Smart dynamic hybrid membranes with self-cleaning capability.

The growing freshwater scarcity has caused increased use of membrane desalination of seawater as a relatively sustainable technology that promises to provide long-term solution for the increasingly water-stressed world. However, the currently used membranes for desalination on an industrial scale are inevitably prone to fouling that results in decreased flux and necessity for periodic chemical cleaning, and incur unacceptably high energy cost while also leaving an environmental footprint with unforeseeable long-term consequences. This extant problem requires an immediate shift to smart separation approaches with self-cleaning capability for enhanced efficiency and prolonged operational lifetime. Here, we describe a conceptually innovative approach to the design of smart membranes where a dynamic functionality is added to the surface layer of otherwise static membranes by incorporating stimuli-responsive organic crystals. We demonstrate a gating effect in the resulting smart dynamic membranes, whereby mechanical instability caused by rapid mechanical response of the crystals to heating slightly above room temperature activates the membrane and effectively removes the foulants, thereby increasing the mass transfer and extending its operational lifetime. The approach proposed here sets a platform for the development of a variety of energy-efficient hybrid membranes for water desalination and other separation processes that are devoid of fouling issues and circumvents the necessity of chemical cleaning operations.

A workflow for the development of template-assisted membrane crystallization downstream processing for monoclonal antibody purification.

Monoclonal antibodies (mAbs) are commonly used biologic drugs for the treatment of diseases such as rheumatoid arthritis, multiple sclerosis, COVID-19 and various cancers. They are produced in Chinese hamster ovary cell lines and are purified via a number of complex and expensive chromatography-based steps, operated in batch mode, that rely heavily on protein A resin. The major drawback of conventional procedures is the high cost of the adsorption media and the extensive use of chemicals for the regeneration of the chromatographic columns, with an environmental cost. We have shown that conventional protein A chromatography can be replaced with a single crystallization step and gram-scale production can be achieved in continuous flow using the template-assisted membrane crystallization process. The templates are embedded in a membrane (e.g., porous polyvinylidene fluoride with a layer of polymerized polyvinyl alcohol) and serve as nucleants for crystallization. mAbs are flexible proteins that are difficult to crystallize, so it can be challenging to determine the optimal conditions for crystallization. The objective of this protocol is to establish a systematic and flexible approach for the design of a robust, economic and sustainable mAb purification platform to replace at least the protein A affinity stage in traditional chromatography-based purification platforms. The procedure provides details on how to establish the optimal parameters for separation (crystallization conditions, choice of templates, choice of membrane) and advice on analytical and characterization methods.

Template-Assisted Crystallization Behavior in Stirred Solutions of the Monoclonal Antibody Anti-CD20: Probability Distributions of Induction Times.

We present a method to determine the template crystallization behavior of proteins. This method is a statistical approach that accounts for the stochastic nature of nucleation. It makes use of batch-wise experiments under stirring conditions in volumes smaller than 0.3 mL to save material while mimicking larger-scale processes. To validate our method, it was applied to the crystallization of a monoclonal antibody of pharmaceutical interest, Anti-CD20. First, we determined the Anti-CD20 phase diagram in a PEG-400/Na2SO4/water system using the batch method, as, to date, no such data on Anti-CD20 solubility have been reported. Then, the probability distribution of induction times was determined experimentally, in the presence of various mesoporous silica template particles, and crystallization of Anti-CD20 in the absence of templates was compared to template-assisted crystallization. The probability distribution of induction times is shown to be a suitable method to determine the effect of template particles on protein crystallization. The induction time distribution allows for the determination of two key parameters of nucleation, the nucleation rate and the growth time. This study shows that the use of silica particles leads to faster crystallization and a higher nucleation rate. The template particle characteristics are shown to be critical parameters to efficiently promote protein crystallization.

Tuning the Electrochemical Properties of Novel Asymmetric Integral Sulfonated Polysulfone Cation Exchange Membranes.

In this study, novel asymmetric integral cation exchange membranes were prepared by the wet phase inversion of sulfonated polysulfone (SPSf) solutions. SPSf with different degrees of sulfonation (DS) was synthesized by variation in the amount of chlorosulfonic acid utilized as a sulfonating agent. The characterization of SPSf samples was performed using FTIR and 1H-NMR techniques. SPSf with a DS of 0.31 (0.67 meq/g corresponding ion exchange capacity) was chosen to prepare the membranes, as polymers with a higher DS resulted in poor mechanical properties and excessive swelling in water. By a systematic study, the opportunity to tune the properties of SPSf membranes by acting on the composition of the polymeric solution was demonstrated. The effect of two different phase inversion parameters, solvent type and co-solvent ratio, were investigated by morphological and electrochemical characterization. The best properties (permselectivity of 0.86 and electrical resistance of 6.3 Ω∙cm2) were obtained for the membrane prepared with 2-propanol (IPA):1-Methyl-2-pyrrolidinone (NMP) in a 20:80 ratio. This membrane was further characterized in different solution concentrations to estimate its performance in a Reverse Electrodialysis (RED) operation. Although the estimated generated power was less than that of the commercial CMX (Neosepta) membrane, used as a benchmark, the tailor-made membrane can be considered as a cost-effective alternative, as one of the main limitations to the commercialization of RED is the high membrane price.

