Frederico Castelo Ferreira

Recovery and reuse of ionic liquids and palladium catalyst for Suzuki reactions using organic solvent nanofiltration

The separation, post-reaction, of ionic liquids and catalysts from reaction products is an unresolved challenge in the application of ionic liquids to organometallic catalysis. This paper addresses this challenge using organic solvent nanofiltration technology. Suzuki reactions were carried out in a homogeneous solution, comprising 50∶50 wt% ethyl acetate and ionic liquid. The post reaction mixture was diluted further with ethyl acetate and then separated by nanofiltration into a permeate fraction and a retained (retentate) fraction. The product was recovered in the nanofiltration permeate, while the ionic liquid and palladium catalyst were retained by the membrane and recycled into subsequent consecutive reactions. Thus, the organic solvent nanofiltration was able to separate the Suzuki reaction product from both catalyst and ionic liquid. Three ionic liquids were tested: cocosalkyl pentaethoxi methyl ammonium methosulfate (ECOENG™500), tetrabutylammonium bromide (TBAB) and trihexyl(tetradecyl)phosphonium chloride (CyPhos®101). All the ionic liquids screened showed positive effects on the catalytic stability, significantly reducing the formation of palladium black and providing high reaction yields over consecutive recycles. The best performance was observed for the CyPhos®101 system. Additional investigations employing this ionic liquid showed that the reaction–recycle process can be successfully performed at lower catalyst–substrate ratios, leading to higher catalyst turnover numbers.

Neural stem cell differentiation by electrical stimulation using a cross-linked PEDOT substrate: Expanding the use of biocompatible conjugated conductive polymers for neural tissue engineering.

BACKGROUND The use of conjugated polymers allows versatile interactions between cells and flexible processable materials, while providing a platform for electrical stimulation, which is particularly relevant when targeting differentiation of neural stem cells and further application for therapy or drug screening. METHODS Materials were tested for cytotoxicity following the ISO10993-5. PEDOT PSS was cross-linked. ReNcellVM neural stem cells (NSC) were seeded in laminin coated surfaces, cultured for 4 days in the presence of EGF (20 ng/mL), FGF-2 (20 ng/mL) and B27 (20 μg/mL) and differentiated over eight additional days in the absence of those factors under 100Hz pulsed DC electrical stimulation, 1V with 10 ms pulses. NSC and neuron elongation aspect ratio as well as neurite length were assessed using ImageJ. Cells were immune-stained for Tuj1 and GFAP. RESULTS F8T2, MEH-PPV, P3HT and cross-linked PEDOT PSS (x PEDOT PSS) were assessed as non-cytotoxic. L929 fibroblast population was 1.3 higher for x PEDOT PSS than for glass control, while F8T2 presents moderate proliferation. The population of neurons (Tuj1) was 1.6 times higher with longer neurites (73 vs 108 μm) for cells cultured under electrical stimulus, with cultured NSC. Such stimulus led also to longer neurons. CONCLUSIONS x PEDOT PSS was, for the first time, used to elongate human NSC through the application of pulsed current, impacting on their differentiation towards neurons and contributing to longer neurites. GENERAL SIGNIFICANCE The range of conductive conjugated polymers known as non-cytotoxic was expanded. x PEDOT PSS was introduced as a stable material, easily processed from solution, to interface with biological systems, in particular NSC, without the need of in-situ polymerization.

Gelatin porous scaffolds fabricated using a modified gas foaming technique: characterisation and cytotoxicity assessment.

The current study presents an effective and simple strategy to obtain stable porous scaffolds from gelatin via a gas foaming method. The technique exploits the intrinsic foaming ability of gelatin in the presence of CO2 to obtain a porous structure stabilised with glutaraldehyde. The produced scaffolds were characterised using physical and mechanical characterisation methods. The results showed that gas foaming may allow the tailoring of the 3-dimensional structure of the scaffolds with an interconnected porous structure. To assess the effectiveness of the preparation method in mitigating the potential cytotoxicity risk of using glutaraldehyde as a crosslinker, direct and in-direct cytotoxicity assays were performed at different concentrations of glutaraldehyde. The results indicate the potential of the gas foaming method, in the preparation of viable tissue engineering scaffolds.

