Microbial- and seaweed-based biopolymers: Sources, extractions and implications for soil quality improvement and environmental sustainability - A review.

Improving soil quality without creating any environmental problems is an unescapable goal of sustainable agroecosystem management, according to the United Nations 2030 Agenda for Sustainable Development. Therefore, sustainable solutions are in high demand. One of these is the use of biopolymers derived from microbes and seaweed. This paper aims to provide an overview of the sources of extraction and use of microbial (bacteria and cyanobacteria) and seaweed-based biopolymers as soil conditioners, the characteristics of biopolymer-treated soils, and their environmental concerns. A preliminary search was also carried out on the entire Scopus database on biopolymers to find out how much attention has been paid to biopolymers as biofertilizers compared to other applications of these molecules until now. Several soil quality indicators were evaluated, including soil moisture, color, structure, porosity, bulk density, temperature, aggregate stability, nutrient availability, organic matter, and microbial activity. The mechanisms involved in improving soil quality were also discussed.

Adding Color to Mass Spectra of Biopolymers: Charge Determination Analysis (CHARDA) Assigns Charge State to Every Ion Peak.

Traditionally, mass spectrometry (MS) output is the ion abundance plotted versus the ionic mass-to-charge ratio m/z. While employing only commercially available equipment, Charge Determination Analysis (CHARDA) adds a third dimension to MS, estimating for individual peaks their charge states z starting from z = 1 and color coding z in m/z spectra. CHARDA combines the analysis of ion signal decay rates in the time-domain data (transients) in Fourier transform (FT) MS with the interrogation of mass defects (fractional mass) of biopolymers. Being applied to individual isotopic peaks in a complex protein tandem (MS/MS) data set, CHARDA aids peptide mass spectra interpretation by facilitating charge-state deconvolution of large ionic species in crowded regions, estimating z even in the absence of an isotopic distribution (e.g., for monoisotopic mass spectra). CHARDA is fast, robust, and consistent with conventional FTMS and FTMS/MS data acquisition procedures. An effective charge-state resolution Rz ≥ 6 is obtained with the potential for further improvements.

Development of basil seed mucilage (a heteropolysaccharide) - Polyvinyl alcohol biopolymers incorporating zinc oxide nanoparticles.

The effect of zinc oxide nanoparticles (ZnONPs) on various properties of basil seed mucilage (a heteropolysaccharide)-polyvinyl alcohol (BSM-PVA) films was investigated. Increasing concentration of ZnONPs in BSM-PVA films reduced moisture content (from 24.73 to 17.09 %), water solubility (from 36.36 to 27.65 %), water vapor permeability (from 4.66 × 10-7 to 2.55 × 10-7 g·m/m2·Pa·h), oxygen permeability (from 2.96 to 2.13 cm3 µm/m2 d kPa), and elongation at break (from 40.34 to 29.44 %), and increased ultimate tensile strength (from 16.81 to 21.48 MPa). Color and light transmission were affected by ZnONPs concentrations. The lack of formation of new peaks, along with the displacement of peaks, indicated the formation of hydrogen bonds between ZnONPs and the film matrix. Dynamic mechanical-thermal analysis showed that storage modulus and glass transition temperature increased with the concentration of ZnONPs. Scanning electron microscopy images illustrated that the addition of ZnONPs improved film integrity. X-ray diffraction pattern showed that the crystal shape of nanoparticles was preserved in the film matrix. Films containing ZnONPs showed good antibacterial activity against Staphylococcus aureus (reductions ≥ 3 log CFU/cm2) and Escherichia coli (reductions ≥ 4 log CFU/cm2). Films containing ZnONPs also showed a suitable antifungal activity during the storage of wheat bread.

Identification and Quantification of Monosaccharides from Fungal Cell Walls and Exopolysaccharides by Gas Chromatography Coupled to Mass Spectrometry.

The fungal cell wall and secreted exopolysaccharides play an important role in the interactions between fungi and their environment. Despite their central role in fungal biology, ecology, and host-pathogen interactions, the composition of these polymers and their synthetic pathways are not well understood. The protocols presented in this article describe an approach to isolate fungal cell wall polysaccharides and to identify and quantify the monosaccharide composition of these polymers by gas chromatography-mass spectrometry (GC-MS). © 2023 The Authors. Current Protocols published by Wiley Periodicals LLC. Basic Protocol: O-methyl trimethylsilyl monosaccharide derivatives composition analysis by GC-MS Support Protocol: Fungal cell wall extraction.

