SEPARATION OF ENANTIOMERS USING ALPHA-CHYMOTRYPSIN-SILICA AS A CHIRAL STATIONARY PHASE

Direct separations of enantiomeric acids and N-substituted amino acids using alpha-chymotrypsin adsorbed or covalently immobilized on silica as the chiral stationary phase are presented. Phosphate buffer was used as the mobile phase. Alpha-Chymotrypsin covalently bound to an aldehyde-activated silica resulted in a stable phase as opposed to alpha-chymotrypsin adsorbed on silica. The covalently bound stationary phase maintained the chiral discrimination ability even after more than 4000 column volumes of mobile phase had passed through the column. An increase in the amount of alpha-chymotrypsin from 110 to 140 mg/g silica was found to affect the retention by a factor of two without significantly influencing the enantioselectivity. The effect of pH, ionic strength and charged modifiers was studied in order to find optimum conditions for chiral separations. The chiral resolution could be optimized by adjusting the pH and by addition of octanesulphonate to the mobile phase.