Understanding the effect of urea on the phase transition of poly(N-isopropylacrylamide) in aqueous solution by temperature-dependent near-infrared spectroscopy

Regulating the folding state by denaturants is essential for the structure and function of proteins. Poly(N-isopropyl acrylamide) (PNIPAM) is usually regarded as a model for protein denaturation. The effects of urea, as a denaturant, on the aggregations of PNIPAM was studied by temperature-dependent near-infrared (NIR) spectroscopy, and particularly the variation of water structures was analyzed. The NIR spectra of the polymer-urea solutions containing different polymer concentrations were measured at different temperatures. N-way principal component analysis (NPCA) was performed to observe the spectral information. Three principal components (PCs) containing the spectral information of CH groups were obtained, showing three kinds of CH in the system. Obvious dehydration of the three CH groups occurs at 27.5 °C in solution, but the temperature turns to 27 °C for two kinds of the CH and 26.5 °C for the third one, respectively, in the urea-add solution. The effect of urea on the formation of the intramolecular hydrogen bonds that promotes polymer folding is suggested. The spectral information of NH in urea molecule indicates that the direct interaction of urea and polymer facilitates the stability of the polymer globule state. Furthermore, the spectral information of OH shows that the release of the water molecules with three hydrogen bonds (S 3 ), which may connect the NH and CO groups in PNIPAM in solution, leads to the phase transition. When urea is added, urea may reduce the content of the S 3 water to facilitate the release, making the phase transition at a low temperature.