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In wastewater treatment, sulfide or hydrogen sulfide is a major problem in refining process and can cause the pollution in the wastewater plant. In this study, acclimatized activated sludge was used to remove sulfide from the petrochemical wastewater. Five independent factors which affecting the degradation of sulfide were investigated in this paper - wastewater/acclimatized activated sludge (WW/AS) ratio, type of system, temperature, agitation and hydraulic retention time (HRT). Fractional factorial design was used to investigate the effect of independent factors as well as the interaction factors on the sulfide and COD removals (%). Some of the independent factors were shown to have significant effect on sulfide removals (%). The results obtained here showed that the order of significance in sulfide removal (%) is as follows: agitation > WW/AS ratio > type of system > temperature > HRT. Amongst of the operation factors in question, it was found that the interaction factor of temperature*WW/AS ratio had the strongest effect on the sulfide removal (%).