Membrane systems for separation of complex protein mixtures: Whey protein purification.

摘要

Although recent studies have demonstrated the potential of using membrane systems for protein separations, most investigations were limited to model binary mixtures, making it difficult to determine the applicability of this technology to real-world feed streams. The overall objective of this thesis was to examine the application of membrane systems for the purification of complex protein mixtures. Experimental studies were performed for the separation of alpha-lactalbumin (alpha-LA) and beta-lactoglobulin (beta-LG), both in a binary mixture and from whey protein isolate, and the purification of an antigen binding fragment of a monoclonal antibody (Fab'2) produced in E. coli. Experimental studies were complemented by theoretical analyses of staged and cascaded membrane systems.;High resolution protein separations were obtained by proper selection of solution pH, ionic strength, membrane, and filtrate flux. Whey protein isolate was separated using two strategies employing different membrane combinations in a 2-stage diafiltration process. In both cases, the purification factor for alpha-LA was greater than 10-fold at 90% yield. The recovery of beta-LG was more challenging due to the presence of multiple impurities. Purified beta-LG was obtained in the filtrate solution at 70% yield and 8-fold purification.;Experimental data were also obtained for the purification of Fab'2 using high performance tangential flow filtration. Product retention and impurity clearance depend on the membrane pore size and surface charge, and the solution conditions, including the presence of specific buffer additives.;Theoretical analyses for multistage and cascaded systems were developed to examine the trade-off between yield and purification factor for the different protein products in the separation of a multi-component mixture. The performance depends on the selectivities for the different components, feed composition, extent of diafiltration, and the ratio of stage volumes. Model predictions are in good agreement with experimental results for the whey protein separation. The behavior of the intermediate component (beta-LG) was particularly interesting, showing a maximum in the yield-purification factor diagram for the staged system and an unusual loop for the cascaded system. The resulting process diagrams provide a framework that can be used to design and optimize multistage membrane processes for the separation of complex protein mixtures.