We analyzed the effect of a small heat shock protein HspB6 on the solvent features of aqueous media. To this end, several solvent features in the HspB6 aqueous solutions were quantified using the so-called solvatochromic comparison method. The studied solvent features included solvent dipolarity/polarizability π*, hydrogen-bond donor acidity α, and hydrogen-bond acceptor basicity β. Using the method of partitioning of HspB6 alone, or in the presence of dyes and partitioning of dyes alone, or in the presence of HspB6 in an aqueous two-phase system, we showed that dyes utilized in this study do not bind to HspB6 and hence may be used as the molecular probes of the solvent features of aqueous media in HspB6 solutions. Our analysis revealed that the effects of HspB6 on the water hydrogen-bond donor acidity are displayed at significantly lower concentrations than those of neutral polymers, such as Ficoll-70, dextran-75, dextran-40, and PEG-10,000 or concentrations of several common osmolytes (glucose, sucrose, sorbitol, and xylitol). Our data suggest that the function of the protein chaperone HspB6 in vivo can be at least partially attributed to its effects on the solvent features of aqueous media.