Evaluation of a hydrazide-linked alpha1-acid glycoprotein chiral stationary phase: separation of R- and S-propranolol.

The binding and chiral separation of R- and S-propranolol was investigated on a new type of alpha1-acid glycoprotein (AGP) column. This column was prepared through the controlled and mild oxidation of AGP, followed by the immobilization of this protein to hydrazide-activated silica. The effects of temperature, pH, ionic strength, and organic modifiers on the retention and separation of R- and S-propranolol were investigated on this column. Both the association equilibrium constants and number of binding sites for R/S-propranolol on the AGP column were found to increase with temperature and affect the measured retention factors for these compounds. Regarding the other factors, a change in the organic modifier concentration was found to give the largest change in retention and separation. It was found through these studies that both coulombic and hydrophobic interactions played important roles in determining the retention of R- and S-propranolol on the AGP column. The efficiency and separation impedance of this system were also considered. Under the final optimum conditions identified in this study, it was possible to separate R- and S-propranolol with a resolution of greater than 1.38 in less than 5 min on a 4.1 mm I.D. x 5 cm column.