a b s t r a c tA model is proposed for predicting the intrinsic rejections of linear flexible molecules, of high molecular dimensions, in microfiltration and ultrafiltration membranes. The developed model is based on the available theory of hindered transport of solutes through narrow pores, assuming that convection is the only relevant transport mechanism. For the application of the model, partition coefficients of the solutes between the solution and the membrane were estimated by a stochastic simulation, assuming suction of the macrosolutes into the pore. In these simulations, the macromolecules were modeled as freely jointed chains. Different methods for the estimation of the molecular dimensions of the solutes are discussed. The validity of the model was tested using dextran T2000 and plasmid pUC19 in its linear isoform, as model solutes; the tests were performed using membranes of different pore size, that include two ultrafiltration membranes previously characterized, having 4.1 and 10.5 nm of pore radius, and two track-etched polycarbonate membranes, having 15 and 40 nm of pore radius. Filtration was performed in a stirred cell, at controlled permeate flux. Different conditions of stirring speed and permeate flux were tested to check the consistency of the model. The results show that very satisfactory predictions are obtained, considering the simplicity of the model, especially in the case of the plasmid.

a b s t r a c tMicrofiltration and ultrafiltration operations were used in tandem to isolate and purify a 6050 bp plasmid DNA (pDNA). To achieve primary isolation of the plasmid from fermentation broths, immediately after cell lysis, a 0.2 mm microfiltration membrane was selected for solid/liquid separation, which was performed in a diafiltration mode, as an alternative to centrifugation. Then to attain plasmid concentration and purification, an ultrafiltration membrane with a pore radius of 4.1 nm was selected. Permeation of pDNA and RNA in the two membrane steps was modeled using recently published mass transfer models applicable to the permeation of closed segmented chains and freely-jointed chains, respectively. The permeation of proteins and genomic DNA (gDNA) was also studied in these operations.The microfiltration operation allowed high plasmid and RNA permeation, as expected. It was observed that significant amounts of gDNA, previously precipitated during the cell lysis step, reappear in solution during the diafiltration step. The effect of the ionic strength on this apparent re-solubilization was investigated, by testing the addition of two different salts to the diafiltration buffer: CH 3 COOK and CaCl 2 . The results show that these salts can be used to control gDNA apparent re-solubilization. During the ultrafiltration operation high plasmid retention with low adsorption was obtained under low ionic strength conditions. The results also show that a significant removal proteins and the re-solubilized gDNA is achieved, as well as some purification in respect to low molecular weight RNA, since all these components permeate through the ultrafiltration membrane.

A mass transfer model is proposed for predicting sieving coefficients, S obs , of supercoiled plasmid DNA (pDNA), in the presence of a salt, in membranes with narrow pores, i.e., pores smaller than the gyration radii of the plasmids to be considered for a certain application. The model assumes that permeation occurs due to plasmid suction at the membrane surface as a result of the convective flow, being the probability of permeation also dependent on the instantaneous molecular conformation of the plasmid, when getting close to the pore. Two different approaches are tested to model plasmid structure, that of a closed segmented chains (CSC) of double stranded DNA, and that of considering the superhelical chain as a freely jointed chains (FJC). Both approaches were used to estimate the radius of gyration, r g , of different plasmids by statistical simulation, and the obtained values were compared with experimental data available in the literature. A 6050 bp plasmid, pVAX1-LacZ, was used in the experimental work, in which filtration tests were performed using three different ultrafiltration membranes of known pore size, in a 10 ml stirred cell. At constant ionic strength, sieving coefficients were determined as a function of the permeate flux, J v , at two different values of stirring speed, ω. The results are in very good agreement with the model predictions at the highest stirring speed and the observed deviations found at the lowest stirring speed were interpreted with the aid of the developed model by considering the possibility of plasmid adsorption. Then, it was investigated the effect of changing the ionic strength of the medium at constant J v and ω. The obtained results clearly agree with the model predictions.

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a b s t r a c tThe demanding ever-increasing quantities of highly purified biomolecules by bio-industries, has triggered the development of new, more efficient, purification techniques. The application of membrane-based technologies has become very attractive in this field, for their high throughput capability, simplicity of operation and scale-up.Herein we report the production of a bi-layer membrane by electrospinning (ES), in which a support of poly e-caprolactone nanofibers was coated with a polyethylene oxide/sodium alginate layer, and subsequently cross-linked with calcium chloride. The membranes were characterized by SEM, ATR-FTIR, contact angle measurements, and were applied in the recovery process of a plasmid. The results show that membranes retained the suspended solids while allowing the permeation of plasmid DNA, with high recovery yields and improved RNA retention. Moreover, they also showed a very low fouling tendency. To the best of our knowledge it is the first time that ES membranes are applied in this type of bioprocess.