Environmental and economic analysis for selection and engineering sustainable API degenotoxification processes

The present paper discusses a comparative study concerning performance efficiency and sustainable impact of three purification processes for degenotoxification of Active Pharmaceutical Ingredients (APIs) post reaction streams: recrystallization, flash chromatography and organic solvent nanofiltration (OSN). Two case studies in each process were selected for evaluation of the separation technologies featuring the same model API being Mometasone furoate (Meta) glucocorticoid and two genotoxic impurities (GTIs). Methyl mesylate (MeMS) and 4-dimethylaminopyridine (DMAP) were chosen as model impurities due to their incidental appearance in glucocorticoids based on a common methanesulfonylation manufacturing step. Successful degenotoxification was achieved in all cases concerning DMAP and MeMS reaching final GTI levels below the regulatory thresholds with the exception of DMAP using recrystallization. API losses were 5% and 6.4% for OSN, 6.4% and 11.9% for flash chromatography and 14.9% and 16.4% for recrystallization during the removal of DMAP and MeMS, respectively. The API loss occurring during the purification processes has a significant impact on the outcome of cost analysis. Mass and carbon intensity values are highest for OSN and lowest for recrystallization, while flash chromatography has intermediate values. Although recrystallization is more time consuming, it should not be discarded without careful analysis since API loss often declines at larger scales and it generally gives the API in the desired crystallographic form. Solvent recycling has a significant impact on the sustainability of all the processes by the reduction of mass intensity by two orders of magnitude (from 400–1300 to 14–63 kg/kg-API, depending on the process) and narrowing down carbon intensity to the range of 100–200 kg-CO2/kg-API. OSN requires the use of 7 diavolumes, therefore its high performance is achieved at the cost of high solvent usage. Hence, solvent recycling also has a higher positive environmental impact on OSN.

Genotoxic Impurities in Pharmaceutical Manufacturing: Sources, Regulations, and Mitigation

Regulations, and Mitigation Gyorgy Szekely,*,† Miriam C. Amores de Sousa,‡ Marco Gil, Frederico Castelo Ferreira,*,‡ and William Heggie* †School of Chemical Engineering & Analytical Science, The University of Manchester, The Mill, Sackville Street, Manchester M13 9PL, United Kingdom ‡Department of Bioengineering and Institute for Bioengineering and Biosciences (iBB), Instituto Superior Tećnico, Universidade de Lisboa, Avenida Rovisco Pais, 1049-001, Lisbon, Portugal Hovione FarmaCiencia SA, R&D, Sete Casas, 2674-506, Loures, Portugal

Membrane aromatic recovery system (MARS): lab bench to industrial pilot scale

This article describes a novel process for recovery of aromatic amines and phenolic compounds form wastewaters, the membrane aromatic recovery aromatic system (MARS). Laboratory work on wastewaters containing aniline and phenol will be presented, including data demonstrating removal and recovery of each chemical in a sufficiently pure form to allow recycling into a chemical production process. This article also describes successful scale-up and operation of the process through pilot trials at Solutia, UK. Process economics are discussed and data showing the potential for application of the process to a wide range of organic chemicals are presented.

Organic solvent nanofiltration in asymmetric hydrogenation: enhancement of enantioselectivity and catalyst stability by ionic liquids

This communication describes the enhancement of the enantioselectivity and the stability of Ru-BINAP with the ionic liquid trihexyl(tetradecyl)phosphonium chloride (CyPhos101), and the use of organic solvent nanofiltration for the efficient separation of the catalyst and ionic liquid from the hydrogenation product, followed by simultaneous recycling of the catalyst and ionic liquid.

Dynamical Characterization of a Cellulose Acetate Polysaccharide

This work brings together dynamical and structural information at a molecular level for cellulose acetate being an original contribution to the general description of polysaccharide properties. In particular, it allowed reinterpreting the secondary relaxation mechanisms that are still controversial in the literature; a compilation of data provided by different authors is provided. Detailed dynamical information is provided by dielectric relaxation spectroscopy (DRS) (10(-1)-10(6) Hz) for cellulose acetate (CA) in the sub-T(g) region below ambient temperature; results were compared with cellulose acetate structured as an asymmetric membrane (CAmb). In samples with low water content, two secondary relaxation processes between 173 and 298 K were identified by DRS, associated with localized mobility. The process located at the lowest temperatures (process I) has a different mobility in CA relative to CAmb. The identical crystalline/amorphous state of both materials allowed rationalizing the distinct behavior in terms of polymeric arrangement and ability for water uptake. The looser structure of the CA relative to CAmb as confirmed by FTIR, TGA, and DSC analysis makes more sites accessible to water molecules, resulting in a higher water retention in CA (2.73% w/w) relative to CAmb (1.60% w/w) and an increased molecular mobility in the former due to a plasticizing effect. In both materials, process I is significantly influenced by hydration, shifting to higher frequencies and lower temperatures upon water uptake. This process seems to be associated with mobility occurring within the monomeric unit, which embraces the two anhydroglucose rings connected by the glycosidic linkage and the polar groups directly attached to it. It should involve a very limited length scale, as suggested by its location, far below the glass transition, and the tau(infinity) value with a low entropic effect. The relaxation process that emerges later, process II, is similar for both samples being much less influenced by water but experiencing a slight antiplasticizing effect shifting to lower frequencies and higher temperatures upon hydration. It should involve side group motions, strongly coupled to the mobility of the anhydroglucose rings, which become hindered probably due to establishment of H-bonds with water molecules. The plasticizing/antiplasticizing effect is being discussed only on the basis of the frequency position of the relaxation peak. Processes I and II merge into a broad relaxation (gamma(dry)) upon water removal in both CA and CAmb, however evolving slower in the former with drying, due to a more disordered structure of CA that allows water to interact with more internal sites in the polymer. At higher temperatures (T > or = 353 K), a process emerges in the high frequency side of the dynamic alpha-relaxation which is compatible with a beta(JG)-relaxation. The structured specimen CAmb provided an additional way to probe the morphological changes undergone by the material when annealed to temperatures higher than 353 K, originating an increase in the dielectric response. This effect can be associated with a skin densification and partial collapse of the membrane porous network, as observed by SEM.

