Experimental Method to Distinguish between a Solution and a Suspension

Abstract Dispersion of objects in a fluid phase can be classified as solutions (Gibbs free energy of mixing, ΔGmix < 0) or suspensions (ΔGmix > 0) depending on their thermodynamic stability. Small objects tend to form solutions, larger ones suspensions, e.g., molecules versus micrometer-sized colloids. Proteins and nanomaterials fall between these two size regimes. The long-standing issue of whether proteins and nanoparticles are dissolved or suspended remains an important research question. Here, a simple, versatile, and experimentally robust method, based on sedimentation equilibrium analytical ultracentrifugation (SE-AUC), which can determine whether proteins, nanoparticles, or polymers form solutions or suspensions, is presented. SE-AUC determines the osmotic pressure profile for a dispersion. Such a profile for solutions (equilibrium one-phase systems) is independent of the initial and the operating conditions. The opposite is true for suspensions that are nonequilibrium two-phase systems. This study proves that bovine serum albumin and lysozyme form solutions while ferritin and apoferritin form suspensions.