Energy Harvesting from Brines by Reverse Electrodialysis Using Nafion Membranes.

Ion exchange membranes (IEMs) have consolidated applications in energy conversion and storage systems, like fuel cells and battery separators. Moreover, in the perspective to address the global need for non-carbon-based and renewable energies, salinity-gradient power (SGP) harvesting by reverse electrodialysis (RED) is attracting significant interest in recent years. In particular, brine solutions produced in desalination plants can be used as concentrated streams in a SGP-RED stack, providing a smart solution to the problem of brine disposal. Although Nafion is probably the most prominent commercial cation exchange membrane for electrochemical applications, no study has investigated yet its potential in RED. In this work, Nafion 117 and Nafion 115 membranes were tested for NaCl and NaCl + MgCl2 solutions, in order to measure the gross power density extracted under high salinity gradient and to evaluate the effect of Mg2+ (the most abundant divalent cation in natural feeds) on the efficiency in energy conversion. Moreover, performance of commercial CMX (Neosepta) and Fuji-CEM 80050 (Fujifilm) cation exchange membranes, already widely applied for RED applications, were used as a benchmark for Nafion membranes. In addition, complementary characterization (i.e., electrochemical impedance and membrane potential test) was carried out on the membranes with the aim to evaluate the predominance of electrochemical properties in different aqueous solutions. In all tests, Nafion 117 exhibited superior performance when 0.5/4.0 M NaCl fed through 500 µm-thick compartments at a linear velocity 1.5 cm·s-1. However, the gross power density of 1.38 W·m-2 detected in the case of pure NaCl solutions decreased to 1.08 W·m-2 in the presence of magnesium chloride. In particular, the presence of magnesium resulted in a drastic effect on the electrochemical properties of Fuji-CEM-80050, while the impact on other membranes investigated was less severe.

On the Aggregation and Nucleation Mechanism of the Monoclonal Antibody Anti-CD20 Near Liquid-Liquid Phase Separation (LLPS).

The crystallization of Anti-CD20, a full-length monoclonal antibody, has been studied in the PEG400/Na2SO4/Water system near Liquid-Liquid Phase Separation (LLPS) conditions by both sitting-drop vapour diffusion and batch methods. In order to understand the Anti-CD20 crystallization propensity in the solvent system of different compositions, we investigated some measurable parameters, normally used to assess protein conformational and colloidal stability in solution, with the aim to understand the aggregation mechanism of this complex biomacromolecule. We propose that under crystallization conditions a minor population of specifically aggregated protein molecules are present. While this minor species hardly contributes to the measured average solution behaviour, it induces and promotes crystal formation. The existence of this minor species is the result of the LLPS occurring concomitantly under crystallization conditions.

Anything but Conventional Chromatography Approaches in Bioseparation.

While packed bed chromatography, known as conventional chromatography, has been serving the biopharmaceutical industry for decades as the bioseparation method of choice, alternative approaches are likely to take an increasing leading role in the next few years. The high number of new biological drugs under development, and the need to make biopharmaceuticals widely accessible, has been driving the academia and industry in the quest of anything but conventional chromatography approaches. In this perspective paper, these alternative approaches are discussed in view of current and future challenges in the downstream processing field.

Chemical Vapor Deposition of Photocatalyst Nanoparticles on PVDF Membranes for Advanced Oxidation Processes.

The chemical binding of photocatalytic materials, such as TiO2 and ZnO nanoparticles, onto porous polymer membranes requires a series of chemical reactions and long purification processes, which often result in small amounts of trapped nanoparticles with reduced photocatalytic activity. In this work, a chemical vapor deposition technique was investigated in order to allow the nucleation and growth of ZnO and TiO2 nanoparticles onto polyvinylidene difluoride (PVDF) porous membranes for application in advanced oxidation processes. The thickness of obtained surface coatings by sputtered nanoparticles was found to depend on process conditions. The photocatalytic efficiency of sputtered membranes was tested against both a model drug and a model organic pollutant in a small continuous flow reactor.

Enzyme Crystals and Hydrogel Composite Membranes as New Active Food Packaging Material.

