Browsing by Author "Freitas, Filomena"
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- Cation-mediated gelation of the fucose-rich polysaccharide FucoPol: preparation and characterization of hydrogel beads and their cytotoxicity assessmentPublication . Fialho, Letícia; Araújo, Diana; Delgado Alves, Vitor; Roma-Rodrigues, Catarina; Baptista, Pedro V.; Fernandes, Alexandra R.; Freitas, Filomena; Reis, Maria A.M.This study describes for the first time the iron- and copper-mediated gelation of FucoPol, fucoserich bacterial polysaccharide. The ability of FucoPol to gel in the presence of metal cations, including iron(III) and copper(II), was used for the preparation of hydrogel beads. Iron mediated the formation of stable and not cytotoxic gel beads, while copper resulted in fragile and cytotoxic ones. Copper-mediated beads coated with an iron-mediated gel layer were more stable and had reduced cytotoxicity. The resulting polymeric structures had differing morphology, physical properties and cytotoxicity, which support their use in several applications, including biomedicine, agriculture and bioremediation
- Characterisation of Films Based on Exopolysaccharides from Alteromonas Strains Isolated from French Polynesia Marine EnvironmentsPublication . Concórdio-Reis, Patrícia; Pereira, João R.; Delgado Alves, Vitor; Nabais, Ana R.; Neves, Luísa A.; Marques, Ana C.; Fortunato, Elvira; Moppert, Xavier; Guezennec, Jean; Reis, Maria A.M.; Freitas, FilomenaThis work assessed the film-forming capacity of exopolysaccharides (EPS) produced by six Alteromonas strains recently isolated from different marine environments in French Polynesia atolls. The films were transparent and resulted in small colour alterations when applied over a coloured surface (ΔEab below 12.6 in the five different colours tested). Moreover, scanning electron microscopy showed that the EPS films were dense and compact, with a smooth surface. High water vapour permeabilities were observed (2.7–6.1 × 10−11 mol m−1 s−1 Pa−1), which are characteristic of hydrophilic polysaccharide films. The films were also characterised in terms of barrier properties to oxygen and carbon dioxide. Interestingly, different behaviours in terms of their mechanical properties under tensile tests were observed: three of the EPS films were ductile with high elongation at break (ε) (35.6–47.0%), low tensile strength at break (Ꞇ) (4.55–11.7 MPa) and low Young’s modulus (εm) (10–93 MPa), whereas the other three were stiffer and more resistant with a higher Ꞇ (16.6–23.6 MPa), lower ε (2.80–5.58%), and higher εm (597–1100 MPa). These properties demonstrate the potential of Alteromonas sp. EPS films to be applied in different areas such as biomedicine, pharmaceuticals, or food packaging
- Characterization and biotechnological potential of extracellular polysaccharides synthesized by Alteromonas strains isolated from French Polynesia marine environmentsPublication . Concórdio-Reis, Patrícia; Delgado Alves, Vitor; Moppert, Xavier; Guezennec, Jean; Freitas, Filomena; Reis, Maria A.M.Marine environments comprise almost three quarters of Earth’s surface, representing the largest ecosystem of our planet. The vast ecological and metabolic diversity found in marine microorganisms suggest that these marine resources have a huge potential as sources of novel commercially appealing biomolecules, such as exopolysaccharides (EPS). Six Alteromonas strains from different marine environments in French Polynesia atolls were selected for EPS extraction. All the EPS were heteropolysaccharides composed of different monomers, including neutral monosaccharides (glucose, galactose, and mannose, rhamnose and fucose), and uronic acids (glucuronic acid and galacturonic acid), which accounted for up to 45.5 mol% of the EPS compositions. Non-carbohydrate substituents, such as acetyl (0.5–2.1 wt%), pyruvyl (0.2–4.9 wt%), succinyl (1–1.8 wt%), and sulfate (1.98–3.43 wt%); and few peptides (1.72–6.77 wt%) were also detected. Thermal analysis demonstrated that the EPS had a degradation temperature above 260 C, and high char yields (32–53%). Studies on EPS functional properties revealed that they produce viscous aqueous solutions with a shear thinning behavior and could form strong gels in two distinct ways: by the addition of Fe2+, or in the presence of Mg2+, Cu2+, or Ca2+ under alkaline conditions. Thus, these EPS could be versatile materials for different applications
