Use of a ceramic membrane to improve the performance of two-separate-phase biocatalytic membrane reactor

Biocatalytic membrane reactors (BMR) combining reaction and separation within the same unit have many advantages over conventional reactor designs. Ceramic membranes are an attractive alternative to polymeric membranes in membrane biotechnology due to their high chemical, thermal and mechanical resistance. Another important use is their potential application in a biphasic membrane system, where support solvent resistance is highly needed. In this work, the preparation of asymmetric ceramic hollow fibre membranes and their use in a two-separate-phase biocatalytic membrane reactor will be described. The asymmetric ceramic hollow fibre membranes were prepared using a combined phase inversion and sintering technique. The prepared fibres were then used as support for lipase covalent immobilization in order to develop a two-separate-phase biocatalytic membrane reactor. A functionalization method was proposed in order to increase the density of the reactive hydroxyl groups on the surface of ceramic membranes, which were then amino-activated and treated with a crosslinker. The performance and the stability of the immobilized lipase were investigated as a function of the amount of the immobilized biocatalytst. Results showed that it is possible to immobilize lipase on a ceramic membrane without altering its catalytic performance (initial residual specific activity 93%), which remains constant after 6 reaction cycles

Pectin as a non-toxic crosslinker for durable and water-resistant biopolymer-based membranes with improved mechanical andfunctional properties

Biopolymer-based hydrophilic membranes with very high stability in aqueous systems were developed by using pectin as a non-toxic crosslinker. The unique properties of pectin as an efficient crosslinker were demonstrated using poly(vinyl alcohol) as a model for a highly water-soluble biopolymer. The chemical crosslinking strategy using glutaraldhehyde has proven successful in improving the stability of poly(vinyl alcohol) membranes. However, the use of non-toxic biological crosslinking agents has not been fully explored. We hypothesized that pectin, as a biopolymer bearing numerous carboxyl groups, could be a very efficient crosslinker compared to carboxylic acids, promoting unprecedented membrane stability. A systematic characterization of the chemical, thermal, mechanical, and functional properties of membranes prepared from poly(vinyl alcohol) crosslinked with pectin confirmed the excellent stability of the membranes in water, tested at the boiling point and at acidic and basic pH. The use of pectin also resulted in membranes with very high tensile strength, resistance to microbial degradation, antiradical and antibacterial activity, and improved water vapor barrier propertie

Biorefinery of olive leaves to produce dry oleuropein aglycone:use of homemade ceramic capillary biocatalytic membranes in a multiphase system

Oleuropein aglycone is an important antioxidant compound produced during oleuropein hydrolysis, not yet commercially available. Its production from renewable material by green processes is a challenge because it permits waste re-use and low environmental impact. In this work, homemade asymmetric capillary ceramic membranes were used to develop biocatalytic membranes, which were further used to produce oleuropein aglycone from olive leaves and/or commercial oleuropein. Results indicated that the biocatalytic system (containing covalently immobilized β-glucosidase) promotes the hydrolysis of oleuropein in both monophase and multiphase processes. Furthermore, the multiphase biocatalytic system enables the extraction of the hydrophobic oleuropein aglycone in an organic phase, before its rearrangement in water. This was achieved by the production, of an unstable water-in-oil emulsion (permeate side), on the basis of membrane emulsification process. The intensified biocatalytic/extractor system allowed taking shelter the hydrophobic compound in the organic phase with good efficiency (90%), protecting it from rearrangement

Pectinases immobilization on magnetic nanoparticles and their anti-fouling performance in a biocatalytic membrane reactor

Enzyme immobilization on commercial superparamagnetic nanoparticles (NPSP) was performed using covalent bonding. The biofunctionalized NPSP was then immobilized on the surface of the membrane using an external magnetic field to form a magneto-responsive biocatalytic membrane reactor (BMRSP). The magnetically formed smart nanolayer can be easily re-dispersed and recovered from the membrane when the enzyme is deactivated or whenever cleaning is required due to substrate over-accumulation. The system was used to hydrolyze pectin contained in different streams. Results are supported with complementary data from hydrodynamic, kinetic and morphological characterization in a flow-through reactive filtration. Wavelength-dispersive X-ray spectroscopy (WDS) elemental mapping revealed that the NPSP are uniformly dispersed on the surface of the membrane forming a thin biocatalytic layer. Both results of hydrodynamic studies and SEM micrographs of the membrane with the enzyme layer under various operating conditions, show that the immobilized enzyme effectively reduced membrane–foulant interaction. Comparison of filtration data using this commercial NPSP reveals good agreement with our previously used home-made NPSP. This implies that the scaling-up and commercialization of the developed BMRSP can be straightforward

