Insulin has a key role in the treatment of diabetes mellitus, which is growing into epidemic proportions worldwide. For the treatment of type I diabetes and many patients with type II diabetes, injection of one or more doses of intermediate- or long-acting insulin is necessary to satisfy the patient’s basal requirement of insulin. This mode of administration has many disadvantages, such as physiological stress, pain, and inconvenience. Therefore there has been great interest in developing an insulin formulation that could provide a controlled release profile of the drug for longer periods of time. For the maximum protection of the drug the integrity of the encapsulating material should be maintained until permeation through the intestine wall. This can be best achieved by the use of polymers, which must be biodegradable and biocompatible. In this study insulin-loaded nanoparticles were prepared by the W/O/W multiple emulsion technique using different blends of PLGA, PLA, PCL and Eudragit® RS100. The AFM results of the insulin-loaded polymeric nanoparticles showed a minimum particle size of 300 nm and maximum particle size of 900 nm. The polymer ratios did not have much effect on the encapsulation efficiency and all formulations had approximately the same encapsulation efficiency values, with the average 81.0% and maximum difference of 5.4%. The in vitro release of insulin from various formulations was evaluated using phosphate-buffered saline (, pH=7.4). For the formulations containing PLGA and PLA, it was observed that with increase in the PLGA percent of the formulation to more than 50%, the amount of insulin released in the first day was significantly increased, due to the low glass transition temperature and the more hydrophilicity of PLGA compared to PLA. The formulation with PLGA/PLA: 45/55 had the minimum burst release, releasing only 16.3% of the encapsulated insulin in the first 24 hours, followed by a smooth and uniform drug release in the next days. For blends containing PCL and Eudragit® RS100, insulin release was increased with the increase of Eudragit® RS100 percentage in the formulation. This may be attributed to the lower molecular weight of Eudragit® RS100 compared to PCL, leading to a less dense encapsulating film, which provides less resistance to diffusion. In general, the amount of insulin released from blends of three and four polymers was more than the optimum PLGA/PLA: 45/55 blend. This may be because of the increase in polymer branches and a less dense encapsulating film with the use of more polymers, and also the reduction of nanoparticles’ sizes, which results in higher diffusion of insulin in polymeric coatings. The in vitro release profiles were modeled with the Higuchi model (average R 2 =0.887) and also the Diffusion model (average R 2 =0.941) and were in better agreement with the