Extraction of the Bacterial Extracellular Polysaccharide FucoPol by Membrane-Based Methods: Efficiency and Impact on Biopolymer Properties

LA/P/0140/2020In this study, membrane-based methods were evaluated for the recovery of FucoPol, the fucose-rich exopolysaccharide (EPS) secreted by the bacterium Enterobacter A47, aiming at reducing the total water consumption and extraction time, while keeping a high product recovery, thus making the downstream procedure more sustainable and cost-effective. The optimized method involved ultrafiltration of the cell-free supernatant using a 30 kDa molecular weight cut-off (MWCO) membrane that allowed for a 37% reduction of the total water consumption and a 55% reduction of the extraction time, compared to the previously used method (diafiltration-ultrafiltration with a 100 kDa MWCO membrane). This change in the downstream procedure improved the product’s recovery (around 10% increase) and its purity, evidenced by the lower protein (8.2 wt%) and inorganic salts (4.0 wt%) contents of the samples (compared to 9.3 and 8.6 wt%, respectively, for the previously used method), without impacting FucoPol’s sugar and acyl groups composition, molecular mass distribution or thermal degradation profile. The biopolymer’s emulsion-forming and stabilizing capacity was also not affected (emulsification activity (EA) with olive oil, at a 2:3 ratio, of 98 ± 0% for all samples), while the rheological properties were improved (the zero-shear viscosity increased from 8.89 ± 0.62 Pa·s to 17.40 ± 0.04 Pa·s), which can be assigned to the higher purity degree of the extracted samples. These findings demonstrate a significant improvement in the downstream procedure raising FucoPol’s recovery, while reducing water consumption and operation time, key criteria in terms of process economic and environmental sustainability. Moreover, those changes improved the biopolymer’s rheological properties, known to significantly impact FucoPol’s utilization in cosmetic, pharmaceutical or food products.publishersversionpublishe

Dimebon enhances hippocampus-dependent learning in both appetitive and inhibitory memory tasks in mice

Dimebon, a compound recently proposed for a treatment of Alzheimer’s disorder was suggested to have memory enhancing properties in pre-clinical and clinical studies. We investigated whether dimebon at doses acutely (0.1 mg/kg and 0.5 mg/kg) or repeatedly (0.1 mg/kg) administered to mice via i.p. injections, increases memory scores respectively in an appetitive and an inhibitory learning task. Acute treatment with dimebon at the dose 0.1 mg/kg did not affect learning scores in either 3-month-old C57BL/6N or CD1 mice. Acute treatment with higher dose of dimebon (0.5mg/kg) was found to enhance inhibitory learning in 3- and 7-month-old mice as shown in the step-down avoidance paradigm in C57BL/6N mice. No effects on learning were seen in CD1 mice. In a model of appetitive learning, a spatial version of the Y-maze, repeated treatment with dimebon increased the rate of correct choices and decreased the latency of accessing a water reward after water deprivation. Repeated administration of dimebon also increased the duration of drinking behaviour during training/testing procedures although behaviours in others tests or water consumption were not altered. Acute treatment of water-deprived and non-water-deprived mice with dimebon also did not affect their water intake. Our data suggest that dimebon enhances hippocampus-dependent learning in both appetitive and inhibitory tasks in mice

Subcritical Water as a Pre-Treatment of Mixed Microbial Biomass for the Extraction of Polyhydroxyalkanoates

This work was financed by national funds from FCT-Fundação para a Ciência e a Tecnologia, I.P., in the scope of projects UIDP/04378/2020 and UIDB/04378/2020 of the Research Unit,on Applied Molecular Biosciences—UCIBIO and the project LA/P/0140/2020 of the Associate,Laboratory Institute for Health and Bioeconomy—i4HB, and by the Associate Laboratory for Green Chemistry—LAQV (UIDB/50006/2020). Liane Meneses, Asiyah Esmail, and Alexandre Paiva also acknowledge FCT I.P. for financial support through SFRH/BD/148510/2019, 2021.05014.BD, and IF/01146/2015, respectively.Polyhydroxyalkanoate (PHA) recovery from microbial cells relies on either solvent extraction (usually using halogenated solvents) and/or digestion of the non-PHA cell mass (NPCM) by the action of chemicals (e.g., hypochlorite) that raise environmental and health hazards. A greener alternative for PHA recovery, subcritical water (SBW), was evaluated as a method for the dissolution of the NPCM of a mixed microbial culture (MMC) biomass. A temperature of 150 degrees C was found as a compromise to reach NPCM solubilization while mostly preventing the degradation of the biopolymer during the procedure. Such conditions yielded a polymer with a purity of 77%. PHA purity was further improved by combining the SBW treatment with hypochlorite digestion, in which a significantly lower hypochlorite concentration (0.1%, v/v) was sufficient to achieve an overall polymer purity of 80%. During the procedure, the biopolymer suffered some depolymerization, as evidenced by the lower molecular weight (M-w) and higher polydispersity of the extracted samples. Although such changes in the biopolymer's molecular mass distribution impact its mechanical properties, impairing its utilization in most conventional plastic uses, the obtained PHA can find use in several applications, for example as additives or for the preparation of graft or block co-polymers, in which low-M-w oligomers are sought.publishersversionpublishe

