up flow packed --bed reactor was evaluated under different concentrations of alkalinity and organic matter. The reactor was filled with granular elemental sulfur particles with diameters of 2.8-5.6 mm and porosity of 40%. The initial culture was prepared from sludge of Shahrak Ghods wastewater treatment plant. First, the stoichiometry ratios, ?SO4/?NO3-N (sulfate produced to nitrogen nitrate removed) and ?ALK. /?NO3-N (alkalinity consumed to nitrogen nitrate removed) were determined 7.05 and 2.92 mg/mg respectively by varying the influent nitrate concentration. Under sufficient alkalinity, complete nitrate removal occurred with 650 mg NO3-N/lit influent nitrate concentration and 0.792 Kg NO3-N/(m3-day) volumetric loading rate. As influent alkalinity decreased to values less than stoichiometry requirements, the nitrate removal efficiency decreased to a final percentage of 60.2% at 810 mg/lit alkalinity as CACO3. Under this condition, organic matter (methanol) was added to feed at concentrations of 247, 494 and 741 mg/lit, equivalent to 20%, 40% and 60% of stoichiometry requirements for heterotrophic denitrification. Results showed that by increasing the methanol concentration, the above stoichiometry ratios decreased to a final ratio of 2.66 and 0. 04 respectively and the overall nitrate efficiency increased up to 100%. Increasing effluent pH and turbidity (as an indirect determinant of effluent biological mass) and almost complete removal of influent methanol was also observed. All these changes imply the co-occurrence simultaneous sulfur based autotrophic and heterotrophic denitrification in bio reactor and show that the addition of controlled concentrations of organic matter to high nitrate wastewater with insufficient alkalinity not only compensates for the lack of alkalinity but also improves the performance of sulfur utilizing autotrophic denitrification