In the present study, culture conditions of Streptococcus equi was optimized through Box-Behnken experimental design for hyaluronic acid production. About 0.87gL(-1) of hyaluronic acid was produced under the determined conditions and optimal conditions were found as 38.42 degrees C, 24hr and 250rpm. The validity and practicability of this statistical optimization strategy were confirmed relation between predicted and experimental values. The hyaluronic acid obtained under optimal conditions was characterized. The effects of different conditions such as ultraviolet light, temperature and enzymatic degradation on hyaluronic acid produced under optimal conditions were determined. 118 degrees C for 32min of autoclaved HA sample included 63.09 mu g mL(-1) of d-glucuronic acid, which is about two-fold of enzymatic effect. Cytotoxicity of hyaluronic acid on human dermal cells (HUVEC, HaCaT), L929 and THP-1 cells was studied. In vitro effect on pro or anti-inflammatory cytokine release of THP-1 cells was determined. Although it varies depending on the concentration, cytotoxicity of hyaluronic acid is between 5 and 30%. However, it varies depending on the concentration of hyaluronic acid, TNF- release was not much increased compared to control study. Consequently, purification procedure is necessary to develop and it is worth developing the bacterial hyaluronic acid.