Synthesis of magnetic silica based boronate affinity chromatography sorbents and investigation of their chromatographic performance

In this study, β-nicotinamide adenine dinucleotide (β-NAD) was molecularly imprinted onto plain and magnetic silica microspheres in the monodisperse-porous form, by core-shell polymerization method. For this purpose, ethylene glycol dimethacrylate was used as crosslinking agent. For both sorbent species, at alkali pH values, β-NAD showed high binding affinity against phenylboronic boronic acid group and the molecular imprinting was achieved by cyclic boronate ester formation between diol and boronic acid moieties. Following to imprinting, β -NAD was removed from the imprinted microspheres with lauryl sulfate-acetic acid solution. As a control group, bare and magnetic silica microspheres were prepared by the same method without using β-NAD molecule as the template. Molecularly imprinted and non- imprinted silica microspheres were analyzed in terms of their size, morphology, the porous properties through Scanning Electron Microscopy, Transmission Electron Microscopy, Fourier Transform Infrared Spectroscopy and nitrogen adsorption-desorption method. The target molecule isolation behavior was investigated by using borate affinity chromatography with β-NAD-imprinted, bare and magnetic silica microspheres as the sorbents. First, the isolation behavior was investigated by changing the target molecule concentration, sorbent type and concentration in the batch system. The imprinting factors ranging between 2.30-3.38 for bare and magnetic silica microspheres were obtained in batch system. Subsequently, β-NAD-imprinted monodisperse silica microspheres were used as a stationary phase in a microcolumn and β-NAD isolation was carried out by borate affinity micro-chromatography in the continuous system. The results showed that the developed sorbent could be successfully used in the continuous system for the isolation of biomolecules containing diol groups by micro-borate affinity chromatography.