Infrared Absorption Intensity Analysis as a New Tool for Investigation of Salt Effect on Proteins

The native protein structures in buffer solution are maintained by the electrostatic force as well as the hydrophobic force, salt ions play an important role in maintaining the protein native structures, and their effect on the protein stability has attracted tremendous interests. Infrared spectroscopy has been generally used in molecular structure analysis due to its fingerprint resolution for different species including macromolecules as proteins. However spectral intensities have received much less attention than the vibrational frequencies. Here we report that the spectral intensities of protein amide I band, the finger prints for the protein secondary structures, are very sensitive to the local electric field known as Onsager reaction field caused by salt ions. IR absorbance thermal titrations have been conducted for a series of samples including simple water soluble amino acids, water soluble monomeric protein cytochrome c and dimeric protein DsbC and its single-site mutant G49R. We found that at lower temperature range （10-20℃）, there exists a thermal activated salting-in process, where the IR intensity increases with a rise in the temperature, corresponding to the ions binding of the hydrophobic surface of protein. This process is absent for the amino acids. When further raising the temperature, the IR intensity decreases, this is interpreted as the thermal activated breaking of the ion-protein surface binding. Applying Van＇t Hoff plot to the thermal titration curves, the thermodynamic parameters such as AH and AS for salting-in and ion unbinding processes can be derived for various protein secondary structural components, revealing quantitatively the extent of hydrophobic interaction as well as the strength of the ion-protein binding.

Determination of the thermodynamic dissociation constant of p-nitrophenol in {NaCl+EtOH+H_2O} mixture by ultra-violet spectrophotometry

The thermodynamic ionization constant K° of p-nitrophenol in {10 mass% ethanol-water} mixed solvent is derived from measurements of its absorption spectrum and pH by the two methods: Debye--Huckel extrapolation and polynomial approximation proposed by us in suitable buffer solution at constant ionic strength from 0.1 to 2.0 mol.kg^(-1) at 298.2 ± 0.2 K. The results from both methods are pK° =7.406 and 7.415, respectively. The effect of medium on the ultraviolet spectra of p-nitrophenol has been discussed.

Thermodynamic models based on Pitzer-NRTL and Pitzer-Margules equations for the extraction of tungstic acid with primary amine N1923

The thermodynamic model of the extraction of W with primary amine under near neutral conditions is reported in this paper. The activity coefficients of the nonelectrolytes in organic phase are calculated by the Margules and NRTL equations which are based on previously tested liquid-liquid extraction equilibrium data in combination with mass balances and charge balance formula. The activity coefficients of the electrolytes in aqueous phase are calculated by the Pitzer equation. The thermodynamic model of the extraction of W by a primary amine is constructed from the calculated activity coefficients of electrolytes and nonelectrolytes. The extraction of W using primary amine is also predicated, and the data is compared to the calculated results of the thermodynamic model. It is concluded that Margules equation proves to be suitable and reliable for calculating the activity coefficients of nonelectrolytes in complex organic phase systems.