Reversal of elution order of N‐(2,4‐dinitrophenyl)‐proline and N‐(2,4‐dinitrophenyl)‐serine in HPLC by BSA chiral stationary phase

Abstract

N-(2,4-dinitrophenyl)-proline and N-(2,4-dinitrophenyl)-serine were enantiomerically resolved on the BSA chiral stationary phase by HPLC in reversed-phase mode. Effects of chromatographic conditions on enantioseparation and elution order have been investigated in detail. For these two samples, reversal of enantiomer elution order was observed by changing buffer pH, the content of acetonitrile, or alcohol modifiers in mobile phase, which is firstly reported in the BSA chiral stationary phase studies. More interestingly, combined effect between buffer pH and the content of acetonitrile was also observed. In addition, coelution range of enantiomers varied along with the content of acetonitrile in mobile phase.

Development of Bovine Serum Albumin-Bonded Silica as a Chiral Stationary Phase and Its Application in Quantitative Direct Enantiomeric Resolution

“…Verification of the Chemical Natures of (1)−( 5). The presence of Br in (1), the absence of Br in (2), and the absence of free BSA and the covalent attachment of BSA to silica in (5) were verified by different chemical tests at each step (as described in the Experimental Section). The results of elemental analysis (C, H, N) of the synthesis products (2)− (4) showed a corresponding increase in comparison with pure silica gel, which verified that alkyl and amino groups had been successfully bonded onto the silica surface.…”

“…The results of elemental analysis (C, H, N) of the synthesis products (2)− (4) showed a corresponding increase in comparison with pure silica gel, which verified that alkyl and amino groups had been successfully bonded onto the silica surface. The presence of sulfur in (5) inferred that BSA molecules were successfully bonded to the silica surface. FTIR spectra of the BSA−silica CSP showed ν OH stretching for silanols 11,12 in the region of 3200−3800 cm −1 .…”

“…The presence of N was observed in (4), and the N content was found to have increased significantly in (5) as a result of the binding of BSA. Small sulfur content was also observed in (5). The characteristics of the CSP materials obtained using three different batches/ samples of BSA were identical.…”

“…The following tests were performed on the new BSA−silica CSP (5) obtained from three different batches of BSA:…”

“…Furthermore, reversal of the enantiomer elution order was observed in BSA CSP studies through changing the mobile phase pH, the content of acetonitrile, or alcohol modifiers. 5 None of these is suitable for large-scale enantioseparation.…”