Nur, Mokhamad (2019) Tragacanth as a novel excipient in oral insulin delivery. PhD thesis, Victoria University.

Abstract

Diabetes mellitus is one of the most grave and lethal non-communicable diseases. Insulin is normally used to medicate diabetes. Due to bioavailability issues, the most regular route of administration is through injection, which may pose compliance problems to treatment. The oral administration thus appears as a suitable alternative, but with several important problems. Low stability of insulin in the gastrointestinal tract and low intestinal permeation are some of the issues. Encapsulation of insulin into polymer-based particles emerges as a plausible strategy. Different encapsulation approaches and polymers have been used in this regard. Polymers with different characteristics from natural or synthetic origin have been assessed to attain this goal, with natural polymers being preferable. Natural polymer such as tragacanth, an anionic polysaccharide gum, can be alternative polymeric carrier for physiologically important peptides and proteins like insulin. Characterisation of tragacanth was explored in the first stage of the study, for providing a foundation for possible applications. Rheological studies colloidal solution of tragacanth at pH 3, 5 or 7 were carried out by means of steady shear and small amplitude oscillatory measurements. From preliminary study, 0.5% tragacanth was selected as optimum colloidal solution and 0.2 mg/ml insulin was chosen as concentration for a model protein. Tragacanth mucoadhesivity was also analysed using an applicable rheological method and compared to chitosan, alginate and PVP. The particle size and zeta potential were measured by a zetasizer. Thermal properties of solutions were obtained using a differential scanning calorimetry. The solution exhibited shearthinning characteristics. The value of the storage modulus (G′) and the loss modulus (G″) increased with an increase in angular frequency (Ω). In all cases, loss modulus values were higher than storage values (G″ > G′) and viscous character was, therefore, dominant. Tragacanth and alginate showed a good mucoadhesion. Tragacanth upon dispersion created particles of a submicron size with z-average diameters (mean) ranging between roughly 431 and 581 nm, with a negative zeta potential (-7.98 to -11.92 mV). These properties were pH dependant resulting in acid gel formation at pH 3.5. Tragacanth has thus a potential to be used as an excipient for peptide/protein delivery. Since tragacanth has a promising result to be used as a carrier in protein/peptide delivery and needs a further application, in the second study, insulin microparticles were prepared by the inclusion of insulin into a tragacanth hydrogel followed by freeze drying. The effect of the pH and concentration relationship involving polyelectrolytes offering individual particle size and zeta potential was assessed by zetasizer and scanning electron microscopy (SEM). Insulin– tragacanth interactions were prepared at varying pH (3.7, 4.3, 4.6, or 6), and concentration (0.1, 0.5, or 1% w/w) to optimize the conditions for optimal delivery of insulin. The pI of insulin can vary from 5.5 to 6.4, based on its origin. The pH 4.3; 4.6 and 6 was selected because these pH is below pI of insulin. At a pH lower than its pI value, insulin will be mainly positively charged. This insulin characteristic could be utilised to facilitate insulin–biopolymer complexes through electrostatic attraction with tragacanth (negatively charged). Individual and smaller particles with z-average diameters approximately 601 ± 19 nm (mean ± S.D.), were acquired at pH 4.6 with 0.5% of tragacanth. The acid gelation test indicated that insulin could be entrapped in the physical hydrogel of tragacanth. DSC thermograms of insulin–tragacanth showed shifts on the same unloaded tragacanth peaks and proposed polyelectrolyte–protein interactions at a pH close to 4.3– 4.6. FTIR spectra of tragacanth–insulin complexes exhibited amide absorption bands featuring in the protein spectra and revealed the creation of a new chemical substance. In the previous stage, tragacanth microparticles seem to have potential functional characteristics for oral insulin delivery by creating a complex with insulin under defined conditions followed by freeze drying. Since freeze-drying is up to 30–50 times more expensive than spray-drying and to make the overall process more industrially applicable, spray drying method has been explored in the third research. A spray-drying process was utilized to create microparticles from insulin/tragacanth GDL acidified solutions. The complexation process was performed at two tragacanth concentrations (0.5; 1%w/w) and several pH values (3.7; 4.3; 4.6; or 6). The SEM analysis indicated that almost spherical or sub-spherical microparticles were created with a diameter of less than 10 μm. The in vitro insulin release of microparticles prepared at a pH 4.3 and 4.6 was substantially minimized in comparison to other pH indicating improved retention of insulin. The selection of complexation pH appears to have an impact on insulin release profile and be an important parameter in protecting against peptic digestion. This finding stem from a possible creation of an insulin/tragacanth complex and hydrogel system. The evaluation of the interaction between insulin and tragacanth at different pH values by ATR-Fourier transform infrared and differential scanning calorimetry analysis verified this hypothesis. This finding suggests that these microparticles may act as a potentially promising device for oral insulin delivery.

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