Hybrid process for genotoxics removal from active pharmaceutical ingredients combining organic solvent nanofiltration with polybenzimidazole adsorbents

Active pharmaceutical ingredients (APIs), as most of medicines, are obtained through chemical synthesis, using highly reactive reagents and usually, low levels of reagents, fractions of catalysts, or by-products are present in the final API or drug product as impurities. Some of these impurities have unwanted toxicities, including genotoxicity and carcinogenicity, and therefore related API administration risks for patient’s health has become an increasing concern of pharmaceutical companies, regulatory authorities, patients and doctors. European Medicines Agency (EMA) was the first agency to implement guidelines to control genotoxic impurities (GTIs), followed by the Food and Drug Administration (FDA). Both authorities agreed on setting as “Threshold of Toxicological Concern” (TTC) a limit at 1.5 micrograms per day for known and potential carcinogens [1,2]. Herein we report an approach based on the thermal treatment of a polybenzimidazole polymer providing novel adsorbent properties for genotoxic removal to purify API post-reaction streams. These novel adsorbents were tested for GTIs, such as methyl p-toluenesulfonate, p-toluenesulfonic acid, 4,4-dimethylamino pyridine and ethanesulfonyl chloride, from API solutions of Mometasone furoate (Meta). The current work will present a strategy of a hybrid process combining organic solvent nanofiltration (OSN) of API post-reaction streams with the adsorbers studied. The aim to include an adsorber stage, after the OSN operation, is to polish the retentate API solution for further purification or/and recovery of API lost through the permeate stream. Results will illustrate process efficiency gains. References [1] Teasdale A.; Elder D.; Chang S. J.; Wang S.; Thompson R.; Benz N.; Flores I. H. S., Org. Process Res. Dev. 17, 2013, 221-230. [2] EMEA Guidelines on the “Limits on Genotoxic Impurities”, EMEA/CHMP/QWP/251344/2006, 2006. Acknowledgements: We thank financial support from Fundação para a Ciência e Tecnologia (FCT) through the Project SelectHost (PTDC/QEQ-PRS/4157/2014) and iBB-Institute for Bioengineering and Biosciences (UID/BIO/04565/2013), from Programa Operacional Regional de Lisboa 2020 (Lisboa-01-0145-FEDER-007317) and from Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq – Brasil). We thank to Hovione PharmaScience Ltd for supplying the API and technical know-how

A membrane bioreactor for biotransformations of hydrophobic molecules using organic solvent nanofiltration (OSN) membranes

Thiswork reports the application of organic solvent nanofiltration (OSN) membranes to a membrane bioreactor for biotransformations (MBB). An organic solvent phasewas employed, allowing high substrate loadings and efficient product removal. The aqueous and organic phases were separated by an OSN membrane. The biotransformation of geraniol to R-citronellol by baker’s yeast was used as the model reaction, and n-hexadecane and toluene as the organic solvents. The performance of the MBB was compared to that of a direct contact biphasic (DCB) bioreactor. The MBB system resulted in lower productivities than the DCB system due to mass transfer limitations. For the n-hexadecane system, the membrane was the main mass transfer resistance, whereas for the toluene system the contribution of the aqueous liquid film mass transfer resistance became predominant. Further investigations are needed to improve the substrate transfer rates. Despite this, the MBB system prevented aqueous breakthrough, and thus the formation of two-phase emulsions. Toluene toxicity to the biocatalyst was also minimized, although it caused a reduction in the reaction enantiospecificity. This work showed that OSN-MBB systems avoid the formation of emulsions, thus reducing downstream separation and allowing increased substrate loadings

