Abstract
Potentilla kleiniana Wight et Arn is widely used as a herbal medicine to treat type 2 diabetes. However, detailed information about its active compounds is lacking. To develop an efficient method for the rapid screening and separation of α-glucosidase inhibitors from Potentilla kleiniana Wight et Arn. Potential α-glucosidase inhibitors from Potentilla kleiniana Wight et Arn were rapidly screened out through ultrafiltration high-performance liquid chromatography mass spectrometry (HPLC-MS), and then followed by a target-guided high-speed counter-current chromatography (HSCCC) separation using two-phase solvent systems composed of n-hexane/ethyl acetate/methanol/water (1:10:1:10, v/v/v/v and 1:10:5:6, v/v/v/v), and adopting increasing flow-rate from 1.5 to 3.0mL/min after 200min. Their structures were identified by ultraviolet (UV), MS, proton nuclear magnetic resonance (1 H-NMR) and carbon-13 (13 C)-NMR, and their α-glucosidase inhibitory activities were assessed by in vitro assay. Five α-glucosidase inhibitors including gallic acid (25.7mg, 98.2%, 1), brevifolincarboxylic acid (9.86mg, 95.3%, 2), ethyl evifolincarboxylate (13.26mg, 97.6%, 3), 3,3'-di-O-methylellagic acid-4'-O-β-d-glucopyranoside (16.26mg, 95.1%, 4), and 3,3'-di-O-methylellagic acid (10.54mg, 96.8%, 5) were successfully purified from 250mg n-butanol extract in a single run. Compounds 1, 2, 4 and 5 exhibited stronger α-glucosidase inhibitory activities[half maximal inhibition concentration (IC50 ) values at 173.41±6.35, 323.46±8.08, 44.63±2.50, and 20.73±2.56μM, respectively] than acarbose (IC50 value at 332.12±5.52μM, reference compound). Notably, compounds 2-5 were reported in the Potentilla kleiniana Wight et Arn for the first time. The results indicated that the proposed method could be applied for the rapid screening and preparative separation of α-glucosidase inhibitors from a complex matrix.
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