| Lactoferrin (LF) and immunoglobulin G (IgG) have become increasingly important due to the diverse range of biological activities. It is of great significance to exploit LF and IgG from bovine colostrum to fuLFill the expanding market demand. The specific synthesis process and unique separation mechanism provide SPEC 70 SLS cation exchange absorbent and MEP Hypercel mixed mode absorbent significant potential for capturing LF and IgG, respectively. In this paper, one-chromatography-step isolation of LF with SPEC 70 SLS resin and efficient purification of IgG with MEP Hypercel resin from bovine colostrum were investigated. Based on the process parameters received in the separation experiments, a novel liquid-solid circulating fluidized beds (LSCFB) ion-exchange system was illustrated for continuous recovery and purification of bioactive proteins from bovine colostrum.Firstly, we investigated the adsorption characteristics of SPEC 70 SLS resin for LF standard solutions, and the results showed that a good dispersion of resins in the bioreactor could reduce the equilibrium adsorption time. Stirring at 50 r/min, the equibrium adsorption time of SPEC 70 SLS resin for LF standard solution and bovine colostrum were 30 and 50 min, respectively. After that, LF and LP could be desorbed within 5 min by NaCl solutions with different concentration. Regrading the binding capacity, the adsorption increased at higher pH near to the pi of Lactoferrin. However, in order to preserve the physical and chemical properties and the flavor of bovine colostrum, the optimal adsorption pH was determined to be 7. The adsorption process could be well described by a Langmuir adsorption isotherm model, with a maximum adsorption capacity of 21.73 mg/g resin and the equilibrium dissociation constant of 0.01463 mg/mL. In batch fractionation of defatted milk with mechanical stirring, the binding capacity of SPEC 70 SLS resin for adsorbing lactoferrin and lactoperoxidase simultaneously was 8.34 and 7.29 mg/g, respectively. In addition, the recovery yield of LF after SPEC 70 SLS resin separation process was 74.50, which was superior to the reported LF separation process.The IgG sample solutions with 80%(w/w) purity, which was obtained from 40%(w/w) ammonium suLFate precipitation of bovine whey, was employed as the substrate for optimizing the spearatoin process of MEP Hypercel chromatography. Dynamic binding studies indicated that MEP Hypercel resin could efficiently capture IgG even at low concentration. Besides, the adsorption was independent to the ionic strength up to 0.5 M NaCl. The optimum binding of MEP Hypercel chromatography for IgG was found to be at pH 7 with a residence time of 15 min, which ensured IgG recovery higher than 90%. Elution process of MEP Hypercel resin chromatography was studied by pH gradient elution method at different ionic strength. It was found that impurities such as low molecular weight proteins could be eluted with pH 5.5 sodium acetate (CH3COONa) buffer solution containing 0.5 M NaCl. As a result, IgG with purity of 93.9% was received in the resulting solution eluted by pH 4.5 CH3COONa buffer and the product recovery of 91.5% was observed in the mixed-mode chromatography process. Moreover, MEP Hypercel resin was regenerated by pH 4 CH3COONa buffer solution.Based on the results of the optimized separation process of SPEC 70 SLS and MEP Hypercel resins, their applications in the LSCFB system were analysed. The density of SPEC 70 SLS resin under wet weight condition was 1065 kg/m3 and the terminal velocity was 0.0085 m/s, which satisfied the requirement of LSCFB referring to related literature. Compared with SPEC 70 SLS resin, MEP Hypercel resin was not suitable to be applied in LSCFB due to its similar density with the bovine whey. At last, in order to achieve the continuous elution process of SPEC 70 SLS resin, a novel LSCFB ion-exchange system with multi-riser was designed. |
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