4.15: Dialysis
Dialysis is a diffusion-based purification process that separates analyte molecules from a complex matrix. This is accomplished by allowing molecules in the solution to pass through a semipermeable membrane into a liquid on the other side. The membrane is usually made of cellulose acetate or cellulose nitrate, and the second liquid must be miscible with the solution. Ions (e.g., chloride or sodium) or organic molecules (e.g., glucose) can pass through the membrane pores, which generally have diameters ranging from 1 to 5 nm. Larger molecules (e.g., proteins, hormones, enzymes) with diameters significantly greater than the pore diameter are retained in the original solution.
Dialysis is an inexpensive technique that is commonly used to purify proteins, hormones, and enzymes. During the procedure, a sample is placed in a dialysis bag, and both ends are sealed. The sealed bag is then immersed into a beaker filled with a solution called the dialysate, which has a composition different from the original solution. As dialysis proceeds, smaller molecules freely diffuse across the membrane, migrating from areas of high concentration to those of low concentration, until equilibrium is reached. Larger molecules that cannot enter the pores are left behind. Dialysis is complete once no net movement of the smaller molecules can occur.
Dialysis is a slow process, so it is often performed overnight or over several days. The driving force for diffusion is provided by the concentration gradient created by differences in analyte concentration across the dialysis membrane. The rate of molecular diffusion can be increased by raising the temperature and decreasing the thickness of the membrane. Because dialysis relies on equilibrium, it often needs to be repeated a few times to decrease the concentration of unwanted small molecules to an acceptable level.
