The preparation of megaporous bodies, with potential applications in biotechnology, was attempted by following several strategies. As a first step, naive and robust scaffolds were produced by polymerization of selected monomers in the presence of a highly soluble cross-linker agent. Ion-exchange function was incorporated by particle embedding, direct chemical synthesis, or radiation-induced grafting. The total ionic capacity of such systems was 1.5 mmol H(+)/g, 1.4 mmol H(+)/g, and 17 mmol H(+)/g, respectively. These values were in agreement with the ability to bind model proteins: observed dynamic binding capacity at 50% breakthrough was ≅7.2 mg bovine serum albumin/g, ≅7.4 hen egg-white lysozyme (HEWL) mg/g, and ≅108 HEWL mg/g. In the later case, total (static) binding capacity reached 220 mg/g. It was observed that the structure and size of the megapores remained unaffected by the grafting procedure which, however, allowed for the highest protein binding capacity. Lysozyme supported on grafted body showed extensive clarification activity against a Micrococcus lysodekticus suspension in the flow-through mode, i.e., 90% destruction of suspended microbial cells was obtained with a residence time ≈ 18 min. Both protein capture and biocatalysis applications are conceivable with the 3D-megaporous materials described in this work.

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Fibrous materials are prominent among novel chromatographic supports for the separation and purification of biomolecules. In this work, strong anion exchange, quaternary ammonium (Q) functional fibrous adsorbents were evaluated with regards to their physical and functional characteristics. A column packed with Q fibrous adsorbent illustrated the good column packing efficiency of theoretical plate height (H) values and higher permeability coefficients (>0.9 × 10 −7 cm 2 ) than commercial adsorbents. For pulse experiments with acetone and lactoferrin as tracers under nonbinding conditions, the total porosity (for acetone) and the interstitial porosity (for lactoferrin) measured 0.97 and 0.47, respectively. The total ionic capacity of the chemically-functionalized Q fiber was 0.51 mmol/mL. The results indicated that the Q fiber had a static binding capacity of 140 mg/mL and a dynamic binding capacity (DBC) of 76 mg/mL for bovine serum albumin (BSA) and showed a linearly-scalable factor (~110 mL) for a column volume with high capacity and high throughput. Furthermore, this adsorptive material had the ability to bind OPEN ACCESSProcesses 2015, 3 205 the high molecular weight protein, thyroglobulin, with a capacity of 6 mg/mL. This work demonstrated the column-packed Q fibrous adsorption system as a potential chromatography support that exhibits high capacity at higher flow rates.

BACKGROUND: Monoclonal antibodies are becoming a leading class of biopharmaceuticals but to increase their accessibility by the general population, new production processes must be developed in particular for the downstream processing. RESULTS: In this work, an alternative and innovative affinity chromatographic method using quaternary amine matrices is proposed. Separation is driven by the dual affinity ligand LYTAG-Z, composed of a choline binding polypeptide (LYTAG) and the synthetic antibody binding Z domain. A two-elution method was developed for the purification of mAbs and the performance of different anion exchangers containing quaternary amines that act as choline analogues -CIMmultus Q, Q Sepharose and gPore Q -were tested and compared, with both CIMmultus Q and Q Sepharose allowing a recovery of more than 94% of mAbs from a CHO cell supernatant with a purity greater than 95%.An integrated platform combining an initial affinity extraction step for the clarification and capture of mAbs and a subsequent chromatographic separation using Q-matrices for the polishing of mAbs is also proposed. LYTAG-Z triggers the extraction of 94.7 ± 1.7% mAbs to the PEG-rich phase, as opposed to 26.9 ± 0.6% in the absence of the ligand, using 7% PEG 3350 and 6% dextran 500 k. Further purification using Q Sepharose allowed a mAb recovery of 95.3 ± 1.4% with a purity level of 91.4 ± 13.0%. CONCLUSION: An integrated platform based on two purification steps -affinity extraction and affinity chromatography -results in an overall process yield of 90%, allowing the processing of mAbs directly from a non-clarified CHO cell culture. ATPS formulationPEG/dextran aqueous two-phase systems consisting of 7% (w/w) PEG 3350 Da and 6% (w/w) dextran 500 kDa were prepared by weighing the appropriate amounts of each component, namely PEG from stock solutions of 50% (w/w), dextran from a stock solution of 25% (w/w), biological sample and water, to a final weight of 10 g.The biological sample (pure human serum antibodies or CHO cell culture supernatant) was loaded at 35% (w/w). The LYTAG-Z ligand was loaded to the systems at double the mass concentration of the antibody (to a concentration of 100 mg L -1 of antibody, the concentration of LYTAG-Z was 200 mg L -1 ), to assure a molar ratio /jctb CHO genomic DNA quantification CHO genomic DNA (gDNA) content was determined by Quant-iT™ PicoGreen ® dsDNA reagent. Genomic DNA standards were prepared by serial dilution in elution buffer of genomic DNA extracted wileyonlinelibrary.com/jctb © 2017 Society of Chemical Industry J Chem Technol Biotechnol 2018; 93: [1966][1967][1968][1969][1970][1971][1972][1973][1974] The chromatographic profiles obtained with both convective flow devices are present in Fig. 3(A), and are very similar to the one obtained with Q Sepharose (in Fig. 2(A)), with (i) a large peak in the FT corresponding to most impurity proteins, (ii) small elution peaks obtained during low pH elution, and (iii) a larger peak

