Interpenetrating alginate network as drug delivery matrix: Effects on protein stability and release.

The study investigates the rheological properties and protein release capacity of a uniform hydrogel composed of sodium alginate (SA) and poloxamer (P407). The hydrogel is prepared through the sustained release of calcium ions, resulting in a reinforced and homogeneous interpenetrating networks (IPNs) of SA and P407 polymeric chains. By adjusting the amount of crosslink agent, the hydrogel exhibites an adjustable dissolution ratio and adaptable gelling time. Moreover, the composite showed a well-structured network and superior mechanical strength, enabling the sustained release of both calcium ions and Soybean Trypsin Inhibitor (STI) protein, a model of Bone Morphogenic Protein (BMP). Importantly, the protein release kinetic can be tuned based on the SA content in the polymeric blend, highlighting the versatile nature of this hydrogel for drug delivery purposes.

Selenium Bio-Nanocomposite Based on Alteromonas macleodii Mo169 Exopolysaccharide: Synthesis, Characterization, and In Vitro Antioxidant Activity.

In this study, the novel exopolysaccharide (EPS) produced by the marine bacterium Alteromonas macleodii Mo 169 was used as a stabilizer and capping agent in the preparation of selenium nanoparticles (SeNPs). The synthesized nanoparticles were well dispersed and spherical with an average particle size of 32 nm. The cytotoxicity of the EPS and the EPS/SeNPs bio-nanocomposite was investigated on human keratinocyte (HaCaT) and fibroblast (CCD-1079Sk) cell lines. No cytotoxicity was found for the EPS alone for concentrations up to 1 g L-1. A cytotoxic effect was only noticed for the bio-nanocomposite at the highest concentrations tested (0.5 and 1 g L-1). In vitro experiments demonstrated that non-cytotoxic concentrations of the EPS/SeNPs bio-nanocomposite had a significant cellular antioxidant effect on the HaCaT cell line by reducing ROS levels up to 33.8%. These findings demonstrated that the A. macleodii Mo 169 EPS can be efficiently used as a stabilizer and surface coating to produce a SeNP-based bio-nanocomposite with improved antioxidant activity.

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

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 °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 (Mw) 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-Mw oligomers are sought.

Polyhydroxyalkanoates from a Mixed Microbial Culture: Extraction Optimization and Polymer Characterization.

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.

Characterization of the Thermostable Biosurfactant Produced by Burkholderia thailandensis DSM 13276.

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 × 107 Da) with low polydispersity (1.66), and forms aqueous solutions with shear-thinning behavior, which remained after autoclaving. The biopolymer has demonstrated a good emulsion-stabilizing 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.

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

Tissue 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.

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

In 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.

Sustainable use of agro-industrial wastes as potential feedstocks for exopolysaccharide production by selected Halomonas strains.

Large quantities of waste biomass are generated annually worldwide by many industries and are vastly underutilized. However, these wastes contain sugars and other dissolved organic matter and therefore can be exploited to produce microbial biopolymers. In this study, four selected Halomonas strains, namely, Halomonas caseinilytica K1, Halomonas elongata K4, Halomonas smyrnensis S3, and Halomonas halophila S4, were investigated for the production of exopolysaccharides (EPS) using low-cost agro-industrial wastes as the sole carbon source: cheese whey, grape pomace, and glycerol. Interestingly, both yield and monosaccharide composition of EPS were affected by the carbon source. Glucose, mannose, galactose, and rhamnose were the predominant monomers, but their relative molar ratio was different. Similarly, the average molecular weight of the synthesized EPS was affected, ranging from 54.5 to 4480 kDa. The highest EPS concentration (446 mg/L) was obtained for H. caseinilytica K1 grown on cheese whey that produced an EPS composed mostly of galactose, rhamnose, glucose, and mannose, with lower contents of galacturonic acid, ribose, and arabinose and with a molecular weight of 54.5 kDa. Henceforth, the ability of Halomonas strains to use cost-effective substrates, especially cheese whey, is a promising approach for the production of EPS with distinct physicochemical properties suitable for various applications.

Preparation and Characterization of Porous Scaffolds Based on Poly(3-hydroxybutyrate) and Poly(3-hydroxybutyrate-co-3-hydroxyvalerate).

