Objectives Because of the less studies in this field in Iraq, this study aimed to use local Streptococcus pyogenes isolates to produce hyluronic acid. Methods The quantitative estimation of hyaluronic acid (HA) produced from eight local S. pyogenes isolates at different pH (6.3, 6.6, 6.9, 7.2, and 7.5) and glucose concentration (4%, 6%, 8%, and 1%) were done using the HA ELISA kit. Results This study showed that the maximum yield of HA was obtained at pH 7.5, and it was found that the differences in pH of the production media of HA enhanced the HA production, but the glucose concentration has no beneficial effect for HA production. Conclusion There were differences in HA production among local isolates at the same pH.