Micellar Solvent Accessibility of Esterified Polyoxyethylene Chains as Crucial Element of Polysorbate Oxidation: A Density Functional Theory, Molecular Dynamics Simulation and Liquid Chromatography/Mass Spectrometry Investigation.

Given that the amphiphilicity of polysorbates represents a key factor in the protection of proteins from particle formation, the loss of this property through degradative processes is a significant concern. Therefore, the present study sought to identify the factors that contribute to the oxidative cleavage of the polysorbate (PS) molecule and to ascertain the preferred sites of degradation. In order to gain insight into the radical susceptibility of the individual polysorbate segments and their accessibility to water, conceptual density functional theory calculations and molecular dynamics simulations were performed. The behavior of monoesters and diesters was examined in both monomer form and within the context of micelles. The theoretical results were corroborated by experimental findings, wherein polysorbate 20 was subjected to 50 ppb Fe²⁺ and 100,000 lx·h of visible light, and subsequently stored at 25 °C/60% r.h. or 40 °C/75% r.h. for a period of 3 months. Molecular dynamics simulations demonstrated that unesterified polyoxyethylene(POE) chains within a polysorbate 20 molecule exhibited the greatest water accessibility, indicating their heightened susceptibility to oxidation. Nevertheless, the oxidative cleavage of esterified polyoxyethylene chains of a polysorbate 20 molecule is highly detrimental to the protective effect on protein particle formation. This occurs presumably at the oxyethylene (OE) units in the vicinity of the sorbitan ring, leaving a nonamphiphilic molecule in the worst case. Consequently, the critical degradation sites were identified, resulting in the formation of degradation products that indicate a loss of amphiphilicity in PS.