A new series of Pyrazolo[3,4-d]pyrimidine containing amide derivatives (8 a-l) were designed, synthesized, and evaluated for their in vitro α-amylase inhibitory activity. The IC50 values of the target compounds ranged from 1.60±0.48 to 2.04±1.20 μM as compared to the standard acarbose 1.73±0.05 μM. All the Pyrazolo[3,4-d]pyrimidine amide derivatives displayed good inhibitory activities, while seven analogs (8 d, 8 f, 8 g, 8 h, 8 i, 8 j and 8 k) exhibited more or less equipotent activity with IC50 values 1.77±2.84, 1.65±0.45, 1.66±2.24, 1.73±0.37, 1.60±0.48, 1.75±0.36 and 1.64±0.03 μM respectively. Further, the most potent α-amylase inhibitors 8 d and 8 k were also screened for their in vivo antidiabetic activity against alloxan induced diabetic rat model at the dose of 25 and 50 mg/kg. Oral administration of these tested compounds significantly reduced the fasting blood glucose levels in dose dependent manner. The hypoglycemic effects of these compounds were more evident at 3 h and 5 h after administration of tested compounds which was similar to the effect displayed by the positive control. In addition, the binding energies calculated from the docking studies with the α-amylase enzyme (PDB ID: 1HNY) and biological activities indicate that the compounds containing nitro moiety on the phenyl group contributed significantly towards the antidiabetic activity. © 2019 Wiley-VCH Verlag GmbH & Co. KGaA, Weinheim