Lipase purification by affinity precipitation with a thermo-responsive polymer immobilized Cibacron Blue F3GA ligand

A thermo-responsive polymer (P-NNB) was synthesized with lower critical solution temperature 27.5A degrees C and over 95% recovery. The adsorption of porcine pancreatic lipase on Cibacron Blue F3GA-conjugated P-NNB (P-NNB-CB) closely followed the bi-Langmuir adsorption isotherm. The maximum adsorption capacity was found at pH 5.0, with a ligand density of 18.4 mu mol/g polymers. The optimized eluent was a 0.01 M phosphate buffer solution at pH 8.0 containing 20% ethylene glycol. Six adsorptiondesorption recycles indicated excellent reusability of the affinity adsorbent. P-NNB-CB was applied to separate porcine pancreatic lipase from its crude material giving a lipase activity recovery of 81.6% with a 16-fold purification factor. Lipase could be purified to single-band purity, according to gel electrophoresis. The purification strategy is therefore feasible and efficient for purifying proteins of interest.