CHROMATOGRAPHIC BEHAVIORS OF PROTEINS AND AMINO-ACIDS ON A GEL-FILTRATION MATRIX, TSK-GEL TOYOPEARL

The chromatographic behaviors of proteins and amino acids on a gel filtration matrix, TSK-GEL Toyopearl have been examined, and the effects interfering with their elution from Toyopearl gel were analyzed individually. Many proteins are retarded in elution to various extents in 25 mM Tris-HCl buffer at pH 7.5. All the proteins examined of isoelectric points (pI) higher than the pH value of elution buffer (pH 7.5) are retarded in comparison with the elution of the proteins of pI less than 7.5. The retardation can be diminished almost to nothing by the addition of 0.3-0.5 M NaCl, suggesting that the electrostatic interaction works between the proteins and gel matrix. On the other hand, the adsorption of some proteins can be reduced by the addition of 30% ethanol (v/v) to the eluent. These proteins were supposed to be adsorbed to the gel matrix by hydrophobic interaction. Aromatic amino acids such as tryptophan and tyrosine are adsorbed to the gel matrix strongly, but no aliphatic hydrophobic amino acids (e.g., leucine and isoleucine) and no charged amino acids (e.g., lysine, arginine, aspartic acid and glutamic acid) are adsorbed to the matrix at all. By considering the effects interfering with the elution of proteins and amino acids from Toyopearl gel, it is possible to separate them more effectively by the subtle differences in their properties by changing pH, ionic strength, and dielectric constant of the elution buffer.