Extraction Optimization and Polymer Characterization

Funding: This work was funded by the INGREEN project that has received funding from the Bio-based Industries Joint Undertaking (JU) under the European Union’s Horizon 2020 research and innovation programme under grant agreement No 838120. The JU receives support from the European Union’s Horizon 2020 research and innovation programme and the Bio-based Industries Consortium. This work was financed by national funds from FCT—Fundação para a Ciência e a Tecno-logia, I.P., in the scope of the project and the project LA/P/0140/2020 of the Associate Laboratory Institute for Health and Bioeconomy—i4HB. A.M. Rodrigues acknowledges the financial support of FCT through the PhD grant 2020.08574.BD. Publisher Copyright: © 2022 by the authors. Licensee MDPI, Basel, Switzerland.Polyhydroxyalkanoates (PHA) are biopolymers with potential to replace conventional oil-based plastics. However, PHA high production costs limit their scope of commercial applications. Downstream processing is currently the major cost factor for PHA production but one of the least investigated aspects of the PHA production chain. In this study, the extraction of poly(3-hydroxybutyrate-co-3-hydroxyvalerate) produced at pilot scale by a mixed microbial culture was performed using sodium hydroxide (NaOH) or sodium hypochlorite (NaClO) as digestion agents of non-PHA cellular mass. Optimal conditions for digestion with NaOH (0.3 M, 4.8 h) and NaClO (9.0%, 3.4 h) resulted in polymers with a PHA purity and recovery of ca. 100%, in the case of the former and ca. 99% and 90%, respectively, in the case of the latter. These methods presented higher PHA recoveries than extraction by soxhlet with chloroform, the benchmark protocol for PHA extraction. The polymers extracted by the three methods presented similar PHA purities, molecular weights and polydispersity indices. Using the optimized conditions for NaOH and NaClO digestions, this study analyzed the effect of the initial intracellular PHA content (40–70%), biomass concentration (20–100 g/L) and biomass pre-treatment (fresh vs. dried vs. lyophilized) on the performance of PHA extraction by these two methods.publishersversionpublishe

Characterization of the Thermostable Biosurfactant Produced by Burkholderia thailandensis DSM 13276