- Chitin-glucan complex – Based biopolymeric structures using biocompatible ionic liquidsPublication . Ferreira, Inês C.; Araújo, Diana; Voisin, Pierre; Delgado Alves, Vitor; Rosatella, Andreia A.; Afonso, Carlos A.M.; Freitas, Filomena; Neves, Luísa A.This work explores the novelty of dissolving chitin-glucan complex (CGC), from two fungal strains, Komagataella pastoris (CGCP) and Aspergillus niger (CGCKZ) (KiOnutrime-CG™), using biocompatible ionic liquids (ILs). Three cholinium-based ILs were tested, choline acetate, choline propionate and choline hexanoate. Although all tested ILs resulted in the dissolution of the co-polymer at a concentration of 5 % (w/w), distinct polymeric structures, films or gels, were obtained from CGCP and CGCKZ, respectively. CGCP films were dense, flexible and elastic, with high swelling capacity (> 200 %). The IL anion alkyl chain length influenced the polymeric structures’ properties, namely, the CGCP films elongation at break and swelling degree. CGCKZ resulted in weak gels. For both polymeric structures, exposure to the ILs under the dissolution conditions caused significant changes in the co-polymers’ chemical structure, namely, reduction of their glucan moiety and reduction of the degree of acetylation, thus yielding chitosan-glucan complexes (ChGC) enriched in glucosamine (53.4 ± 0.3–60.8 ± 0.3 %)
- Chitin-Glucan Complex Hydrogels: Optimization of Gel Formation and Demonstration of Drug Loading and Release AbilityPublication . Araújo, Diana; Rodrigues, Thomas; Delgado Alves, Vitor; Freitas, FilomenaChitin-glucan complex (CGC) hydrogels were fabricated through a freeze–thaw procedure for biopolymer dissolution in NaOH 5 mol/L, followed by a dialysis step to promote gelation. Compared to a previously reported methodology that included four freeze–thaw cycles, reducing the number of cycles to one had no significant impact on the hydrogels’ formation, as well as reducing the total freezing time from 48 to 18 h. The optimized CGC hydrogels exhibited a high and nearly spontaneous swelling ratio (2528 +- 68%) and a water retention capacity of 55 +- 3%, after 2 h incubation in water, at 37 ºC. Upon loading with caffeine as a model drug, an enhancement of the mechanical and rheological properties of the hydrogels was achieved. In particular, the compressive modulus was improved from 23.0 +-0.89 to 120.0 +- 61.64 kPa and the storage modulus increased from 149.9 +- 9.8 to 315.0 +- 76.7 kPa. Although the release profile of caffeine was similar in PBS and NaCl 0.9% solutions, the release rate was influenced by the solutions’ pH and ionic strength, being faster in the NaCl solution. These results highlight the potential of CGC based hydrogels as promising structures to be used as drug delivery devices in biomedical applications
- Chitin-Glucan complex hydrogels: physical-chemical characterization, stability, in vitro drug permeation, and biological assessment in primary cellsPublication . Araújo, Diana; Rodrigues, Thomas; Roma-Rodrigues, Catarina; Alves, Vitor D.; Fernandes, Alexandra R.; Freitas, FilomenaChitin-glucan complex (CGC) hydrogels were fabricated by coagulation of the biopolymer from an aqueous alkaline solution, and their morphology, swelling behavior, mechanical, rheological, and biological properties were studied. In addition, their in vitro drug loading/release ability and permeation through mimic-skin artificial membranes (Strat-M) were assessed. The CGC hydrogels prepared from 4 and 6 wt% CGC suspensions (Na51*4 and Na51*6 hydrogels, respectively) had polymer contents of 2.40 0.15 and 3.09 0.22 wt%, respectively, and displayed a highly porous microstructure, characterized by compressive moduli of 39.36 and 47.30 kPa and storage moduli of 523.20 and 7012.25 Pa, respectively. Both hydrogels had a spontaneous and almost immediate swelling in aqueous media, and a high-water retention capacity (>80%), after 30 min incubation at 37 C. Nevertheless, the Na51*4 hydrogels had higher fatigue resistance and slightly higher-water retention capacity. These hydrogels were loaded with caffeine, ibuprofen, diclofenac, or salicylic acid, reaching entrapment efficiency values ranging between 13.11 0.49% for caffeine, and 15.15 1.54% for salicylic acid. Similar release profiles in PBS were observed for all tested APIs, comprising an initial fast release followed by a steady slower release. In vitro permeation experiments through Strat-M membranes using Franz diffusion cells showed considerably higher permeation fluxes for caffeine (33.09 g/cm2/h) and salicylic acid (19.53 g/cm2/h), compared to ibuprofen sodium and diclofenac sodium (4.26 and 0.44 g/cm2/h, respectively). Analysis in normal human dermal fibroblasts revealed that CGC hydrogels have no major effects on the viability, migration ability, and morphology of the cells. Given their demonstrated features, CGC hydrogels are very promising structures, displaying tunable physical properties, which support their future development into novel transdermal drug delivery platforms.
