A new biocatalyst: Penicillin G acylase immobilized in sol-gel micro-particles with magnetic properties.
The present work focuses on the development and basic characterization of a new magnetic biocatalyst, namely penicillin G acylase (PGA), immobilized in sol-gel matrices with magnetic properties, ultimately aimed for application in cephalexin (CEX) synthesis. A mechanically stable carrier, based on porous xerogels silica matrixes starting from tetramethoxysilane (TMOS), was prepared leading to micro-carriers with medium sized particles of 30 microm, as determined by scanning electron microscopy. An immobilization yield of 95-100% and a recovered activity of 50-65% at 37 degrees C, as determined by penicillin G (PG) hydrolysis (pH STAT method), were observed. These results clearly exceed those reported in a previous work on PGA immobilization in sol-gel, where only 10% of activity was recovered. The values of activity were kept constant for 6 months. Immobilized PGA (682 U/g(dry weight)) retained high specific activity throughout ten consecutive runs for PG hydrolysis, suggesting adequate biocatalyst stability. The CEX synthesis was performed at 14 degrees C, using the free and immobilized PGA in aqueous medium. Phenylglycine methyl ester was used as acyl donor at 90 mM and 7-aminodeacetoxycephalosporanic acid was the limiting substrate at 30 mM. The CEX stoichiometric yield after 1-h reaction was close to 68% (23 mM CEX/h) and 65% (19 mM CEX/h), respectively.
ESTM - Escola Superior de Turismo e Tecnologia do Mar, Instituto Politécnico de Leiria, Peniche, Portugal. susanab@estm.ipleiria.pt
Susana M S A
SM
Bernardino
Pedro
P
Fernandes
Luís P
LP
Fonseca
D055162:Biocatalysis; D002506:Cephalexin / Q000378:metabolism; D004574:Electromagnetic Fields; D004795:Enzyme Stability; D004800:Enzymes, Immobilized / Q000737:chemistry / Q000378:metabolism*; D004926:Escherichia coli / Q000201:enzymology; D006863:Hydrogen-Ion Concentration; D006868:Hydrolysis; D008855:Microscopy, Electron, Scanning; D008863:Microspheres; D010399:Penicillin Amidase / Q000737:chemistry / Q000378:metabolism*; D010400:Penicillin G / Q000378:metabolism; D044367:Phase Transition; D012822:Silicon Dioxide / Q000737:chemistry; D013696:Temperature
D004800:Enzymes, Immobilized; D012822:Silicon Dioxide; D010399:Penicillin Amidase; D002506:Cephalexin; D010400:Penicillin G