Poly(hydroxyalkanoates) (PHAs) with different material properties, namely, the homopolymer poly(3-hydroxybutyrate), P(3HB), and the copolymer poly(3-hydroxybutyrate-co-3-hydroxyvalerate, P(3HB-co-3HV), with a 3HV of 25 wt.%, were used for the preparation of porous biopolymeric scaffolds. Solvent casting with particulate leaching (SCPL) and emulsion templating were evaluated to process these biopolymers in porous scaffolds. SCPL scaffolds were highly hydrophilic (>170% swelling in water) but fragile, probably due to the increase of the polymer's polydispersity index and its high porosity (>50%). In contrast, the emulsion templating technique resulted in scaffolds with a good compromise between porosity (27-49% porosity) and hydrophilicity (>30% water swelling) and without impairing their mechanical properties (3.18-3.35 MPa tensile strength and 0.07-0.11 MPa Young's Modulus). These specifications are in the same range compared to other polymer-based scaffolds developed for tissue engineering. P(3HB-co-3HV) displayed the best overall properties, namely, lower crystallinity (11.3%) and higher flexibility (14.8% elongation at break. Our findings highlight the potency of our natural biopolyesters for the future development of novel porous scaffolds in tissue engineering, thanks also to their safety and biodegradability.

Polysaccharides as Stabilizers for Polymeric Microcarriers Fabrication.

Biodegradable polymeric microparticles are widely used in drug delivery systems with prolonged-release profiles and/or cell microcarriers. Their fabrication via the oil/water emulsion solvent evaporation technique has normally required emulsifiers in the aqueous phase. The present work aims to evaluate the effectiveness of various polysaccharides, such as chitosan, hyaluronic acid, cellulose, arabinogalactan, guar and their derivatives, as an alternative to synthetic surfactants for polylactide microparticle stabilization during their fabrication. Targeted modification of the biopolymer's chemical structure was also tested as a tool to enhance polysaccharides' emulsifying ability. The transformation of biomacromolecules into a form of nanoparticle via bottom-up or top-down methods and their subsequent application for microparticle fabrication via the Pickering emulsion solvent evaporation technique was useful as a one-step approach towards the preparation of core/shell microparticles. The effect of polysaccharides' chemical structure and the form of their application on the polylactide microparticles' total yield, size distribution and morphology was evaluated. The application of polysaccharides has great potential in terms of the development of green chemistry and the biocompatibility of the formed microparticles, which is especially important in biomedicine application.

Oxygen Plasma Treated-Electrospun Polyhydroxyalkanoate Scaffolds for Hydrophilicity Improvement and Cell Adhesion.

Poly(hydroxyalkanoates) (PHAs) with differing material properties, namely, the homopolymer poly(3-hydroxybutyrate), P(3HB), the copolymer poly(3-hydroxybutyrate-co-3-hydroxyvalerate), P(3HB-co-3HV), with a 3HV content of 25 wt.% and a medium chain length PHA, and mcl-PHA, mainly composed of 3-hydroxydecanoate, were studied as scaffolding material for cell culture. P(3HB) and P(3HB-co-3HV) were individually spun into fibers, as well as blends of the mcl-PHA with each of the scl-PHAs. An overall biopolymer concentration of 4 wt.% was used to prepare the electrospinning solutions, using chloroform as the solvent. A stable electrospinning process and good quality fibers were obtained for a solution flow rate of 0.5 mL h-1, a needle tip collector distance of 20 cm and a voltage of 12 kV for P(3HB) and P(3HB-co-3HV) solutions, while for the mcl-PHA the distance was increased to 25 cm and the voltage to 15 kV. The scaffolds' hydrophilicity was significantly increased under exposure to oxygen plasma as a surface treatment. Complete wetting was obtained for the oxygen plasma treated scaffolds and the water uptake degree increased in all treated scaffolds. The biopolymers crystallinity was not affected by the electrospinning process, while their treatment with oxygen plasma decreased their crystalline fraction. Human dermal fibroblasts were able to adhere and proliferate within the electrospun PHA-based scaffolds. The P(3HB-co-3HV): mcl-PHA oxygen plasma treated scaffold highlighted the most promising results with a cell adhesion rate of 40 ± 8%, compared to 14 ± 4% for the commercial oxygen plasma treated polystyrene scaffold AlvetexTM. Scaffolds based on P(3HB-co-3HV): mcl-PHA blends produced by electrospinning and submitted to oxygen plasma exposure are therefore promising biomaterials for the development of scaffolds for tissue engineering.

Production of medium-chain-length polyhydroxyalkanoates by Pseudomonas chlororaphis subsp. aurantiaca: Cultivation on fruit pulp waste and polymer characterization.

