Andrographolide is the main bioactive component of the medicinal plant, Andrographis paniculata. Callus and suspension cultures of Andrographis paniculata are known to produce only paniculides (sesquiterpene lactones) and not andrographolides (diterpene lactone) that are produced by the intact plant. In this study, Indole-3 acetic acid (IAA), Napthalene-acetic-acid (NAA) and Gibberellic acid (GA) were used at different concentrations such as 25, 50, 75, 100mg/l to find their effect on andrographolide content in leaves of A. paniculata. Treatment of detached leaves with growth regulators showed increase in andrographolide content and maximum enhancement was observed with NAA at 100 mg/l. Spectrophotometric and HPLC analysis proved andrographolide induction in callus by treatment with NAA. Present study thus indicates that andrographolide production can be induced in the callus of A. paniculata by treatment with plant growth hormones.