alpha-Amylase from Bacillus amyloliquefociens was purified by the immobilized metal ion affinity adsorbent, beta-CDcl-IDA-Cu2+. The adsorbent was prepared by reacting the cross-linked beta-cyclodextrin (beta-CD) with the ligand, iminodiacetic acid (IDA). The copper ion was further linked to the adsorbent. Poly(ethylene glycol) (PEG) was added to the fermentation broth to improve the adsorption efficiency of the adsorbent toward alpha-amylase. The effort was to provide hydrophobic interactions with the impurities which might interfere with the adsorption of alpha-amylase. It also provided a polymer shielding effect to prevent non-specific interactions. With the addition of PEG, the adsorption efficiency could be increased to 98%. Imidazole containing a phosphate buffer and NaCl was used to elute the bound alpha-amylase. By consecutive adsorption/desorption steps, up to 81% of the alpha-amylase activity could be recovered. Regarding the reutilization of the affinity adsorbents, alpha-amylase could be adsorbed and desorbed six times consecutively without a significant loss of alpha-amylase activity. (C) 2009 Elsevier Ltd. All rights reserved.
