Identification of Polyphenols from Mulberry Leaves and Their Antioxidant and α-Amylase Inhibitory Activity

Objectives: Mulberry leaves are used in traditional medicine as hypoglycaemic and hypotensive agents and their mechanism is related to the content of bioactive ingredients. To evaluate the different biological activities of mulberry leaf extracts, including the content of bioactive ingredients, antioxidant activity and α-amylase inhibitory activity. Methods: The crude extracts of polyphenols were separated and purified into three fractions: Anthocyanins, non-anthocyanin polyphenols and water fractions by SPE (solid phase extraction), and the polyphenols in mulberry leaves were systematically characterized and quantified by HPLC-PDA and HPLC-ESI/MS². The antioxidant activities of the different extracts were determined using three methods: DPPH (1,1-diphenyl-2-picrylhydrazyl), ABTS (2,2'-azino-bis (3-ethylbenzothiazoline-6-sulfonic acid)) and FRAP (ferric reducing antioxidant power). The total phenolic, total anthocyanin and total proanthocyanidin contents of the mulberry leaf extracts were determined by the Folin-Ciocalteu's, pH-differential and colorimetric methods. Results: 6 anthocyanins and 42 non-anthocyanin polyphenols were identified in mulberry leaves, among which, petunidin-3-glucoside and proanthocyanidin derivatives were in high content. The total phenolic, total anthocyanin and total proanthocyanidin contents of the mulberry leaves were 14.09 mg GAE/g DW, 0.17 mg C3G/g DW and 17.24 μg PA2/g DW, respectively. The IC₅₀ (semi-inhibitory concentration) values of the polyphenol crude extracts, anthocyanins, non-anthocyanin polyphenols and water fraction against α-amylase were 8.31, 13.70, 0.25 and 12.00 mg/mL, respectively, which showed that the non-anthocyanin polyphenols had the highest α-amylase inhibitory activity. Conclusion: Mulberry leaves had strong antioxidant activity, and anthocyanins showed the highest antioxidant activity. These results can provide an important reference for the use of Mongolian mulberry leaves as functional foods.