European Journal of Pharmaceutics and Biopharmaceutics
Research paperFormulation and characterization of an oily-based system for oral delivery of insulin
Introduction
Insulin is an important therapeutic protein due to its role in the treatment of diabetes, which is growing into epidemic proportions in many countries [1]. Despite the advances in development of injectable insulin analogues, the goal of optimal glycemic control has remained elusive. Substantial progress has been made recently in non-invasive methods for insulin administration such as pulmonary, nasal, rectal and oral in order to replace parenteral therapy [2]. The oral route is recognized as the natural and the safest route for drug administration. Generally, oral administration can improve disease management, enhance patient compliance and reduce of long-term complications of diabetes [3]. However, the oral route continues to be a challenge to deliver proteins and various barriers must be overcome to obtain an adequate bioavailability. These barriers are the permeability across gastrointestinal tract (GIT), the enzymatic barriers and protein stability in GIT environment [4].
Different formulation approaches have been investigated to overcome the GIT barriers for the delivery of insulin via oral route such as the use of liposomes, microemulsion, microspheres and nanoparticles. Nanosize carriers have a large specific surface area, and their protection power against gastrointestinal environment is mostly dependent on the degree of protein encapsulation within these nanostuctures. Such nanoparticles are thought to be the most promising solution for oral delivery of peptides and proteins [5]. However, it is well known that proteins and peptides undergo a loss of activity following encapsulation during their processing [6]. A loss of insulin activity was observed following encapsulation in polylactic acid and poly (lactide-co-glycolide) microspheres [7], [8]. In addition, the use of some components used in polymerization during nanoparticles preparation may affect biocompatibility of the final formulation [9]. Therefore, researchers recently focused on the use of naturally occurring polymers especially polysaccharide such as chitosan for the preparation of carriers in an aqueous environment avoiding heating, organic solvents and severe mechanical stress [10], [11]. Previously, insulin-loaded chitosan nanoparticles were prepared by ionotropic gelation of chitosan with tripolyphosphate anions [12] or by polyelectrolyte complexation with polyanionic polymers such as poly (γ-glutamic acid) and dextran [13], [14]. In those studies the in vitro release tests showed a very rapid initial burst effect. The released insulin was likely to be degraded by proteolysis in the GIT resulting in a low pharmacological availability [12].
Recently, polyelectrolyte complexation method was utilized to prepare insulin–trimethyl chitosan nanoparticles for intranasal delivery of insulin [15]. Interaction of chitosan and polyanions leads to spontaneous formation of nanoparticles. However, these polyelectrolyte complexes (PECs) are easily dissociated in acidic medium, because both insulin and chitosan are soluble at lower pH such as that of the stomach. Sadeghi et al. [16] observed a prompt release of insulin in 0.01 N HCl from nanoparticles made by PEC method, where the released insulin in the first few minutes was about 70% of the incorporated quantity, while the rest was released after 30 min. In order to protect proteins from the unfavorable environment of the GIT, some researchers proposed the use of oily vehicles [5], where the most promising oily-based preparations are water in oil (w/o) microemulsions. These microemulsions have been proposed to enhance the oral bioavailability of peptides by protecting them from the enzymatic breakdown in the GIT, and increasing their permeability through the intestinal wall [17]. In spite of this when insulin was incorporated into a w/o microemulsion high doses were used (200 IU/kg) to elicit a pharmacological response [18].
In order to overcome the aforementioned shortcomings a new formulation strategy was adopted, where a polyelectrolyte complex of insulin–chitosan was dispersed in a microemulsion. The resultant w/o nanosized system was characterized and tested on diabetic rats.
Section snippets
Materials
Recombinant human (rh) insulin powder was purchased from Biocon, Bangalore, India. Low molecular weight chitosan was prepared by acidic depolymerization of high molecular weight chitosan HCl (MWT ∼ 250 kDa and DDA 93%, Xiamen Xing, Shanghai, China). Oleic acid was obtained from Merck, Germany. Plurol oleique® (polyglyceryl-6-dioleate) and Labrasol® (PEG 8 caprylic/capric glycerides) were purchased from Gattefosse S.A., Priest, France. Streptozotocin, and pepsin were obtained from Sigma–Aldrich,
Depolymerization, molecular weight and degree of deacetylation determination
Depolymerization of high molecular weight chitosan by hydrochloric acid was carried out according to previously reported methods [21], [22], [23] This yielded fractions of low molecular weight chitosans. Identification of theses fractions were carried out by FTIR which showed similar distinct peaks as appeared in the parent compound. This is evidenced that depolymerization process did not change the basic chemical structure.
In this work, a molecular weight of 13 kDa with a degree of
Conclusions
The present investigation explored the possibility of formulating an oral insulin delivery system by combining the advantages of nanoencapsulation and the use of an oily vehicle. The preparation was found to have a unimodal particle size distribution with a mean diameter of 108 ± 9 nm. Insulin was protected from gastric enzymes by incorporation into lipid-based formulation. The results of RP-HPLC and ELISA indicated that insulin was able to withstand the preparation procedure. Insulin in the
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