The great antimicrobial and antioxidant potential of enzymes makes them prone to be used as active packaging materials to preserve food from contamination or degradation. Major drawbacks are connected to the use of enzymes freely dispersed in solution, due to reduced protein stability. The immobilization of enzymes on solid supports to create biocatalytic interfaces has instead been proven to increase their stability and efficiency. In this work, it is shown that enzymes crystallized on hydrogel composite membranes (HCMs) can exert an effective antimicrobial action, thus making the composite membrane and crystals biofilm a potential active substrate for food packaging applications. The antimicrobial hen egg white lysozyme is crystallized on the surface of the hydrogel layer of HCMs, and its activity is determined by measuring the decrease in absorbance of Micrococcus lysodeikticus culture incubated with the specimen. The overall catalytic efficiency of the antimicrobial HCMs increases by a factor of 2 compared to the pure enzyme dissolved in solution at the same quantity. Because the enzyme in crystalline form is present in higher concentration and purity than in the solution, both its overall catalytic efficiency and antimicrobial action increase. Moreover, the hydrogel environment allows a better protein stabilization and retention during crystals dissolution.

Photothermal Membrane Distillation for Seawater Desalination.

Thermoplasmonic effects notably improve the efficiency of vacuum membrane distillation, an economically sustainable tool for high-quality seawater desalination. Poly(vinylidene fluoride) (PVDF) membranes filled with spherical silver nanoparticles are used, whose size is tuned for the aim. With the addition of plasmonic nanoparticles in the membrane, the transmembrane flux increases by 11 times, and, moreover, the temperature at the membrane interface is higher than bulk temperature.

Bioinspired Synthesis of CaCO3 Superstructures through a Novel Hydrogel Composite Membranes Mineralization Platform: A Comprehensive View.

Hydrogel composite membranes (HCMs) are used as novel mineralization platforms for the bioinspired synthesis of CaCO3 superstructures. A comprehensive statistical analysis of the experimental results reveals quantitative relationships between crystallization conditions and crystal texture and a strong selectivity toward complex morphologies when monomers bearing carboxyl and hydroxyl groups are used together in the hydrogel layer synthesis in HCMs.

Insights into the polymorphism of glycine: membrane crystallization in an electric field.

In this work we studied glycine crystallization with two main objectives: (i) to get improved control of crystal growth and polymorphic selectivity of organic molecules; (ii) to achieve additional insights into the nucleation mechanisms of glycine polymorphs. To reach these goals, membrane crystallization technology, a tool which allows improved control of supersaturation in solution crystallization, was used under different operating conditions: the variable solvent removal rate, acidic and almost neutral pH, the presence of a pulsed electric field. The traditional explanation for the crystallization of α and γ glycine polymorphs from aqueous solution is based on the general cyclic dimer hypothesis and the self-poisoning mechanism. In contrast with both the conventional theories, experimental results suggest that the relative nucleation rates with respect to the relative growth kinetics of the two forms under the different conditions play a dominant role in determining the polymorphic outcome. Our results instead support a molecular nucleation route where open chain dimers can behave as building units for both γ- and α-glycines in the rate determining structuring step of the two-step nucleation mechanism.

Multivariate analysis of quaternary carbamazepine-saccharin mixtures by X-ray diffraction and infrared spectroscopy.

Co-crystallization brings new opportunities for improving the solubility and dissolution rate of drugs with the chance of finely tuning some relevant chemical-physical properties of mixtures containing bioactive compounds. As co-crystallization process involves several molecular species, which are generally solid at room conditions, its control requires accurate knowledge and monitoring of the different phase that might appear during the formulation stage. In the present study the suitability of X-ray powder diffraction (XRPD) and Fourier-transformed infrared (FTIR) spectroscopy in quantifying mixtures of carbamazepine polymorphs (forms I and III), saccharin, and carbamazepine-saccharin cocrystals (form I) is assessed. Quaternary crystalline mixtures typically produced in the process of co-crystal production were analyzed by multivariate methods. Principal component analysis (PCA) was used for the identification of the crystal phases, while unsupervised simultaneous fitting of the spectra from pure phases, or supervised partial least squares (PLS) methods were used for their quantitative determination. The performance of data analysis was enhanced by applying peculiar pre-processing methods, such as SNIP filtering in case of FTIR and PCA filtering in case of XRPD. It was found that, for XRPD data, the automatic multi-fitting procedures and PLS models developed in this study are able to quantify single phases in mixtures to an accuracy level comparable to that obtained by the widely used Rietveld method, which, however, requires knowledge of the crystal structures. For FTIR data the results here obtained prove that this technique can be used as a fast method for polymorph characterization.

Energetics of protein nucleation on rough polymeric surfaces.