BACKGROUND: Complex polysaccharides are important in the pharmaceutical industry, yet, due to their large molecular weight and reduced charges, their purification is a highly demanding process that requires binding matrices with unique properties. This work demonstrates for the first time that complex polysaccharides biosynthesized by microalga Porphyridium purpureum can be adsorbed onto Q fibrous anion exchangers. RESULTSWhen the polysaccharides were characterized, the extent of sulfation was higher in native polysaccharides than in ethanol-or alkali-extracts. The zeta potentials increased with increasing pH and the highest charge was observed at pH 8, while the Z-average diameters of the polysaccharide at pH 6 were highest for alkali-extracts. Instead of pellicular resins, Q fibrous adsorbents were used to determine Langmuir thermodynamic properties and dynamic binding capacities. The parameters included static binding capacity and dissociation constant of 13.47 ± 1.02 mg g -1 and 0.141 ± 0.027 mg mL -1 , and 10 and 50% breakthrough capacities of 4.46 ± 0.22 and 5.51 ± 0.28 mg g -1 , respectively. The antiviral activity of the polysaccharides was demonstrated by minimizing bacteriophage lysis of Streptococcus thermophilus.CONCLUSION This work demonstrates that polysaccharide extraction can be optimized and the adsorption and desorption of a complex polysaccharide onto Q fibrous matrix is feasible. These parameters could be exploited for up-scaling of polysaccharides for nutraceutical and pharmaceutical applications. Primary recovery operations of sulfated polysaccharidesMicroalgae were pelleted by centrifugation at 3250 g and 4 • C for 45 min followed by addition of three parts of cold 99% ethanol to the microalgal supernatant. Likewise, polysaccharides were alkali extracted following the protocol of Miyajima and co-workers. 19 Both mixtures were shaken at 100 rpm for 10 min then kept at 4 • C for overnight extraction. The precipitates were separated from the aqueous phases by centrifugation as mentioned above. Pellets were resuspended in demineralized water, dialyzed through Spectra/Por regenerated cellulose membranes (MWCO, 12-14 kDa) overnight against double deionized water followed by lyophilization using an ALPHA 1-2 LD Plus (Martin Christ Gefriertrocknungsanlagen GmbH, Osterode am Harz, Germany). The ethanol-and alkali-extracted polysaccharides were designated 'Extract 1' and 'Extract 2', respectively.Determination of zeta potential and Z-average diameter of sulfated polysaccharide by light scattering P. purpureum polysaccharides were dissolved in 20 mmol L -1 citrate-phosphate buffer at pH 4, 5, 6, 7 and 8 to a concentration wileyonlinelibrary.com/jctb