Enzyme catalysis coupled with artificial membranes towards process intensification in biorefinery- a review

In this review, for the first time, the conjugation of the major types of enzymes used in biorefineries and the membrane processes to develop different configurations of MBRs, was analyzed for the production of biofuels, phytotherapics and food ingredients. In particular, the aim is to critically review all the works related to the application of MBR in biorefinery, highlighting the advantages and the main drawbacks which can interfere with the development of this system at industrial scale. Alternatives strategies to overcome main limits will be also described in the different application fields, such as the use of biofunctionalized magnetic nanoparticles associated with membrane processes for enzyme re-use and membrane cleaning or the membrane fouling control by the use of integrated membrane process associated with MBR

Comparison between Lipase Performance Distributed at the O/W Interface by Membrane Emulsification and by Mechanical Stirring

Multiphase bioreactors using interfacial biocatalysts are unique tools in life sciences such as pharmaceutical and biotechnology. In such systems, the formation of microdroplets promotes the mass transfer of reagents between two different phases, and the reaction occurs at the liquid–liquid interface. Membrane emulsification is a technique with unique properties in terms of precise manufacturing of emulsion droplets in mild operative conditions suitable to preserve the stability of bioactive labile components. In the present work, membrane emulsification technology was used for the production of a microstructured emulsion bioreactor using lipase as a catalyst and as a surfactant at the same time. An emulsion bioreaction system was also prepared by the stirring method. The kinetic resolution of (S,R)-naproxen methyl ester catalyzed by the lipase from Candida rugosa to obtain (S)-naproxen acid was used as a model reaction. The catalytic performance of the enzyme in the emulsion systems formulated with the two methods was evaluated in a stirred tank reactor and compared. Lipase showed maximum enantioselectivity (100%) and conversion in the hydrolysis of (S)-naproxen methyl ester when the membrane emulsification technique was used for biocatalytic microdroplets production. Moreover, the controlled formulation of uniform and stable droplets permitted the evaluation of lipase amount distributed at the interface and therefore the evaluation of enzyme specific activity as well as the estimation of the hydrodynamic radius of the enzyme at the oil/water (o/w) interface in its maximum enantioselectivity

New developments for the controlled fabrication of microstructured multiphase bioreactor using membrane emulsification technology

Over the past 20 years, there has been a growing interest in 'membrane emulsification' (ME). The distinguishing feature is that the resulting droplet size is controlled by the choice of the microporous membrane and not by drop breakup using shear or impact stress generated by mechanical agitation. The technique is highly attractive given its potentially lower energy demands and greater control over drop size distribution. Benefits of membrane emulsification for phase transfer biocatalysis could derive from low shear properties and structured phase with fine droplets. In this work, oil-in-water (O/W) emulsions with an heterogenized enzyme was produced by ME. The lipase was used as model enzyme. Lipase is an enantioselective phase transfer biocatalyst frequently used in esterifications, transesterifications and hydrolysis reactions accepting a broad range of even hydrophobic substrates with vast industrial importance. The enantioselective hydrolysis of racemic naproxen methyl ester was used a reaction model to produce (S)-naproxen, a member of the arylacetic acid group of nonsteroidal anti-inflammatory drugs. When emulsion is prepared with ME, lipase is distributed at the O/W interface while drops grow at the membrane pore opening, whereas the substrate is dissolved in the dispersed phase. Both droplets micromanufacturing and biocatalysts immobilization can be performed simultaneously and continuously. The interface with controlled and uniform size provided a constant reaction interface at steady-state. This methodology permits a controlled fabrication of monodispersed microstructured biocatalytic emulsion interface for a highly efficient enzymatic reaction. This new precisely controlled methodology to fabricate microstructured multiphase bioreactor was compared with conventional methods. Low shear stress and enzyme optimal spatial arrangement at the stable and constant oil/water interface permitted to obtain very high enantioselectivity (100%) at high conversion degree (up to 90% of the (S)-ester). In addition, thanks to the highly droplet uniformity and stability the methodology offered a possibility to accurately evaluate catalyst basic parameters such as the hydrodynamic diameter. This information is useful in the bioreactor optimization design. In this work, macromolecule diameter at the interface was evaluated and compared to the molecular diameter calculated from crystallographic data. The ME technology offers a novel, easy and scalable technique for producing microfabricated reactor for a large variety of process implementation in biotechnology, biomedicine, food, waste water treatment. The flexibility of the method permits to produce a wide variety of functional microparticles having targeted properties in terms of size distribution and composition by mixing various chemical or biological material and controlling membrane process parameters