An Insight into the Release Kinetics

The authors acknowledge the financial support from Fundação para a Ciência e a Tecnologia (FCT-MEC), Portugal, through the dedicated project (PTDC/EDM-EDM/30828/2017) (BeLive) Publisher Copyright: © 2023 by the authors.This work explores the unique features of magnetic-responsive hydrogels to obtain liposomal hydrogel delivery platforms capable of precise magnetically modulated drug release based on the mechanical responses of these hydrogels when exposed to an external magnetic field. Magnetic-responsive liposomal hydrogel delivery systems were prepared by encapsulation of 1,2-dipalmitoyl-sn-glycero-3-phosphocoline (DPPC) multilayered vesicles (MLVs) loaded with ferulic acid (FA), i.e., DPPC:FA liposomes, into gelatin hydrogel membranes containing dispersed iron oxide nanoparticles (MNPs), i.e., magnetic-responsive gelatin. The FA release mechanisms and kinetics from magnetic-responsive liposomal gelatin were studied and compared with those obtained with conventional drug delivery systems, e.g., free liposomal suspensions and hydrogel matrices, to access the effect of liposome entrapment and magnetic field on FA delivery. FA release from liposomal gelatin membranes was well described by the Korsmeyer–Peppas model, indicating that FA release occurred under a controlled diffusional regime, with or without magnetic stimulation. DPPC:FA liposomal gelatin systems provided smoother controlled FA release, relative to that obtained with the liposome suspensions and with the hydrogel platforms, suggesting the promising application of liposomal hydrogel systems in longer-term therapeutics. The magnetic field, with low intensity (0.08 T), was found to stimulate the FA release from magnetic-responsive liposomal gelatin systems, increasing the release rates while shifting the FA release to a quasi-Fickian mechanism. The magnetic-responsive liposomal hydrogels developed in this work offer the possibility to magnetically activate drug release from these liposomal platforms based on a non-thermal related delivery strategy, paving the way for the development of novel and more efficient applications of MLVs and liposomal delivery systems in biomedicine.publishersversionpublishe

Protein-imprinted polymers: how far have “plastic antibodies” come?

Antibodies are highly selective and sensitive, making them the gold standard for recognition affinity tools. However, their production cost is high and their downstream processing is time-consuming. Molecularly imprinted polymers (MIPs) are tailor-made by incorporating specific molecular recognition sites in their structure, thus translating into receptor-like activity mode of action. The interest in molecular imprinting technology, applied to biomacromolecules, has increased in the past decade. MIPs, produced using biomolecules as templates, commonly referred to as “plastic antibodies” or “artificial receptors”, have been considered as suitable cheaper and easy to produce alternatives to antibodies. Research on MIPs, designed to recognize proteins or peptides is particularly important, with potential contributions towards biomedical applications, namely biosensors and targeted drug delivery systems. This mini review will cover recent advances on (bio)molecular imprinting technology, where proteins or peptides are targeted or mimicked for sensing and therapeutic applications. Polymerization methods are reviewed elsewhere, being out of the scope of this review. Template selection and immobilization approaches, monomers and applications will be discussed, highlighting possible drawbacks and gaps in research.Peer ReviewedPostprint (author's final draft

Magnetic field dynamic strategies for the improved control of the angiogenic effect of mesenchymal stromal cells

project PTDC/EDM-EDM/30828/2017 SFRH/BD/114043/2015 co-financed by the ERDF under the PT2020 Partnership Agreement (POVI-01-0145-FEDER-007265), as well as from POR Lisboa 2020 grant PRECISE (Project N. 16394). Publisher Copyright: © 2021 by the authors. Licensee MDPI, Basel, Switzerland.This work shows the ability to remotely control the paracrine performance of mesenchymal stromal cells (MSCs) in producing an angiogenesis key molecule, vascular endothelial growth factor (VEGF-A), by modulation of an external magnetic field. This work compares for the first time the application of static and dynamic magnetic fields in angiogenesis in vitro model, exploring the effect of magnetic field intensity and dynamic regimes on the VEGF-A secretion potential of MSCs. Tissue scaffolds of gelatin doped with iron oxide nanoparticles (MNPs) were used as a platform for MSC proliferation. Dynamic magnetic field regimes were imposed by cyclic variation of the magnetic field intensity in different frequencies. The effect of the magnetic field intensity on cell behavior showed that higher intensity of 0.45 T was associated with increased cell death and a poor angiogenic effect. It was observed that static and dynamic magnetic stimulation with higher frequencies led to improved angiogenic performance on endothelial cells in comparison with a lower frequency regime. This work showed the possibility to control VEGF-A secretion by MSCs through modulation of the magnetic field, offering attractive perspectives of a non-invasive therapeutic option for several diseases by revascularizing damaged tissues or inhibiting metastasis formation during cancer progression.publishersversionpublishe

Magnetic stimulation of the angiogenic potential of mesenchymal stromal cells in vascular tissue engineering

