The present study investigated the use of chemical extractions for the determination of acid volatile sulfides (AVS) and chromium reducible sulfides (CRS) in sulfidic mine tailings from Au and Cu-Zn mines. The main objectives of the study were to understand the factors affecting AVS and CRS extraction in mine tailings and to use the AVS and CRS data as indicator of biotic reactions, including microbial sulfate-reduction. Tailings from one Cu-Zn mine and two Au mines located in the Timmins area, Ontario, Canada, were extracted for AVS and CRS, along with pure metal sulfides. Tailings were also characterized by XRD and the relative abundance of pyrite and pyrrhotite in the samples was determined. Our results showed that AVS in mine tailings were generally not extractable at cold temperature, but they could be extracted from most tailings samples at ∼ 80 °C. AVS extraction of pure metal sulfides showed that pure pyrrhotite was partially soluble during cold and hot extractions and that it significantly contributed to the overall AVS fraction of the tailings. Such findings complicate the use of AVS as bio-indicators of microbial activity because a large fraction of the pyrrhotite present in the tailings originated from the ore deposit. The CRS fractions were found to be unaffected by the presence of pyrrhotite and truly represented the pyrite content of the tailings. The use of AVS and CRS data from mine tailings can be helpful in assessing the distribution of metal sulfides but attention must be paid to their true significance.