- Deacetylation and Desuccinylation of the Fucose-Rich Polysaccharide Fucopol: Impact on Biopolymer Physical and Chemical PropertiesPublication . Baptista, Sílvia; Araújo, Diana; Concórdio-Reis, Patrícia; Marques, Ana C.; Fortunato, Elvira; Delgado Alves, Vitor; Freitas, FilomenaFucoPol is an acylated polysaccharide with demonstrated valuable functional properties that include a shear thinning fluid behaviour, a film-forming capacity, and an emulsion forming and stabilizing capacity. In this study, the different conditions (concentration, temperature, and time) for alkaline treatment were investigated to deacylate FucoPol. Complete deacetylation and desuccinylation was achieved with 0.02 M NaOH, at 60 ºC for 15 min, with no significant impact on the biopolymer’s sugar composition, pyruvate content, and molecular mass distribution. FucoPol depyruvylation by acid hydrolysis was attempted, but it resulted in a very low polymer recovery. The effect of the ionic strength, pH, and temperature on the deacetylated/desuccinylated polysaccharide, d-FucoPol, was evaluated, as well as its emulsion and film-forming capacity. d-FucoPol aqueous solutions maintained the shear thinning behaviour characteristic of FucoPol, but the apparent viscosity decreased significantly. Moreover, contrary to FucoPol, whose solutions were not affected by the media’s ionic strength, the d-FucoPol solutions had a significantly higher apparent viscosity for a higher ionic strength. On the other hand, the d-FucoPol solutions were not affected by the pH in the range of 3.6–11.5, while FucoPol had a decreased viscosity for acidic pH values and for a pH above 10.5. Although d-FucoPol displayed an emulsification activity for olive oil similar to that of FucoPol (98 +- 0%) for an oil-to-water ratio of 2:3, the emulsions were less viscous. The d-FucoPol films were flexible, with a higher Young0s modulus (798 +- 152 MPa), a stress at the break (22.5 +- 2.5 MPa), and an elongation at the break (9.3 +- 0.7%) than FucoPol (458 +- 32 MPa, 15.5 +- 0.3 MPa and 8.1 +- 1.0%, respectively). Given these findings, d-FucoPol arises as a promising novel biopolymer, with distinctive properties that may render it useful for utilization as a suspending or emulsifier agent, and as a barrier in coatings and packaging films
- Exopolysaccharides enriched in rare suggars: bacterial sources, production and applicationsPublication . Roca, Christophe; Alves, Vitor D.; Freitas, Filomena; Reis, Maria A.M.Microbial extracellular polysaccharides (EPS), produced by a wide range of bacteria, are high molecular weight biopolymers, presenting an extreme diversity in terms of chemical structure and composition. They may be used in many applications, depending on their chemical and physical properties. A rather unexplored aspect is the presence of rare sugars in the composition of some EPS. Rare sugars, such as rhamnose or fucose, may provide EPS with additional biological properties compared to those composed of more common sugar monomers. This review gives a brief overview of these specific EPS and their producing bacteria. Cultivation conditions are summarized, demonstrating their impact on the EPS composition, together with downstream processing. Finally, their use in different areas, including cosmetics, food products, pharmaceuticals, and biomedical applications, are discussed
- Fucose-containing exopolysaccharide produced by the newly isolated Enterobacter strain A47 DSM 23139Publication . Freitas, Filomena; Alves, Vitor D.; Torres, Cristiana A.V.; Cruz, Madalena; Sousa, Isabel; Melo, Maria João; Ramos, Ana M.; Reis, Maria A.M.Enterobacter strain A47 (DSM 23139) was found to produce a fucose-containing exopolysaccharide (EPS). The EPS is composed of fucose, galactose, glucose, pyruvate, succinate and acetate in the molar ratios 1.6:1.3:1.1:1.2:0.7:1.5. It is a high molecular weight (5.8×106) homogeneous biopolymer, as indicated by the low polydispersity value (1.3). The steady shear flow properties of the EPS aqueous solutions are similar to guar gum and fucogel, and its viscoelastic properties indicate the formation of viscous aqueous solutions with entangled polymer chains. Additionally, the EPS has demonstrated good flocculating and emulsion stabilizing capacities, comparable to some commercially available products. These functional propertiesmakethe fucose-containing EPS a good alternative tomanysynthetic polymers, as well as other natural polysaccharides, in several applications in the food, pharmaceutical, cosmetic, textile, paper and petroleum industries.
- Kinetics of production and characterization of the fucose-containing exopolysaccharide from Enterobacter A47Publication . Torres, Cristiana A.V.; Marques, Rodolfo; Antunes, Sílvia; Alves, Vitor D.; Sousa, Isabel; Ramos, Ana Maria; Oliveira, Rui; Freitas, Filomena; Reis, Maria A.M.A fucose-containing exopolysaccharide (EPS) was produced by the bacterium Enterobacter A47 using glycerol byproduct from the biodiesel industry. The analysis of kinetic data suggested a partially growth associated EPS synthesis model. Although the EPS was composed of fucose, galactose and glucose at all cultivation stages, their relative proportion has varied considerably during the run. At the beginning (24 h), glucose was the main component (82.4 wt.%), being fucose and galactose minor components (5.0 wt.% and 10.9 wt.%, respectively), while at the end (96 h) it was composed of 26.0 wt.% fucose, 28.9 wt.% galactose and 43.7 wt.% glucose. The acyl groups content and composition have also changed, reaching their maximum content (19.2 wt.%) at the end of the run. Moreover, the molecular weight has increased linearly during the run (from 8 × 105 to 5 × 106). The changes observed in EPS composition and molecular weight have also had an impact upon the polymer’s intrinsic viscosity, as shown by its linear increase from 3.95 to 10.72 dL g−1. The results suggest that the culture might have synthesized at least two distinct EPS, with different sugar composition and average molecular weight, which predominated at different cultivation stages