The authors acknowledge the financial support from Fundação para a Ciência e a Tecnologia (FCT-MEC), Portugal, through the dedicated project [PTDC/EDM-EDM/30828/2017] (BeLive) and PhD grant [SFRH/BD/114043/2015] and  through the project [EXPL/CTM-POL/1117/1135/2012] Moreover, the authors thanks POR Lisboa 2020 for the research project [PRECISE, Project N. 16394]. We acknowledge Dr. Marta Teixeira and the IPATIMUP facilities for the development of the ex vivo CAM experiments. The authors acknowledge Prof. Reyes Mallada (University of Zaragoza, Spain) for the use of the vibrating sample magnetometer (VSM) equipment and Dr. Pavel Strichovanec (University of Zaragoza, Spain) for the technical assistance provided during the experiments. We also acknowledge the Instituto de Medicina Molecular (IMM, Lisboa) for the services provided concerning the use of the Confocal Scanning Microscopy (Zeiss LSM 710). Publisher Copyright: © 2021 The Author(s). Published by National Institute for Materials Science in partnership with Taylor & Francis Group.The growing prevalence of vascular diseases worldwide has emphasized the need for novel tissue-engineered options concerning the development of vascularized 3D constructs. This study reports, for the first time, the use of external magnetic fields to stimulate mesenchymal stromal cells (MSCs) to increase the production of vascular endothelial growth factor-A (VEGF-A). Polyvinylalcohol and gelatin-based scaffolds, containing iron oxide nanoparticles, were designed for optimal cell magnetic stimulation. While the application of static magnetic fields over 24 h did not impact on MSCs proliferation, viability and phenotypic identity, it significantly increased the production of VEGF-A and guided MSCs morphology and alignment. The ability to enhance MSCs angiogenic potential was demonstrated by the increase in the number of new vessels formed in the presence of MSCs conditioned media through in vitro and in vivo models. Ultimately, this study uncovers the potential to manipulate cellular processes through short-term magnetic stimulation.publishersversionpublishe

Moesziomyces spp. cultivation using cheese whey: new yeast extract-free media, beta-galactosidase biosynthesis and mannosylerythritol lipids production

ABSTRACT: Mannosylerythritol lipids (MELs) are biosurfactants with excellent biochemical properties and a wide range of potential applications. However, high production costs, low productivity and unsatisfactory scale-up production have hampered commercial adoption. Herein, we report for the first time the beta-galactosidase production by Moesziomyces spp. from different sugars (D-galactose, D-glucose and D-lactose), with D-galactose being the best beta-galactosidase inducer, with 11.2 and 63.1 IU/mg(biomass), for Moesziomyces aphidis 5535(T) and Moesziomyces antarcticus 5048(T), respectively. The production of this enzyme allows to break down D-lactose and thus to produce MEL directly from D-lactose or cheese whey (a cheese industry by-product). Remarkably, when CW was used as sole media component (carbon and mineral source), in combination with waste frying oil, MEL productivities were very close (1.40 and 1.31 g(MEL)/L/day) to the ones obtained with optimized medium containing yeast extract (1.92 and 1.50 g(MEL)/g(susbtrate)), both for M. antarcticus and M. aphidis. The low-cost, facile and efficient process which generates large amounts of MELs potentiates its industrialization.info:eu-repo/semantics/publishedVersio

High cellulase-free xylanases production by Moesziomyces aphidis using low-cost carbon and nitrogen sources

ABSTRACT: Background Enzymes involved in xylan hydrolysis have several industrial applications. Selection of efficient microbial hosts and scalable bioreaction operations can lower enzyme production costs and contribute to their commercial deployment. This work aims at investigating the Moesziomyces aphidis yeast cultivation conditions that deliver maximal xylanase titres, yields and productivities using low-cost nitrogen (N) and carbon (C) sources. Results NaNO3 and KNO3 supplementation improved xylanase production 2.9- and 2.7-fold (against 67.2 U mL(-1)), respectively, using xylan as C source. Interestingly, the use of KNO3, instead of NaNO3, results in 2- to 3-fold higher specific activity, highlighting the potassium ion role. In addition, this study investigates synergetic effects on using ionic and organic N sources. A 4.9-fold increase in xylanase production, with high specific activity, is attained combining KNO3 and corn steep liquor (CSL). Exploring the previous findings, this study reports one of the highest extracellular xylanase production titres (864.7 U mL(-1)) by yeasts, using a media formulation containing dilute-acid pre-treated brewery spent grains (BSG), as C source and inducer, supplemented with KNO3 and CSL. Replacement of dilute-acid pre-treatmed BSG by untreated BSG had low impact on xylanase production, of only 6%. Conclusion Efficient production of M. aphidis xylanolytic enzymes, using low-cost N and C sources, is attractive for deployment of on-site enzyme production targeting different biotechnological applications under circular economy and biorefinery concepts. Potential xylanases end-users include industries such as brewing (using BSG as substrate for enzyme production), pulp and paper (benefiting from the cellulase-free xylanase activity) or lignocellulosic ethanol (for cellulase supplementation).info:eu-repo/semantics/publishedVersio