Pseudomonas chlororaphis subsp. aurantiaca DSM 19603 was cultivated on apple pulp, a glucose- and fructose-rich waste generated during juice production, to produce medium-chain length polyhydroxyalkanoates. A cell dry mass of 8.74 ± 0.20 g/L, with a polymer content of 49.25 ± 4.08% were attained. The produced biopolymer was composed of 42.7 ± 0.1 mol% 3-hydroxydecanoate, 17.9 ± 1.0 mol% 3-hydroxyoctanoate, 14.5 ± 1.1 mol% 3-hydroxybutyrate, 11.1 ± 0.6 mol% 3-hydroxytetradecanoate, 10.1 ± 0.5 mol% 3-hydroxydodecanoate and 3.7 ± 0.2 mol% 3-hydroxyhexanoate. It presented low glass transition and melting temperatures (-40.9 ± 0.7 °C and 42.0 ± 0.1 °C, respectively), and a degradation temperature of 300.0 ± 0.1 °C, coupled to a low crystallinity index (12.7 ± 2.7%), a molecular weight (Mw) of 1.34 × 105 ± 0.18 × 105 Da and a polydispersity index of 2.70 ± 0.03. The biopolymer's films were dense and had a smooth surface, as demonstrated by Scanning Electron Microscopy. They presented a tension at break of 5.21 ± 1.09 MPa, together with an elongation of 400.5 ± 55.8% and an associated Young modulus of 4.86 ± 1.49 MPa, under tensile tests. These attractive filming properties of this biopolymer could potentially be valorised in several areas such as the fine chemicals industry, biomedicine, pharmaceuticals, or food packaging.

Silver nanocomposites based on the bacterial fucose-rich polysaccharide secreted by Enterobacter A47 for wound dressing applications: Synthesis, characterization and in vitro bioactivity.

This study demonstrates the potential of a high molecular weight fucose-containing polysaccharide secreted by the bacterium Enterobacter A47, named FucoPol, and its silver nanocomposite as potential bioactive materials for wound dressings applications. A green, simple, light-assisted method was used for the synthesis of silver nanoparticles (AgNP) using FucoPol, as stabilizing and reducing agent. The synthesized nanoparticles were spherical, and the main population had a particle size in number ranging between 13 and 30 nm for percentiles 50 and 90, respectively. FucoPol, as well as the functionalized material, besides having no cytotoxicity towards human skin keratinocytes and mouse fibroblasts, also promoted in vitro keratinocytes migration. These observations not only show the safety of FucoPol and FucoPol/AgNP biocomposite, but also their wound healing ability. Moreover, the biocomposite had a strong antimicrobial activity against Staphylococcus aureus ATCC 6538 and Klebsiella pneumoniae CECT 8453, two representative strains of known skin commensal pathogens. These findings demonstrate for the first time the potential of FucoPol for the development of wound healing formulations. Additionally, the FucoPol/AgNP biocomposite might find use in antimicrobial biomaterials, including antibacterial wound healing formulations, which further strengthens the establishment of FucoPol as a bioactive biopolymer.

Biodegradable Cell Microcarriers Based on Chitosan/Polyester Graft-Copolymers.

Self-stabilizing biodegradable microcarriers were produced via an oil/water solvent evaporation technique using amphiphilic chitosan-g-polyester copolymers as a core material in oil phase without the addition of any emulsifier in aqueous phase. The total yield of the copolymer-based microparticles reached up to 79 wt. %, which is comparable to a yield achievable using traditional emulsifiers. The kinetics of microparticle self-stabilization, monitored during their process, were correlated to the migration of hydrophilic copolymer's moieties to the oil/water interface. With a favorable surface/volume ratio and the presence of bioadhesive natural fragments anchored to their surface, the performance of these novel microcarriers has been highlighted by evaluating cell morphology and proliferation within a week of cell cultivation in vitro.

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 biomaterials.

Correlation between Surface Properties of Polystyrene and Polylactide Materials and Fibroblast and Osteoblast Cell Line Behavior: A Critical Overview of the Literature.

Bone reconstruction remains an important challenge today in several clinical situations, notably regarding the control of the competition occurring during proliferation of osteoblasts and fibroblasts. Polystyrene and polylactide are reference materials in the biomedical field. Therefore, it could be expected from the literature that clear correlations have been already established between the behavior of fibroblasts or osteoblasts and the surface characteristics of these materials. After an extensive analysis of the literature, although general trends could be established, our critical review has highlighted the need to develop a more in-depth analysis of the surface properties of these materials. Moreover, the large variation noticed in the experimental conditions used for in vitro animal cell studies impairs comparison between studies. From our comprehensive review on this topic, we have suggested several parameters that would be valuable to standardize to integrate the data from the literature and improve our knowledge on the cell-material interactions.

