Aqueous two-phase system (ATPS) has great potential in industrial applications of bio-separations and bio-reactions. However, its large-scale application is limited by recovery difficulty of phase systems. In this paper, a recyclable ATPS was prepared by two pH-responsive copolymers (P ADB4.99 and P MDM7.08 ) and applied for purification of porcine circovirus type 2 Cap protein fermentation broth (PCV2 Cap protein). Phase separation mechanism was studied by using low-field nuclear magnetic resonance (LF-NMR). The results showed that relatively desirable ATPS were formed successfully when two copolymer concentrations were 4%–6% (w/w) with pH 7.5–8.6. The main parameters, such as copolymer concentration, temperature, pH, type and concentration of salts, were investigated. The results demonstrated that the best ATPS consisted of 4% (w/w) P ADB4.99 and 6% (w/w) P MDM7.08 , the optimal partition coefficient (K) and extraction recovery (ER) of PCV2 cap protein were 0.25 and 94.2% in the presence of 50 mM Li 2 SO 4 , 3.53 and 93.4% in the presence of 40 mM KCl, respectively, at 30 °C, pH 8.1. The maximum purity of PCV2 Cap protein was 88.4%. In addition, the two phase-forming copolymers could be recycled by adjusting solution pH to their isoelectric points (pI), with recoveries of over 97.5%.
The vaccine against porcine circovirus type II (PCV2) is mainly composed of virus capsid proteins. An efficient separation method of PCV2 Cap protein has not been developed. Affinity precipitation methods with high selectivity and recoverability based on the thermo-responsive affinity polymer (P NDBN -Ni) were introduced to separate the PCV2 Cap protein. When the temperature was above the polymer LCST (36.1 °C), P NDBN -Ni could be precipitated with a recovery of above 95.00%. The affinity ligand immobilized on the polymer could specifically bind the proteins. The optimal conditions for affinity precipitation were a pH of 6.0 with a ligand density of 56.22 μmol/g polymer at 25.0 °C. The final results were detected by electrophoresis. The interactions between proteins and ligands were determined by the change in ligand state as seen using X-ray photoelectron spectra (XPS).
危险品化学品经营许可证（带存储）