Evaluation of the Antidiabetic and Insulin Releasing Effects of A. squamosa, Including Isolation and Characterization of Active Phytochemicals

Abstract

Annona squamosa is generally referred to as a ‘custard apple’. Antidiabetic actions of hot water extract of Annona squamosa (HWAS) leaves together with isolation of active insulinotropic compounds were studied. Insulin release, membrane potential and intracellular Ca2+ were determined using BRIN-BD11 cells and isolated mouse islets. 3T3L1 adipocytes and in vitro models were used to determine cellular glucose uptake, insulin action, starch digestion, glucose diffusion, DPP-IV activity and glycation. Glucose intolerant high-fat fed rats were used for in vivo studies. Active compounds were isolated and characterized by HPLC, LCMS and NMR. HWAS stimulated insulin release from clonal β-cells and mouse islets. Using fluorescent indicator dyes and modulators of insulin secretion, effects could be attributed to depolarization of β-cells and influx of Ca2+. Secretion was stimulated by isobutylmethylxanthine (IBMX), tolbutamide or 30 mM KCl, indicating additional non-KATP dependent pathways. Extract stimulated cellular glucose uptake and insulin action and inhibited starch digestion, protein glycation, DPP-IV enzyme activity and glucose diffusion. Oral HWAS improved glucose tolerance and plasma insulin in high-fat fed obese rats. Treatment for 9 days with HWAS (250 mg/5 mL/kg), partially normalised energy intake, body weight, pancreatic insulin content, and both islet size and beta cell mass. This was associated with improved oral glucose tolerance, increased plasma insulin and inhibition of plasma DPP-IV activity. Isolated insulinotropic compounds, including rutin (C27H30O16), recapitulated the positive actions of HWAS on beta cells and in vivo glucose tolerance and plasma insulin responses. Annona squamosa is attractive as a dietary adjunct in treatment of T2DM and as a source of potential antidiabetic agents including rutin.