Determination of Hansen solubility parameters of water-soluble proteins using UV–vis spectrophotometry

Determination of solubility parameters by dissolution tests are difficult for some valuable molecules, such as proteins, where the quantities available are small, UV–vis spectroscopy can determine dissolved concentrations of even small amounts of material, but accurate determination of dissolution is difficult in relatively poor solvents, due to difficulty with constructing a reliable calibration curve. In this work we report a new simple procedure to determine the relative dissolution of proteins in various solvents using UV vis spectroscopy for the determination of Hansen solubility parameters (HSP) of proteins. This method allows qualitative determination of the amounts of BSA dissolved in various solvents. The amounts of BSA dissolved in each solvent, can then be used to rank solvents as good or bad for HSP calculation purpose, which gives more reliable ranking than observation alone in dissolution tests. To be able to evaluate the HSP of any solid material, the solubility of the tested material in a range of solvents needs to be determined. Solvents are then scored as good or bad based on observations and from the known properties of those solvents, the HSP of the solute molecule in question can be calculated. Here, bovine serum albumin (BSA) was used as a model protein, but the procedure reported here can be applied to any soluble protein or other macromolecule with a clear UV/vis adsorption peak. This procedure requires the tested material to be highly soluble in water, therefore eliminating the need for the preparation of many standard solutions of BSA in different solvents. Only a set of standard solutions of protein in water is required. UV–vis spectroscopy was used to analyze the remaining solid resuspended in water after centrifugation of solutions of BSA dissolved in other solvents to separate any undissolved protein. The HSP of BSA obtained via this procedure was compared to values previously obtained for BSA using other methods. A very good agreement between the HSP obtained in this work with the that reported by Houen et al. which used amino acid analysis for the estimation of the solubility of BSA in various organic solvents.