Improving Oral Bioavailability Of Insulin By Utilizing Nanoemulsions Coated With Alginate-Chitosan

Insulin is the most important regulator of metabolism as well as an anabolic hormone, which is the main effective drug for type I diabetes. As a hydrophilic peptide or protein, the properties of low penetration ability through membrane and high sensitivity to heat and enzyme, causing easily destroyed in gastrointestinal tract limit insulin application by general oral administration. The research of insulin oral formulas is a hot spot recently. W/O microemulsion has been closely fellow these years as a drug delivery system for protein and peptides for it improvements of oral bioavailability. However, w/o microemulsion is easily transformed into o/w emulsion and it may irritate the gastrointestinal tract for large amount surfactants or co-surfactants usage. This paper aims to improve the insulin oral bioavailability using w/o/w nanoemulsion coated by alginate-chitosan, the natural bioadhesive polymer, which can protect the inner nanoemulsion from breakage in gastrointestinal tract by gastric acid or enzymes and prolong the residence time.The first part aims to the preparations of nanoemulsion and then optimization of formula and characterization. Multi-emulsion was prepared by two step method, constituting of Labrafac CC, phospholipid, Span80and Cremorphor EL. After homogenization of multi-emulsion,0.067%alginate sodium,0.1%calcium chloride and0.06%chitosan were added to coat the nanoemulsion by crosslinking. The coated and uncoated nanoemulsions were characterized by size, zeta potential and transmission electron microscopy (TEM). The conformation of insulin was determined by circular dichroism (CD) spectroscopy. The in vitro and in vivo properties were further studied respectively. The size of coated nanoemulsion was488nm and the insulin entrapment ratio was47.3%. CD spectroscopy proved no change of insulin conformation in the preparation process. In vitro stability indicated outstanding protection in simulated gastric juices.The second part is to investigate the in vivo hypoglycemic effects by animal experiments. Wonderful hypoglycemic effects were observed in both normal and diabetic rats. The relative pharmacological bioavailability of25IU/Kg and50IU/Kg dose of coated nanoemulsion were8.42%and5.72%in normal rats,8.19%and7.84%in diabetic rats respectively, higher than uncoated nanoemulsion with relative pharmacological bioavailability which showed no statistical difference compared to blank control.The last part is to explore the basic mechanism of nanoemulsion coated by alginate-chitosan. In intestinal bio-adhesion experiments, the confocal laser scanning microscope photographs show that the absorbed amounts of alginate-chitosan nanoemulsion to intestinal villi is clearly more than general emulsion in jejunum and ileum section. Through cell transportation experiments I find that the nanoemulsion coated by alginate-chitosan didn’t show high values, even more less. The result indicates that the bio-adhesion of materials and the protection of coated shell may play great role in improving insulin bioavailability.