Separation of enantiomers on chiral stationary phase based on cellulase: Effect of preparation method and silica particle diameters on chiral recognition ability

Abstract

Cellulase (Cel) was immobilized onto aminopropyl-silica gels via its amino and carboxy groups, respectively, using N,N′-disuccinimidyl carbonate, and 1-ethyl-3-(3′-dimethylaminopropyl)carbodimide and N-hydroxysulfosuccinimide. They were termed N-Cel and C-Cel, respectively. Despite their smaller retention factors on a C-Cel column, the enantioseparation factors and resolution of β-blockers, propranolol, alprenolol, oxprenolol and pindolol, were similar with N- and C-Cel columns. In addition, C-Cel was prepared using aminopropyl-silica gels, whose nominal particle diameters were 5 and 3, and 2.1μm, respectively. A C-Cel column prepared with 2.1-μm aminopropyl-silica gels gave the highest enantioselectivity and column efficiency among three C-Cel columns. Furthermore, the influence of N,N-dimethyl-n-octylamine (DMOA) or cellobiose concentrations on the retentivity and enantioselectivity for β-blockers on a C-Cel column was investigated. The results indicate that single-site competition of β-blockers with DMOA or cellobiose on the catalytic binding site of Cel and the further bindings at the secondary site in a non-competitive fashion could occur. Furthermore, the enantioselective bindings of β-blockers could occur at the catalytic biding cite of Cel and at the secondary binding site.