Anti-elastase and anti-hyaluronidase of phenolic substance from Areca catechu as a new anti-ageing agent.

We have previously screened 150 medicinal plants for the inhibition of elastase and found significant inhibitory effects of the extracts of Areca catechu L. on the ageing and inflammation of skin tissues. To isolate and identify the compounds having biological activity, they were further purified by each fraction of solvents, silica gel column chromatography, preparative TLC and reversed-phase HPLC. The peak in HPLC, which coincided with the inhibitory activity against elastase, was identified as a phenolic substance by using various colorimetric methods, UV and IR. IC(50) values of this phenolic substance were 26.9 mug mL(-1) for porcine pancreatic elastase (PPE) and 60.8 mug mL(-1) for human neutrophil elastase (HNE). This phenolic substance showed more potent activity than that of reference compounds, oleanolic acid (76.5 mug mL(-1) for PPE, 219.2 mug mL(-1) for HNE) and ursolic acid (31.0 mug mL(-1) for PPE, 118.6 mug mL(-1) for HNE). According to the Lineweaver-Burk plots, the inhibition against both PPE and HNE by this phenolic substance was competitive inhibition with the substrate. The phenolic substance from A. catechu effectively inhibited hyaluronidase activity (IC(50) : 210 mug mL(-1) ). These results suggest that the phenolic substance purified from A. catechu has an anti-ageing effect by protecting connective tissue proteins.