The aim of the present study was to develop nanotechnology-based oral formulations of glipizide to enhance the bioavailability and eliminate the frequent oral administration of the conventional dosage form. Glipizide is an antidiabetic drug with a short biological half-life and limited oral bioavailability. Novel palmitic acid-pluronic F127-palmitic acid (PA-F127) pentablock copolymer-based prolonged release glipizide nanoparticles (GNs) were prepared and screened for in vitro and in vivo studies.GNs were prepared using a novel PA-F127 pentablock copolymer by solvent evaporation technique. The prepared nanoparticles were evaluated for particle size, polydispersity index (PDI), zeta potential, entrapment efficiency, percentage yield, and drug excipient compatibility using fourier transform infrared spectroscopy (FTIR) and differential scanning calorimeter (DSC) analysis, X-ray diffraction, scanning electron microscopy, in vitro drug release studies, stability studies, and in vivo pharmacokinetic studies.The results of FTIR and DSC analysis revealed the absence of drug-excipient interactions. The optimized GN1 had particle size 242.60±4.20 nm, PDI 0.171±0.014, and zeta potential -21.41±0.462 mV. The prepared nanoparticles were spherical and showed semi-amorphous characteristics. The in vitro release studies showed 34.43±4.8% drug was released in the first 8 h and 56.11±4.12% glipizide was released further over 24 h. The GN1 was found to be stable at 5±3°C for up to 3 months. Pharmacokinetic studies showed that the orally administered GN1 was superior with Cmax 2.35-fold, tmax 1.6-fold, area under the curve (AUC0→∞) 3.3-fold, and mean residence time 1.2-fold as compared to pure glipizide (p<0.05).The bioavailability of the newly developed GN1 was successfully increased and the problem of frequent oral administration with the conventional dosage form can be overcome for diabetes treatment.