Antiangiogenic Activity and Bioassay-guided Isolation of Aqueous Extract of Orthosiphon stamineus

Orthosiphon stamineus Benth. (Family: Lamiaceae), a famous traditional medicinal plant in tropical and sub-tropical region, is being used for treating various ailments including inflammatory conditions. Since anti-inflammation and antiangiogenesis involve cyclooxygenase-II pathway, the present study is aimed to carry out antiangiogenic studies and bioassay-guided isolation on aqueous extract of the plant. After antiangiogenic activity studies on the extract, fractions were prepared by partitioning the extract with hexane, chloroform and ethyl acetate, which were then evaluated for antiangiogenic activity. The most active, hexane fraction (HF) upon activity-guided isolation gave three compounds, 1 (betulinic acid), 2 (oleanolic acid) and 3 (ursolic acid). The total contents of these three compounds were determined in the extract and its fractions using HPLC. The extract and HF exhibited 20% and 80% antiangiogenic activity, respectively, whereas, chloroform and ethyl acetate fractions were inactive. The three isolated compounds exhibited 100% antiangiogenic activity. Median effective concentrations (EC50s) of HF, 1, 2 and 3 were found to be 84, 32, 34 and 38 mu g/mL, respectively. The results of this study indicate that hexane fraction and the isolated compounds have promising antiangiogenic activity. Additionally, the isolated compounds may be sued as analytical markers to standardize extracts and formulations prepared from O. stamineus.