Solid-State Synthesis of Water-Soluble Chitosan-g-Hydroxyethyl Cellulose Copolymers.

Graft copolymers of chitosan with cellulose ether have been obtained by the solid-state reactive mixing of chitin, sodium hydroxide and hydroxyethyl cellulose under shear deformation in a pilot twin-screw extruder. The structure and composition of the products were determined by elemental analysis and IR spectroscopy. The physicochemical properties of aqueous solutions of copolymers were studied as a function of the composition, and were correlated to the mechanical characteristics of the resulting films to assess the performance of new copolymers as coating materials, non-woven fibrous materials or emulsifiers for interface stabilization during the microparticle fabrication process.

Demonstration of the ability of the bacterial polysaccharide FucoPol to flocculate kaolin suspensions.

In this study, the flocculation properties of FucoPol, a bacterial extracellular polysaccharide, were investigated. FucoPol is a high molecular weight polymer and negatively charged due to the presence of glucuronic acid and the acyl groups succinyl and pyruvyl. High flocculation rate values (>70%) were achieved with a low bioflocculant dosage of 1 mg/L, for pH values in the range 3-5 and temperature within 15-20°C. The bioflocculant was also shown to be stable after freezing/thawing and heating up to 100°C. Given the polymer's anionic character, the size of flocs formed and their surface profile, bridging seems to be the main flocculation mechanism of FucoPol. This study demonstrated that FucoPol is a promising natural, biodegradable and biocompatible alternative to the currently used synthetic or inorganic hazardous products, with potential to be used as a novel flocculation agent in several applications, such as water treatment, food or mining. Further studies will involve evaluating the reduction of cation dosage on flocculation efficiency, as well as testing the applicability of FucoPol to flocculate different types of suspended solids, such as, for example, activated carbons, soil solids or yeast cells.

Pseudomonas chlororaphis as a multiproduct platform: Conversion of glycerol into high-value biopolymers and phenazines.

Pseudomonas chlororaphis subsp. aurantiaca DSM 19603 was cultivated using glycerol as the sole carbon source for the simultaneous production of medium-chain length polyhydroxyalkanoates (mcl-PHA), extracellular polysaccharide (EPS) and phenazines. A maximum cell dry mass of 11.79 g/L was achieved with a mcl-PHA content of 19 wt%, corresponding to a polymer concentration of 2.23 g/L. A considerably higher EPS production, 6.10 g/L, was attained. Phenazines synthesis was evidenced by the bright orange coloration developed by the culture during the cell growth phase. The mcl-PHA produced by P. chlororaphis was composed mainly of 3-hydroxydecanoate (50 wt%) with lower amounts of 3-hydroxyoctanoate (17 wt%), 3-hydroxytetradecanoate (17 wt%), 3-hydroxydodecanoate (13 wt%) and 3-hydroxyhexanoate (3 wt%). This PHA showed unique thermal features being highly amorphous, with a degree of crystallinity of 27% and a low melting temperature (45.0 °C). The secreted EPS was mostly composed of glucose, glucosamine, rhamnose and mannose, with smaller amounts of three other unidentified monomers. Although the bioprocess can be improved further to define the optimal conditions to produce each bioproduct (mcl-PHA, EPS or phenazines), this study has demonstrated for the first time the ability of P. chlororaphis to simultaneously produce three high-value products from a single substrate.

Phosphodiester Hydrogels for Cell Scaffolding and Drug Release Applications.

Given the major structural role phosphodiesters play in the organism it is surprising they have not been more widely adopted as a building block in sophisticated biomimetic hydrogels and other biomaterials. The potential benefits are substantial: phosphoester-based materials show excellent compatibility with blood, cells, and a remarkable resistance to protein adsorption that may trigger a foreign-body response. In this work, a novel class of phosphodiester-based ionic hydrogels is presented which are crosslinked via a phosphodiester moiety. The material shows good compatibility with blood, supports the growth and proliferation of tissue and presents opportunities for use as a drug release matrix as shown with fluorescent model compounds. The final gel is produced via base-induced elimination from a phosphotriester precursor, which is made by the free-radical polymerization of a phosphotriester crosslinker. This crosslinker is easily synthesized via multigram one-pot procedures out of common laboratory chemicals. Via the addition of various comonomers the properties of the final gel may be tuned leading to a wide range of novel applications for this exciting class of materials.