The assessment of the relaxation behaviour of frozen aqueous solutions of human serum albumin and polyvinylpyrrolidine

Abstract

In this investigation, the structure and behaviour of frozen solutions of human serum albumin (HSA) alone and in combination with the cryoprotectant polyvinylpyrrolidone (PVP) have been studied using low frequency dielectric analysis and modulated temperature differential scanning calorimetry (MTDSC). Solutions of HSA (1–10 w/v%) and combined solutions (from 1 to 5 w/v% of each ingredient) were prepared and studied thermally and dielectrically over a frequency range of 10 5–10 −2 Hz every 10 °C from +20 to −70 °C. Dielectric data were fitted according to the Dissado–Hill theory and the relaxation times calculated. In addition, a relaxation peak was noted for the frozen HSA systems at a frequency approximately one order of magnitude lower than that seen for the PVP systems, with the PVP dominating the response of the mixed systems. The systems showed Arrhenius behaviour, with, for example, the 5% HSA solution showing an activation energy for the relaxation process of 19.34 kJ/mol. In accordance with previous studies on frozen aqueous solutions of PVP, the results suggest that unfrozen water dominated the dielectric response, with the local environment surrounding the HSA being strongly influenced by the PVP. MTDSC data indicated that the PVP and HSA interact in a complex manner in solution, with a glass transition attributable to PVP being seen only in those systems where PVP was present in sufficient excess. In conclusion, the study has suggested that MTDSC and dielectric spectroscopy may be the useful complementary tools with which the structure and molecular mobility of frozen proteinaceous systems may be studied.