Metropolis Monte Carlo (MC) algorithm of the two-dimensional Ising model is used to study the heterogeneous nucleation of protein crystals on rough polymeric surfaces. The theoretical findings are compared to those obtained from classical nucleation theory (CNT), and to experimental data from protein model hen egg white lysozyme (HEWL) crystallized on poly(vinylidene fluoride) or PVDF, poly(dimethylsiloxane) or PDMS and Hyflon homemade membranes. The reduction of the activation energy for the nucleation process on polymeric membranes, predicted to occur at increasing surface roughness, results in a nucleation kinetics that is many orders of magnitude faster than in homogeneous phase. In general, MC stochastic dynamics offers the unique opportunity to investigate the effects of collective molecular aggregation at site level on the nucleation rate and, consequently, allows to identify optimal morphological and structural properties of polymeric membranes for a fine control of the crystallization kinetics.

Antisolvent membrane crystallization of pharmaceutical compounds.

This article describes a modification of the conventional membrane crystallization technique in which a membrane is used to dose the solvent/antisolvent composition to generate supersaturation and induce crystallization in a drug solution. Two operative configurations are proposed: (a) solvent/antisolvent demixing crystallization, where the solvent is removed in at higher flow rate than the antisolvent so that phase inversion promotes supersaturation and (b) antisolvent addition, in which the antisolvent is dosed into the crystallizing drug solution. In both cases, solvent/antisolvent migration occurs in vapor phase and it is controlled by the porous membrane structure, acting on the operative process parameters. This mechanism is different than that observed when forcing the liquid phases through the pores and the more finely controllable supersaturated environment would generate crystals with the desired characteristics. Two organic molecules of relevant industrial implication, like paracetamol and glycine, were used to test the new systems. Experiments demonstrated that, by using antisolvent membrane crystallization in both configurations, accurate control of solution composition at the crystallization point has been achieved with effects on crystals morphology.

Influence of the structural properties of poly(vinylidene fluoride) membranes on the heterogeneous nucleation rate of protein crystals.

In this study, the influence of the morphological parameters of microporous poly(vinylidene fluoride) (PVDF) membranes on the heterogeneous nucleation rate of hen egg white lysozyme (HEWL) crystals has been investigated. Experiments have been carried out on membranes prepared by non-solvent-induced phase inversion method, using PVDF-co-hexafluoropropylene (Kynarflex 2800) and PVDF homopolymer (Kinar 460), and adding LiCl and poly(vinylpyrrolidone) (PVP) in order to modulate the pore structure. From a theoretical point of view, the free Gibbs energy balance for the formation of a critical nucleus has been adapted to nonporous surfaces, thus obtaining a mathematical correlation between the energy nucleation barrier, the membrane porosity, and the contact angle between protein solution and polymeric substrate. The energetic barrier to heterogeneous nucleation was found to increase at higher contact angles-according to the prediction of classical theory-and to decrease at higher porosity. For instance, the predicted deltaG(het)/deltaG(hom) ratio for PVDF-Kynarflex (PVP 2.5%) membrane with porosity of 0.11 was 0.30, 35% lower with respect to the value calculated by the Volmer equation for a dense polymeric matrix having the same contact angle (87.4 +/- 5.8 degrees). In addition, the effect of the membrane pore size, porosity, and thickness on the removal rate of solvent have been discussed. For example, the transmembrane flux through PVDF-Kynar (LiCl 5.0%) membrane was 12% inferior than the one measured under the same experimental conditions through PVDF-Kynarflex (LiCl 7.5%) membrane, the latter having similar pore size and thickness but higher porosity (0.44 vs 0.32). The possibility to achieve rapidly a high level of supersaturation is expected to increase the nucleation rate. In general, measurements performed during crystallization tests carried out at pH 4.5 in NaAc 0.05 M buffer with different precipitant (NaCl) concentrations agree with the predicted trends.

Trypsin crystallization by membrane-based techniques.

To grow protein crystals is not an easy task; moreover, if we need to grow protein crystals with controlled shape, size, and size distribution, depending on their application, the mission becomes even harder. Membrane crystallization has been recognized as an interesting tool for growing protein crystals with enhanced crystallization kinetics, both in static and in forced solution flow configuration, without detrimental effects on crystal quality. In the present work, we have studied the membrane crystallization process of benzamidine inhibited trypsin from bovine pancreas (BPT), with ammonium sulphate (dissolved in Tris-HCl buffer, 0.1 M, pH 8.5), as precipitant agent. We have demonstrated that, by using the membrane crystallization technique, BPT crystals can be obtained in 24-48 h, in static configuration, and in 4-7 days, in a forced solution flow system, depending on the experimental conditions. Furthermore, the kinetics of BPT crystallization have been modulated, to control the morphological characteristics of the crystals produced, by an accurate selection of the operative parameters involved in the process. The active membrane surface and the flow rate of extraction solvent in quiescent configuration, and the solution velocity in forced convection solution experiments, were the parameters investigated. In this respect, membrane crystallization techniques have been assessed as an interesting way for growing proteins, and more specifically enzyme crystals, with high control on the final properties of the crystalline material produced, with potential fundamental implication in the field of structural biology and biotechnology.