Synthesis And Characterization Of PH-sensitive PEGylated Insulin Nanoparticles

Insulin is the most effective and essential drug for the treatment of insulin-dependentdiabetes mellitus (IDDM). Subcutaneous injection is commonly used in clinical，which bringsthe patients great inconvenience and pain. An oral dosage form of insulin is the preferred formof delivery because of easy administration, patient compliance, as well as, in line with thebody’s normal physiological pathways. However, there are still some challenges to developthe oral dose of insulin due to its short half-life and the rapid enzymatic degradation of insulinin gastrointestinal tract. In this work, to overcome these barriers, single-methoxy polyethyleneglycol butyraldehyde was used to modify insulin to prolong its half-life, and the pH-sensitiveand bio-degradable materials were used to encapsulate the PEGylated insulin to prevent theenzymatic degradation for insulin in the gastrointestinal tract.Free amino sites of insulin were modified by single-methoxy polyethylene glycolbutyraldehyde in optimized pH values. The relative content of mono-PEGylated insulin inmodified products was86%, reached above91%after separated and purified by SP Sepharosecation exchange chromatography. By comparing the subcutaneous injections of PEGylatedinsulin and insulin on diabetic rats, we found that the PEGylated insulin could dramaticallyincrease the half-life of insulin. The blood glucose of diabetic rats was still20%of the initiallevel at10h after injection.Biodegradable polylactic acid (PLA) and pH-sensitive hydroxypropyl methyl cellulosephthalate ester (HP55) were selected to wrap the PEGylated insulin by W/O/W multipleemulsion solvent evaporation method. The effects of volume ratios of the external water phaseto oil phase, mass ratios of HP55to PLA, as well as volume ratios of the oil phase to waterphase on the size and encapsulation efficiency of the PEGylated insulin loaded nanoparticleswere evaluated. The optimized conditions as follows: the external phase concentration is0.1%PVA, the volume ratio of external phase to oil phase is8:1, mass ratio of HP55to PLA is3:2,and volume ratio of oil phase and inner phase is10:1.To obtain dried drug loaded nanoparticles with uniform particle sizes, mannitol was usedas the stabilizer in the freez-drying process to prevent the aggregation of drug loadednanoparticles. And we found that drug loaded nanoparticles with spherical and core-shellstructures, mean diameter of285±7nm, and the polydispersity index of0.233±0.027wereobtained at the optimized mannitol and nanoparticles ratio of1:6. From the results of in vitrodrug release experiments, only30%of mPEGylated insulin was released at pH1.2, while about70%of mPEGylated insulin was released at pH7.4, indicating that the insulin loadedmPEGylated insulin showed obvious pH-sensitivities.