Synthesis and characterization of pseudo-affinity ligand for penicillin acylase purification.
The aim of this work was to test a chromatographic affinity support containing methacryloyl antipyrine (MAAP) for penicillin acylase (PA) purification by using pure penicillin acylase and crude extract. First, MAAP as a pseudo-specific ligand was synthesized by using methacryloyl chloride and 4-aminoantipyrine. Polymer beads (average size diameter: 40-120 micro m) were prepared by suspension polymerization of ethylene glycol dimethacrylate (EGDMA) and MAAP. This approach for the preparation of adsorbent has several advantages over conventional preparation protocols. An expensive and time consuming step in the preparation of adsorbent is immobilization of a ligand to the adsorption matrix. In this procedure, affinity ligand MAAP acts as comonomer without further modification steps. Poly(EGDMA-MAAP) beads were characterized by FTIR, NMR and screen analysis. Elemental analysis of MAAP for nitrogen was estimated as 89.3 micro mol/g. The prepared adsorbent was then used for the capture of penicillin acylase in batch system. The maximum penicillin acylase adsorption capacity of the poly(EGDMA-MAAP) beads was found to be 82.2 mg/g at pH 5.0. Chromatography with crude feedstock resulted in 23.2-fold purification and 93% recovery with 1.0 M NaOH.
Keçili
Rüstem
R
Department of Chemistry, Anadolu University, Eskisehir, Turkey.
Say
Ridvan
R
Yavuz
Handan
H
D000327:Adsorption; D000345:Affinity Labels / Q000138:chemical synthesis / Q000737:chemistry; D000983:Antipyrine / Q000031:analogs & derivatives / Q000138:chemical synthesis / Q000737:chemistry; D002846:Chromatography, Affinity; D006863:Hydrogen-Ion Concentration; D008024:Ligands; D019906:Nuclear Magnetic Resonance, Biomolecular; D010316:Particle Size; D010399:Penicillin Amidase / Q000302:isolation & purification* / Q000378:metabolism; D010408:Penicillium chrysogenum / Q000201:enzymology; D011109:Polymethacrylic Acids / Q000138:chemical synthesis / Q000737:chemistry; D017550:Spectroscopy, Fourier Transform Infrared
D000345:Affinity Labels; D008024:Ligands; D011109:Polymethacrylic Acids; C497090:poly((ethylene glycol dimethacrylate)-co-(methacrylic acid)); D010399:Penicillin Amidase; D000983:Antipyrine