Continuous separation of protein loaded nanoparticles by simulated moving bed chromatography.
Bottom Line: In the case of beta casein where no multimers are present we achieved 89% purity and 90% recovery of loaded nanoparticles in the Raffinate and an extract free of particles (92% purity).Using a tangential flow filtration unit with 5kDa cutoff membrane we proved that the extract can be concentrated for recycling of protein and buffer.The calculated space-time-yield for loaded nanoparticles was 0.25g of loaded nanoparticles per hour and liter of used resin.
Affiliation: Department of Biotechnology, University of Natural Resources and Life Sciences, Vienna (BOKU), Muthgasse 18, 1190 Vienna, Austria.Show MeSH
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Mentions: In comparison to BSA as model protein, beta-casein does not build up any multimers, therefore we expected no such problem as seen for BSA. Fig. 6 shows the time traces of separation of 70 nm silica nanoparticles and beta-casein, and as expected the system is also stable after roughly 3 complete cycles with good separation of protein and particles. This system fulfils the assumption of a binary mixture, resulting in almost pure fractions of proteins and nanoparticles in the extract and Raffinate, respectively. Table 4 shows the corresponding purities and recoveries, we achieved good recovery and good purity for protein and particles around 90% and the calculated productivity was 0.25 g/L/h of purified protein loaded nanoparticle per volume of column resin, which makes this process suitable for industrial production. Chromatography and especially SMB offers great scalability advantages over Ultracentrifugation, being only restricted by the available column and pump sizes and not dependent on particle density or size. The scale up itself would be straight forward as only flow rates and column diameters have to be adjusted to achieve the same residence time in small and large scale.
Affiliation: Department of Biotechnology, University of Natural Resources and Life Sciences, Vienna (BOKU), Muthgasse 18, 1190 Vienna, Austria.