A pH-sensitive nanoparticle (NP) system composed of chitosan and poly(gamma-glutamic acid) was prepared for the oral delivery of insulin. The biodistribution study in a rat model showed that some of the orally administered NPs were retained in the stomach for a long duration, which might lead to the disintegration of NPs and degradation of insulin. To overcome these problems, we freeze-dried NPs and filled them in an enteric-coated capsule. The small angle X-ray scattering (SAXS) profiles indicated that the freeze-drying process did not significantly disrupt the internal structure of NPs; additionally, their pH-sensitivity was preserved and the insulin release was pH-dependent. The results obtained in the native PAGE analysis indicated that the released insulin molecules were neither fragmented nor aggregated. Upon oral administration, the enteric-coated capsule remained intact in the acidic environment of the stomach, but dissolved rapidly in the proximal segment of the small intestine. Consequently, all the NPs loaded in the capsule were brought into the small intestine, thus enhancing the intestinal absorption of insulin and providing a prolonged reduction in blood glucose levels. The relative bioavailability of insulin was found to be approximately 20%. These results suggest that the formulation developed in the study might be employed as a potential approach for the oral delivery of insulin.