Developing a co-culture system for effective megakaryo/thrombopoiesis from umbilical cord blood hematopoietic stem/progenitor cells

BACKGROUND AIMS Platelet transfusion can be a life-saving procedure in different medical settings. Thus, there is an increasing demand for platelets, of which shelf-life is only 5 days. The efficient ex vivo biomanufacturing of platelets would allow overcoming the shortages of donated platelets. METHODS We exploited a two-stage culture protocol aiming to study the effect of different parameters on the megakaryo/thrombopoiesis ex vivo. In the expansion stage, human umbilical cord blood (UCB)-derived CD34(+)-enriched cells were expanded in co-culture with human bone marrow mesenchymal stromal cells (BM-MSCs). The megakaryocytic commitment and platelet generation were studied, considering the impact of exogenous addition of thrombopoietin (TPO) in the expansion stage and a cytokine cocktail (Cyt) including TPO and interleukin-3 in the differentiation stage, with the use of different culture medium formulations, and in the presence/absence of BM-MSCs (direct versus non-direct cell-cell contact). RESULTS Our results suggest that an early megakaryocytic commitment, driven by TPO addition during the expansion stage, further enhanced megakaryopoiesis. Importantly, the results suggest that co-culture with BM-MSCs under serum-free conditions combined with Cyt addition, in the differentiation stage, significantly improved the efficiency yield of megakaryo/thrombopoiesis as well as increasing %CD41, %CD42b and polyploid content; in particular, direct contact of expanded cells with BM-MSCs, in the differentiation stage, enhanced the efficiency yield of megakaryo/thrombopoiesis, despite inhibiting their maturation. CONCLUSIONS The present study established an in vitro model for the hematopoietic niche that combines different biological factors, namely, the presence of stromal/accessory cells and biochemical cues, which mimics the BM niche and enhances an efficient megakaryo/thrombopoiesis process ex vivo.

Conversion of cellulosic materials into glycolipid biosurfactants, mannosylerythritol lipids, by Pseudozyma spp. under SHF and SSF processes

BackgroundMannosylerythritol lipids (MEL) are glycolipids with unique biosurfactant properties and are produced by Pseudozyma spp. from different substrates, preferably vegetable oils, but also sugars, glycerol or hydrocarbons. However, solvent intensive downstream processing and the relatively high prices of raw materials currently used for MEL production are drawbacks in its sustainable commercial deployment. The present work aims to demonstrate MEL production from cellulosic materials and investigate the requirements and consequences of combining commercial cellulolytic enzymes and Pseudozyma spp. under separate hydrolysis and fermentation (SHF) and simultaneous saccharification and fermentation (SSF) processes.ResultsMEL was produced from cellulosic substrates, Avicel® as reference (>99% cellulose) and hydrothermally pretreated wheat straw, using commercial cellulolytic enzymes (Celluclast 1.5 L® and Novozyme 188®) and Pseudozyma antarctica PYCC 5048T or Pseudozyma aphidis PYCC 5535T. The strategies included SHF, SSF and fed-batch SSF with pre-hydrolysis. While SSF was isothermal at 28°C, in SHF and fed-batch SSF, yeast fermentation was preceded by an enzymatic (pre-)hydrolysis step at 50°C for 48 h. Pseudozyma antarctica showed the highest MEL yields from both cellulosic substrates, reaching titres of 4.0 and 1.4 g/l by SHF of Avicel® and wheat straw (40 g/l glucan), respectively, using enzymes at low dosage (3.6 and 8.5 FPU/gglucan at 28°C and 50°C, respectively) with prior dialysis. Higher MEL titres were obtained by fed-batch SSF with pre-hydrolysis, reaching 4.5 and 2.5 g/l from Avicel® and wheat straw (80 g/l glucan), respectively.ConclusionsThis work reports for the first time MEL production from cellulosic materials. The process was successfully performed through SHF, SSF or Fed-batch SSF, requiring, for maximal performance, dialysed commercial cellulolytic enzymes. The use of inexpensive lignocellulosic substrates associated to straightforward downstream processing from sugary broths is expected to have a great impact in the economy of MEL production for the biosurfactant market, inasmuch as low enzyme dosage is sufficient for good systems performance.