Evaluation of a monolithic epoxy silica support for penicillin G acylase immobilization

• Published in: Journal of Chromatography A 2004-03, Vol.1031 (1), p.93-100
• Main Authors: Calleri, E, Massolini, G, Lubda, D, Temporini, C, Loiodice, F, Caccialanza, G
• Format: Article
• Language: English
• Subjects: Analysis; Analytical chemistry; Biological and medical sciences; Chemistry; Chiral stationary phases; Chromatographic methods and physical methods associated with chromatography; Chromatography, High Pressure Liquid; Enantiomer separation; Enzymes; Enzymes, Immobilized - chemistry; Epoxy Compounds - chemistry; Exact sciences and technology; General pharmacology; Indicators and Reagents; Magnetic Resonance Spectroscopy; Medical sciences; Microspheres; Monolithic columns; Other chromatographic methods; Particle Size; Penicillin Amidase - chemistry; Penicillin G acylase; Pharmacology. Drug treatments; Protein immobilization; Silicon Dioxide - chemistry; Spectrophotometry, Ultraviolet; Stereoisomerism; Surface Properties
• ISSN: 0021-9673
• DOI: 10.1016/j.chroma.2003.08.076

In this work, a new type of penicillin G acylase (PGA)-based monolithic silica support was developed and evaluated for the chiral separation in HPLC. The preparation procedure consisted of two steps: preparation of an epoxy derivatized monolithic silica column and chemical modification of the epoxide groups with the enzyme chiral selector. The epoxy Silica-Rod column for the immobilization of PGA was prepared with the in situ modification process by using epoxy-silanes and the identification of the species bound to the surface was achieved by solid-state nuclear magnetic resonance. The enzyme was covalently immobilized to the surface of the derivatized monolithic column. The enantioselectivity and the performance of the developed column are discussed and compared to the corresponding experimental data obtained with a PGA-based microparticulate (5 μm) silica column.