There are high amounts of hydrogen sulfide in deep periodontal pockets. This volatile sulfur compound may be formed from L-Cysteine, but only low levels of this amino acid can be expected to be present in periodontal pockets. Glutathione, L-gamma-glutamyl-L-cysteinylglycine, is in high concentration in most tissue cells, and this tripeptide may be more readily available as a source of hydrogen sulfide formation in the pockets. The ability of 37 different species of oral bacteria to utilize glutathione in hydrogen sulfide formation was studied. Of these species, only 2 species of Peptostreptococcus and 5 species of Fusobacterium formed high amounts of hydrogen sulfide from glutathione within 24 h. Since the initial rate of hydrogen sulfide formation was more than 5 times higher in Peptostreptococcus micros than in any of the other bacterial species, the kinetics of sulfide formation from glutathione by P micros was further elucidated. The formation of sulfide followed quite closely hyperbolic Michaelis-Menten kinetics. The maximal initial rate of sulfide formation (V(max)) was 163 +/- 2 nmol sulfide per minute per milligram of cellular protein. Half maximal initial rate (K(m)) was obtained at 7.4 +/- 0.8 muM glutathione. The initial rate of sulfide formation from L-Cysteine was much slower and was almost proportional to L-Cysteine concentration. This difference in kinetics of sulfide formation between glutathione and L-cysteine strongly suggested that glutathione was actively transported into the cell, whereas the transport Of L-Cysteine was more or less controlled by diffusion. The sulfide formation from the dipeptide L-cysteinylglycine also followed quite closely hyperbolic Michaelis-Menten kinetics. The maximal initial rate of sulfide formation (V(max)) from L-cysteinylglycine was 75 +/- 2 nmol sulfide per minute per milligram of cellular protein. Half maximal initial rate (K(m)) was obtained at 26 +/- 4 muM L-cysteinylglycine. Hydrogen sulfide may readily be formed from glutathione by P micros in periodontal pockets. This is the first time that experimental data indicates active transport of di- and tripeptides in anaerobic bacteria.