The effect of chitosan on indirubin production by Polygonum tinctorium cells cultured in shake flasks was investigated. The optimum concentration of chitosan for indirubin production was found to be 200 mg/l, at which the specific indirubin concentration (indirubin concentration per unit weight of cells) was 5.17 mg/g dry cell weight (DCW). The optimum period of elicitation was 5 d in the presence of chitosan. Indirubin production was enhanced by 72% when the cells were cultured in the presence of 200 mg/l of chitosan for 5 d using SH medium supplemented with 2.72 mM calcium chloride and 5 mM indole.