Background: There are many health hazards associated with diabetes mellitus, characterized by high blood glucose levels caused by defects in insulin secretion or insulin action, or both. Among the most life-threatening diseases, cancer presents many health hazards both in developed and developing countries, characterized by irregular cell proliferation. The prevalence of diabetes and cancer are escalating day by day and this shows the urgently require for the management of diabetes and cancer. Methods: In this study was aimed to evaluate the antidiabetic and anticancer potential of Siddha polyherbal formulation Tulasi oil (SPFTO) by glycosylation of hemoglobin assay, glucose uptake by yeast cells, alpha amylase inhibition and MTT assay method, respectively. Results: The inhibition rate in all concentrations of Tulasi oil exhibited higher inhibition of glycosylation, glucose uptake in yeast cells assay and α amylase as compared with the standard drug. The cytotoxic activity for MCF7 cell line was 65.75% at 50 μg/mL concentration by MTT assay method. Conclusion: The results of this study demonstrate that the Siddha polyherbal formulation Tulasi oil has significant antidiabetic and antiproliferative activities.