Effects of solution polarity and viscosity on peptide deamidation

Abstract

Deamidation kinetics were measured for a model hexapeptide (L-Val-L-Tyr-L-Pro-L-Asn-Gly-L-Ala, 0.02 mg/mL) in aqueous solutions containing glycerol (0-50% w/w) and poly(vinyl pyrrolidone) (PVP, 0-20% w/w) at 37 degrees C and pH 10 to determine the effects of solution polarity and viscosity on reactivity. The observed pseudo-first order deamidation rate constants, k(obs), decreased markedly when the viscosity increased from 0.7 to 13 cp, but showed no significant change at viscosities >13 cp. Values of k(obs) also increased with increasing dielectric constant and decreasing refractive index. Molecular dynamics simulations indicated that the free energy associated with Asn side-chain motion is insensitive to changes in dielectric constant, suggesting that the observed dielectric constant dependence is instead related primarily to the height of the transition state energy barrier. An empirical model was proposed to describe the effects of the viscosity, refractive index and dielectric constant on k(obs). Analysis of the regression coefficients suggested that both permanent and induced dipoles of the medium affect the deamidation rate constant, but that solution viscosity is relatively unimportant in the range studied.

Tyrosine and peptide reorientational mobility in polymer solutions: Time‐dependent fluorescence anisotropy measurements

“…This result may be significant for stabilization of peptide drugs, for example, where reduction of peptide mobility may inhibit undesirable reactions. 8 We thank Prof. Elizabeth Topp, Department of Pharmaceutical Chemistry, University of Kansas, for the gift of the sample of VYPNGA, and for helpful insights and discussions. We thank Ken Osborn for writing the computer program glovert to convert data from the multichannel analyzer into a format suitable for data fitting in the Globals software package.…”

“…The model peptide VYPNGA has been studied previously as a model for the dependence of peptide degradation on solvent polarity and viscosity. 8,9 Viscosity and temperature-dependent measurements of reorientational motions of VYPNGA were carried out to elucidate the effect of the solvent environment on peptide mobility in heterogeneous samples containing the polymer PVP as a model excipient.…”

Non-Aqueous Systems for Formulation Development—Proteins

“…The rate of deamidation, in solution, decreased with increasing PVP and glycerol concentrations. Overall, the deamidation rate constants decreased with increasing viscosity (0.7-13.0 cp), but showed no effect above 13 cp (Li et al, 2000). The rate constants also increased with increasing dielectric constant and decreasing refractive index.…”

“…The rate constants also increased with increasing dielectric constant and decreasing refractive index. The pseudo first order rate constants were correlated with viscosity, dielectric constant and refractive index in a multi-dimensional free energy model (Li et al, 2000). The data suggested that a) a permanent dipole effect strongly stabilizes the transition state, as indicated by the dielectric constant, b) an induced dipole effect weakly stabilizes the reactant state, as indicated by the refractive index, and c) dynamic or frictional effects were of minimal importance, as indicated by the viscosity (Li et al, 2000).…”

“…The same hexapeptide, Val-Tyr-Pro-Asn-Gly-Ala (pH 10.0), with PVP and glycerol, was utilized to explore the effect of solvent polarity and viscosity on deamidation rate (Li et al, 2000;Li et al, 2002). The rate of deamidation, in solution, decreased with increasing PVP and glycerol concentrations.…”