Clinically Relevant Insulin Degludec and Its Interaction with Polysaccharides: A Biophysical Examination

Abstract

Protein polysaccharide complexes have been widely studied for multiple industrial applications and are popular due to their biocompatibility. Insulin degludec, an analogue of human insulin, exists as di-hexamer in pharmaceutical formulations and has the potential to form long multi-hexamers in physiological environment, which dissociate into monomers to bind with receptors on the cell membrane. This study involved complexation of two negatively charged bio-polymers xanthan and alginate with clinically-relevant insulin degludec (PIC). The polymeric complexations and interactions were investigated using biophysical methods. Intrinsic viscosity [η] and particle size distribution (PSD) of PIC increased significantly with an increase in temperature, contrary to the individual components indicating possible interactions. [η] trend was X > XA > PIC > A > IDeg. PSD trend was X > A > IDeg > XA > PIC. Zeta (ζ)-potential (with general trend of IDeg < A < XA < X ≈ PIC) revealed stable interaction at lower temperature which gradually changed with an increase in temperature. Likewise, sedimentation velocity indicated stable complexation at lower temperature. With an increase in time and temperature, changes in the number of peaks and area under curve were observed for PIC. Conclusively, stable complexation occurred among the three polymers at 4 • C and 18 • C and the complex dissociated at 37 • C. Therefore, the complex has the potential to be used as a drug delivery vehicle.Polymers 2020, 12, 390 2 of 15 Alginates (A) are linear polymers of 1,4-linked β-D-mannuronic acid residues and 1,4-linked α-L-guluronic residues, containing homo-polymeric sequences [7]. Xanthan (X) is composed of D-glucosyl, D-mannosyl, and D-glucuronyl acid residues in a 2:2:1 molar ratio and variable proportions of O-acetyl and pyruvyl residues. Its main chain consists of β-D-glucose units linked at the 1 and 4 positions. Side-chains consist of a tri-saccharide composed of mannose (β-1,4) glucuronic acid and (β-1,2) mannose, attached to alternate glucose residues in the backbone by α-1,3 linkages [8,9].Xanthan gum and alginate complexes (XA) have been studied previously [10] and used for functional foods [11], tissue engineering [3,12,13] and drug delivery. As negatively charged polymers, xanthan [14][15][16][17] and alginate [18][19][20] have been used separately, and in combination with positively charged polymers such as chitosan, as potential vehicles for insulin delivery.Interactions among different types of protein-polysaccharide complexes (PPCs) have been previously investigated using a variety of methods [21,22]. The formation and dissociation of PPCs and their solubility depend on many factors such as surface charge, pH, temperature and ionic strength of the solvent [23]. These factors greatly influence non-covalent interactions such as electrostatic, H-bonding, hydrophobic, and steric interactions, as well as covalent interactions [24]. Conjugation of polysaccharides with therapeutic proteins is a well-established phenomenon [25]. In particul...