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The ability to process high-concentration monoclonal antibody solutions (> 10 g/L) through small-pore membranes typically used for virus removal can improve current antibody purification processes by eliminating the need for feed stream dilution, and by reducing filter area, cycle-time, and costs. In this work, we present the screening of virus filters of varying configurations and materials of construction using MAb solutions with a concentration range of 4-20 g/L. For our MAbs of interest-two different humanized IgG1s-flux decay was not observed up to a filter loading of 200 L/m(2) with a regenerated cellulose hollow fiber virus removal filter. In contrast, PVDF and PES flat sheet disc membranes were plugged by solutions of these same MAbs with concentrations >4 g/L well before 50 L/m(2). These results were obtained with purified feed streams containing <2% aggregates, as measured by size exclusion chromatography, where the majority of the aggregate likely was composed of dimers. Differences in filtration flux performance between the two MAbs under similar operating conditions indicate the sensitivity of the system to small differences in protein structure, presumably due to the impact of these differences on nonspecific interactions between the protein and the membrane; these differences cannot be anticipated based on protein pI alone. Virus clearance data with two model viruses (XMuLV and MMV) confirm the ability of hollow fiber membranes with 19 +/- 2 nm pore size to achieve at least 3-4 LRV, independent of MAb concentration, over the range examined.

For rapid development of initial solvent extraction processes, knowledge of the solubility and partition behavior of surfactants and solubility enhancers is required. Unfortunately, experimental solubility data for many common surfactants and solubility enhancers in aqueous and organic solvents have not been reported. There are also few references to the partitioning behavior of these additives between water and common extraction solvents. In this paper, the solubility and partition coefficients were measured at 293 K for a range of additives in solvent systems of varying polarities and classes:  ethyl acetate, isobutyl alcohol, toluene, methyl ethyl ketone, methyl tert-butyl ether, and 0.2 mol·L-1 potassium phosphate buffer (pH 7). The additives chosen were based on common usage and represent a cross-section of the surfactant classes:  UCON LB-625, P2000, Triton X-100, sodium dodecyl sulfate (SDS), Tween 20, Tween 80, hexadecyltrimethylammonium bromide (CTAB), ammonium sulfate, and methyl-β-cyclodextrin. The partition behavior of these additives (except Tween 20) was also investigated. The effect of ionic strength, pH, and cosolvents on the partition coefficient was also determined to provide a database for surfactant and solubility enhancer behavior in order to allow for rapid optimization of initial extraction processes. The solubility results showed that the antifoams were extremely soluble in the organic solvents but had limited solubility in water. The nonionic surfactants were soluble in all solvents tested. The anionic surfactant was soluble in all solvents tested, with the exception of toluene. The cationic surfactant and ammonium sulfate had limited solubility in most solvents. The methyl-β-cyclodextrin had varying degrees of solubility depending on polarity. The partition results can largely be predicted from the solubility data, with the exception of the nonionic surfactants. For all of the compounds that partitioned, the behavior could also be predicted based on solvent polarity, with larger partition coefficients for the more polar solvents. These data can be used to design initial extraction processes containing these additives and, by analogy, for other related additives as well.

Enzymes solubilized in organic solvents, hosted within the polar cores of surfactant aggregates, known as reversed micelles, provide many unique opportunities for new biocatalytic synthesis and protein separation processes. Small-angle neutron scattering (SANS) studies have shown that insertion of the protein cytochrome-c in the reversed micelle polar core causes a significant reapportioning of the surfactants and water between the filled and unfilled micelles, leading to an increase in overall size of the filled micelles relative to their empty counterparts. The simple shell and core model assuming single occupancy of the reversed micelles has significant limitations in interpreting data for high protein loadings, and points to the need for more detailed characterization of the protein-reversed micelle interactions.