Effect of urea on the linear and nonlinear rheological properties of human serum albumin hydrogels

We report the linear and nonlinear rheological properties of human serum albumin (HSA) hydrogels with and without urea using small (SAOS) and large (LAOS) amplitude oscillatory shear tests. In SAOS tests, pure HSA and HSA-urea hydrogels exhibited a predominantly solid-like behavior (G' ≫ G''), with viscoelastic properties proportional to HSA concentrations. As urea concentration increased, the viscoelastic properties of the hydrogels decreased and the frequency dependence declined, indicating a sparser cross-linking network. Under LAOS flow, both pure HSA and HSA-urea hydrogels exhibited intra- and inter-cycle strain-stiffening, which became more pronounced with increasing urea concentration. The presence of urea delayed the onset of the nonlinear behavior in HSA-urea hydrogels, in proportion to the urea concentration, decreasing the hydrogel strength. The degree of nonlinearity, quantified by the intrinsic nonlinear parameter Q₀ from Fourier-transform rheology, decreased with increasing urea concentration. Additionally, critical strain amplitudes obtained from LAOS tests indicated that the yield strain of the HSA-urea hydrogels increased with urea concentration. The intracycle behavior was analyzed by sequence of physical processes methods. Molecular dynamics simulations were performed to analyze the hydrogels behavior at the atomic level. Significant changes in the hydrogel network were attributed to the efficient insertion of urea into the HSA hydrogen bond network, which forms the cross-linking network. Thus, the hydrogen bonding between urea and HSA, as well as urea’s contribution to HSA denaturation, affects the gelation, linear, and nonlinear viscoelastic properties of the HSA hydrogels.