Funding Information: Technology: 31961133015; Shandong University: 31961133014). C.V.G., A.E. and A.T.R. acknowledge FCT I.P. for the Ph.D. grants 2021.05798.BD, 2021.05014.BD and 2020.06470.BD, respectively. Funding Information: ThTeh ethgelrymcoalli pstoapbeilpittyid oef bthioesuBr. ftahcatialanntdseencsriestbediobsuyrBfaucrtkahnotl dweraisa ethvaailluanatdeedn sbisyD exSpMos1i3n2g7 6 thwe absiodpeomlyomnsetrr aitne danto apquosesoeusss svoalluutaiobnle tsou raf atecme-apcetriavteuprer oopfe 1r2ti1e s°,Cnainm aenly ,aautloowclaCvMe (C0.9a8nd bar) for 20 min. Interestingly, the treated solution maintained its shear-thinning behavior, with a slight increase in the flow behavior index value (0.46) compared to the untre◦ ated biosurfactant solution (0.44) (refer to Figure S4 for supporting information on fitting the power law model). The thermally treated biosurfactant also maintained a surface tension value of 40.36 ± 0.5 mN/m, which was identical to that of the untreated solution (40.31 ± 0.agent26 mNin/mfood), thus conand/orfirmcosmeticing its prthermoducts/pral stabocessing,ility. and for bioremediation. The emulsifying ability of the treated biosurfactant, on the other hand, was negatively affected, with a reduction in the EA to 50.5 ± 0.9%, which is around half of the value observed for the non-treated biosurfactant (100.0 ± 0.0%). Different biosurfactants (e.g., glAyucothliopridCso)n atrlisbousthioonws:eCdoan scleipgthuta dliezcartieoans,eC i.nV.tGh.e, eF.mF.ualnsdifyCi.nAg.Vc.aTp.;amciettyhoadftoelroghye,aCt .tVre.Ga.t,mAe.Tn.tR ., at similar temperatures [64]. Nevertheless, the value is still within the range reported for good EA (≥50%) [59]. Moreover, Triton X-100 also suffered a similar reduction in its emulsification ability, as the EA of the emulsions stabilized with the autoclaved compound also reduced from 43.9 ± 2.1% to 33.4 ± 5.2%. These results underline the potential of the B. thIa.Pil.a, nindetnhseisscboipoesuorfftahcetapnrot jfeocrts uUseID, fPo/r0e4x37a8m/p20le20, ianntdheU fIDooBd/ 0i4n3d7u8/st2r0y2,0 inofwthheicRhestheaercthemUpneitr-on atAuprepslieadreMeolleevcuatlaerdBdiouscriienngcepsr—oUceCsIsBiInOgaonrd tthhee fpirnoajelcpt rLoAd/uPc/t0 i1s40c/o2n0s2u0mofetdh.e Associate Laboratory Institute for Health and Bioeconomy—i4HB, and by the European Union’s Horizon 2020 research and 4. Coninnovcalustionioprnos gram through Project Bio Innovation of a Circular Economy for Plastics (BioICEP), under The glycolipopeptide biosurfactant secreted by Burkholderia thailandensis DSM 13276 numbers: Institute of Microbiology, Chinese Academy of Sciences: 31961133016; Beijing Institute of was demonstrated to possess valuable surface-active properties, namely, a low CMC and high EA for almond and sunflower oils, and for benzene. Moreover, the biosurfactant showed good thermostability, with a thermal degradation temperature above 200 °C, and the ability to maintain stable rheological and surface-active properties, as well as good EA Publisher Copyright: © 2022 by the authors. Licensee MDPI, Basel, Switzerland.Biosurfactants synthesized by microorganisms represent safe and sustainable alternatives to the use of synthetic surfactants, due to their lower toxicity, better biodegradability and biocompatibility, and their production from low-cost feedstocks. In line with this, the present study describes the physical, chemical, and functional characterization of the biopolymer secreted by the bacterium Burkholderia thailandensis DSM 13276, envisaging its validation as a biosurfactant. The biopolymer was found to be a glycolipopeptide with carbohydrate and protein contents of 33.1 ± 6.4% and 23.0 ± 3.2%, respectively. Galactose, glucose, rhamnose, mannose, and glucuronic acid were detected in the carbohydrate moiety at a relative molar ratio of 4:3:2:2:1. It is a high-molecular-weight biopolymer (1.0×107Da) with low polydispersity (1.66), and forms aqueous solutions with shear-thinning behavior, which remained after autoclaving. The biopolymer has demonstrated a good emulsionstabilizing capacity towards different hydrophobic compounds, namely, benzene, almond oil, and sunflower oil. The emulsions prepared with the biosurfactant, as well as with its autoclaved solution, displayed high emulsification activity (>90% and ~50%, respectively). Moreover, the almond and sunflower oil emulsions stabilized with the biosurfactant were stable for up to 4 weeks, which further supports the potential of this novel biopolymer for utilization as a natural bioemulsifier.publishersversionpublishe

Preparation and characterization of films based on a natural P(3HB)/mcl-PHA blend obtained through the co-culture of Cupriavidus necator and Pseudomonas citronellolis in apple pulp waste

The co-culture of Cupriavidus necator DSM 428 and Pseudomonas citronellolis NRRL B-2504 was performed using apple pulp waste from the fruit processing industry as the sole carbon source to produce poly(3-hydroxybutyrate), P(3HB) and medium-chain length PHA, mcl-PHA, respectively. The polymers accumulated by both strains were extracted from the co-culture’s biomass, resulting in a natural blend that was composed of around 48 wt% P(3HB) and 52 wt% mcl-PHA, with an average molecular weight of 4.3 105 Da and a polydispersity index of 2.2. Two melting temperatures (Tm) were observed for the blend, 52 and 174 C, which correspond to the Tm of the mcl-PHA and P(3HB), respectively. P(3HB)/mcl-PHA blend films prepared by the solvent evaporation method had permeabilities to oxygen and carbon dioxide of 2.6 and 32 Barrer, respectively. The films were flexible and easily deformed, as demonstrated by their tensile strength at break of 1.47 0.07 MPa, with a deformation of 338 19% until breaking, associated with a Young modulus of 5.42 1.02 MPa. This study demonstrates for the first time the feasibility of using the co-culture of C. necator and P. citronellolis strains to obtain a natural blend of P(3HB)/mcl-PHA that can be processed into films suitable for applications ranging from commodity packaging products to high-value biomaterialsinfo:eu-repo/semantics/publishedVersio

