Extraction, in vitro hypoglycaemic activity and active ingredient analysis of polyphenols from walnut green husk

• The hypoglycaemic effect of walnut green husk polyphenol (WGHP) was revealed. • Six bioactive compounds targeting α-glucosidase in walnut green husk were identified by AF-UPLC-MC/MC. • Six bioactive compounds targeting α-glucosidase in WGHP were validated by molecular docking. • Chlorogenic acid was found to be the main polyphenolic compound with hypoglycaemic activity in walnut green husk. A variety of polyphenols in walnut green husk have shown physiological benefits, but their primary hypoglycemic active ingredients remain unclear. We report the extraction of walnut green husk polyphenols (WGHP) and their hypoglycemic effects in vitro . WGHP was extracted using ultrasound-assisted ethanol extraction, with optimization via response surface methodology resulting in 88.0 ± 0.1 % purity after D101 macroporous resin purification. WGHP effectively inhibited α-glucosidase and α-amylase, with IC 50 values of 3.42 μg/mL and 2.56 mg/mL, respectively. It also increased glycogen synthesis in insulin-resistant HepG2 cells and enhanced glucose consumption in 3T3-L1 and insulin-resistant HepG2 cells. Screening six α-glucosidase ligands from WGHP using affinity ultrafiltration and UPLC-MS/MS, combined with molecular docking, identified hydrogen bonds and binding energies between the ligands and the enzyme. These results highlight the hypoglycemic potential of walnut green husk polyphenols and provide a basis for future therapeutic research.