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Andrographis paniculata is an annual herb belongs in the family acanthaceae. It contains the medically secondary metabolite, andrographolide diterpenoid lactone which is used in the treatment of cold cough, disentry etc. Protocol for in vitro production of andrographolide and the method to enhance the andrographolide content by treatment with EMS and SA were carried out in the present study. MS medium supplemented with 2 mg/l NAA and 1 mg/l kinetin were used as the basal medium for callus induction. Andrographolide content in the mutagen treated and control callus were analysed using HPLC method.