Study and development of biocatalytic membrane reactors as biomimic plant cell systems for the production and separations of bioactive molecules from renewable materials

Doctorate Research in Plant Biology,XXII Ciclo,a.a. 2006-2009Università della Calabri

Advances in biocatalytic membrane reactors for the production of non-commercially available pharmacologically active compounds from vegetal material

The use of plant derived products has a considerable impact on the environment in terms of energy saving and waste reduction. Developing low-cost enzymes remains a priority for both the Department of Energy's Office of the Biomass Program and for private industry. To achieve this goals, scientific advance in enzyme technologies, including enzyme production and/or re-use of enzyme, are required. Biocatalytic membrane reactors are extremely compatible with labile molecule, it is possible to work in mild conditions, the equipments need small space, are flexible and easy to scale-up (they are enabling technologies and well respond to the process intensification strategy); operating costs are low; energy used is low; products are of high quality; co-products are also of high quality. In addition it is possible to carry out enzymatic reaction in a continuous way controlling the fluid-dynamic conditions and to re-use the biocatalyst. The two compartments that biocatalytic membranes configuration has, permits to conduct the enzymatic reaction in multiphasic systems, a configuration particularly useful to isolate water insoluble products. In this work advanced nanostructured microporous membranes functionalized with biomolecules of plant origin were developed to demonstrate that biocatalyst properly immobilized in membranes permit the development of catalytic organized systems able to simultaneously carry out the bioconversion and the product separation, in a single unit. The biocatalyst used was vegetal -glucosidase, the substrate was oleuropein, a phenolic waste component coming from olive oil production process that can be also easily extracted from olive leaves. The hydrolysis of oleuropein gives high added value compounds pharmacologically active (aglycon, hydroxytyrosol and dialdehydes). The expensive heterogenized enzyme resulted stable and efficient in the oleuropein hydrolysis. Kinetic parameters of -glucosidase immobilized in the membrane reactor have been evaluated and compared with those of the ones of the free enzyme. Results evidenced that intrinsic kinetics of immobilized enzyme was not negatively affected compared to the free enzyme. This shows that lower catalytic activity of immobilized enzyme, commonly observed in the literature, is not an intrinsic technological drawback and that appropriate control of biohybrid microstructured systems, microenvironment conditions and transpor

Biorefinery of Tomato Leaves by Integrated Extraction and Membrane Processes to Obtain Fractions That Enhance Induced Resistance against Pseudomonas syringae Infection

Tomato leaves have been shown to contain significant amounts of important metabolites involved in protection against abiotic and biotic stress and/or possessing important therapeutic properties. In this work, a systematic study was carried out to evaluate the potential of a sustainable process for the fractionation of major biomolecules from tomato leaves, by combining aqueous extraction and membrane processes. The extraction parameters (temperature, pH, and liquid/solid ratio (L/S)) were optimized to obtain high amounts of biomolecules (proteins, carbohydrates, biophenols). Subsequently, the aqueous extract was processed by membrane processes, using 30–50 kDa and 1–5 kDa membranes for the first and second stage, respectively. The permeate from the first stage, which was used to remove proteins from the aqueous extract, was further fractionated in the second stage, where the appropriate membrane material was also selected. Of all the membranes tested in the first stage, regenerated cellulose membranes (RC) showed the best performance in terms of higher rejection of proteins (85%) and lower fouling index (less than 15% compared to 80% of the other membranes tested), indicating that they are suitable for fractionation of proteins from biophenols and carbohydrates. In the second stage, the best results were obtained by using polyethersulfone (PES) membranes with an NMWCO of 5 kDa, since the greatest difference between the rejection coefficients of carbohydrates and phenolic compounds was obtained. In vivo bioactivity tests confirmed that fractions obtained with PES 5 kDa membranes were able to induce plant defense against P. syringae