Production of Medium-Chain Length Polyhydroxyalkanoates by Pseudomonas citronellolis Grown in Apple Pulp Waste

Background and objective: Apple pulp waste generated by the fruit processing industry is a sugar-rich material with great potential to be used as a feedstock for production of value added microbial products. The aim of this work was to use this feedstock for the cultivation of Pseudomonas citronellolis and production of medium-chain-length polyhydroxyalkanoates, a natural elastomer.Material and methods: The solid fraction of the apple pulp waste was discarded and the soluble fraction, rich in fructose (17.7 g l-1), glucose (7.5 g l-1) and sucrose (1.2 g l-1), was used for the batch bioreactor cultivation of Pseudomonas citronellolis NRRL B-2504.Results and conclusion: Pseudomonas citronellolis reached a polymer content in the biomass of 30% wt and a volumetric productivity of 0.025 g l-1 h-1. The polymer was mainly composed of 3-hydroxydecanoate (68% mol) and 3-hydroxyoctanoate (22% mol), with minor contents of 3-hydroxydodecanoate (5% mol), 3-hydroxytetradecanoate (4% mol) and 3-hydroxyhexanoate (1% mol), and had a molecular weight of 3.7×105 Da. It presented glass transition and melting temperatures of -12 and 53°C, respectively, and a thermal degradation temperature of 296°C. The polymer’s films were dense, ductile and permeable to oxygen and carbon dioxide. These results demonstrated that apple pulp waste is a suitable feedstock for the production of a biopolymer with properties that render it a promising alternative to some synthetic petrochemical-derived polyesters.Conflict of interest: The authors declare no conflict of interest

Poly(2-dimethylamino ethylmethacrylate)-Based Polymers To Camouflage Red Blood Cell Antigens

peer reviewedPoly(2-dimethylamino-ethylmethacrylate) (PDMAEMA) is a cationic polymer when dissolved in a 7.4 pH fluid. Owing to its ionic nature, this polycation interacts with the negatively charged cell membrane surface of red blood cells (RBCs). The electrostatic self-assembly of PDMAEMA on RBCs membrane can be employed for inducing the formation of a polymeric shield camouflaging blood group antigens on RBCs as a valuable strategy for developing “universal RBCs” for blood transfusion. The purpose of this research was to evaluate the camouflaging ability of PDMAEMA homopolymers and PDMAEMA-copoly(nethylene glycol) copolymers differing in molecular weight and architecture. Surprisingly, the PDMAEMAs caused a partially masking, no masking, and sensitization of the same RBCs population. The MW and architecture of the polymers as well as temperature of PDMAEMA-RBCs treatment influenced the results observed. Herein, the very particular reactivity of PDMAEMAs and RBCs is analyzed and discussed

Biodegradable Microparticles for Regenerative Medicine: A State of the Art and Trends to Clinical Application

peer reviewedTissue engineering and cell therapy are very attractive in terms of potential applications but remain quite challenging regarding the clinical aspects. Amongst the different strategies proposed to facilitate their implementation in clinical practices, biodegradable microparticles have shown promising outcomes with several advantages and potentialities. This critical review aims to establish a survey of the most relevant materials and processing techniques to prepare these micro vehicles. Special attention will be paid to their main potential applications, considering the regulatory constraints and the relative easiness to implement their production at an industrial level to better evaluate their application in clinical practices

Occurrence of non-toxic bioemulsifiers during polyhydroxyalkanoate production by Pseudomonas strains valorizing crude glycerol by-product

While screening for polyhydroxyalkanoate (PHA) producing strains, using glycerol rich byproduct as carbon source, it was observed that extracellular polymers were also secreted into the culture broth. The scope of this study was to characterize both intracellular and extracellular polymers, produced by Pseudomonas putida NRRL B 14875 and Pseudomonas chlororaphis DSM 50083, mostly focusing on those novel extracellular polymers. It was found that they fall into the class of bioemulsifiers (BE), as they showed excellent emulsion stability against different hydrocarbons/oils at various pH conditions, temperature and salinity concentrations. Cytotoxicity tests revealed that BE produced by P. chlororaphis inhibited the growth of highly pigmented human melanoma cells (MNT-1) by 50% at concentrations between 150-200 μg/mL, while no effect was observed on normal skin primary keratinocytes and melanocytes. This is the first study reporting mcl-PHA production by P. putida NRRL B-14785 and bioemulsifier production from both P. putida and P. chlororaphis strains