Uso de quitina e quitosana como adsorventes de amônia de efluentes aquícolas: revisão de literatura / Use of chitin and quitosan as adsorvents of ammonia of aquaculture effluents: a literature review / Uso de quitina y quitosana como adsorbentes de amonia de efluentes acuícolas: revisión de literatura

Os resíduos provenientes da aquicultura são derivados da ração e da excreção dos peixes e podem estar sedimentados, suspensos ou dissolvidos, ocasionando elevados valores de DBO, DQO, nitrogênio e fósforo. A produção de camarões no Brasil tem gerado elevadas quantidades de resíduos sólidos, tendo em vista que os exoesqueletos dos camarões correspondem a cerca de 40% do seu peso total, resultando num forte impacto ambiental. Diversas pesquisas envolvendo a quitina estão sendo desenvolvidas na área de tratamento de água, devido principalmente a sua capacidade de formar filme, sendo utilizada em sistemas filtrantes. Este polissacarídeo também pode ser utilizado como agente floculante no tratamento de efluentes, como adsorvente na clarificação de óleos, e principalmente na produção de quitosana. Atualmente a quitosana possui aplicações multidimensionais, desde áreas como a nutrição humana, biotecnologia, ciência dos materiais, indústria farmacêutica, agricultura, terapia genética e proteção ambiental. A quitosana é muito eficiente na remoção de poluentes em diferentes concentrações. Apresenta alta capacidade e grande velocidade de adsorção, boa eficiência e seletividade tanto em soluções que possuem altas ou baixas concentrações. O uso da biotecnologia, através do processo de adsorção utilizando adsorventes naturais e baratos, como a quitina e quitosana, minimiza os impactos ambientais da aquicultura tanto em relação aos provocados pelo lançamento de efluentes no meio ambiente quanto aos causados pelo descarte inadequado dos resíduos do processamento de camarões.(AU)

Aquaculture residues are derived from fish feed and excretion and may be sedimented, suspended or dissolved, resulting in high BOD, COD, nitrogen and phosphorus values. Shrimp production in Brazil has generated high amounts of solid waste, since shrimp exoskeletons account for about 40% of their total weight, resulting in a strong environmental impact. Several researches involving chitin are being developed in the area of water treatment, mainly due to its ability to form film, being used in filter systems. This polysaccharide can also be used as a flocculating agent in the treatment of effluents, as an adsorbent in the clarification of oils, and especially in the production of chitosan. Currently, chitosan has multidimensional applications, from areas such as human nutrition, biotechnology, materials science, pharmaceutical industry, agriculture, gene therapy and environmental protection. Chitosan is very efficient in the removal of pollutants at different concentrations. It presents high capacity and high adsorption velocity, good efficiency and selectivity both in solutions that have high or low concentrations. The use of biotechnology, through the adsorption process using natural and cheap adsorbents such as chitin and chitosan, minimizes the environmental impacts of aquaculture both in relation to those caused by the release of effluents into the environment and those caused by the inappropriate disposal of processing residues of shrimps.(AU)

Los residuos procedentes de la acuicultura se derivan de la ración y de la excreción de los peces y pueden estar sedimentados, suspendidos o disueltos, ocasionando elevados valores de DBO, DQO, nitrógeno y fósforo. La producción de camarones en Brasil ha generado grandes cantidades de residuos sólidos, teniendo en cuenta que los exoesqueletos de los camarones corresponden a cerca del 40% de su peso total, resultando en un fuerte impacto ambiental. Varias investigaciones involucrando la quitina se están desarrollando en el área de tratamiento de agua, debido principalmente a su capacidad de formar película, siendo utilizada en sistemas filtrantes. Este polisacárido también puede ser utilizado como agente floculante en el tratamiento de efluentes, como adsorbente en la clarificación de aceites, y principalmente en la producción de quitosana. Actualmente la quitosana posee aplicaciones multidimensionales, desde áreas como la nutrición humana, biotecnología, ciencia de los materiales, industria farmacéutica, agricultura, terapia genética y protección ambiental. La quitosana es muy eficiente en la eliminación de contaminantes en diferentes concentraciones. Presenta alta capacidad y gran velocidad de adsorción, buena eficiencia y selectividad tanto en soluciones que poseen altas o bajas concentraciones. El uso de la biotecnología, a través del proceso de adsorción utilizando adsorbentes naturales y baratos, como la quitina y quitosana, minimiza los impactos ambientales de la acuicultura tanto en relación a los provocados por el lanzamiento de efluentes en el medio ambiente en cuanto a los causados por el descarte inadecuado de los residuos del procesamiento de camarones.(AU)

A comprehensive review on the recent advancements in natural rubber nanocomposites.

Natural rubber (NR) is an eminent sustainable material and is the only agricultural product among various rubbers. Use of nanofillers in NR matrix as a reinforcing agent has gained huge attention because they offer excellent matrix-filler interaction upon forming a good dispersion in the NR matrix. Nanoscale dispersion of fillers lead to greater interfacial interactions between NR and fillers compared to microfillers, which in turn lead to a conspicuous reinforcing effect. Addition of various nanofillers into NR matrix improves not only the mechanical properties but also the electrical, thermal and antimicrobial properties to an extreme level. The current review describes the reinforcing ability of various nanofillers such as clay, graphene, carbon nanotube (CNT), titanium dioxide (TiO2), chitin, cellulose, barium titanate (BaTiO3) and lignin in NR matrix. Moreover, reinforcement of various hybrid nanofillers in NR is also discussed in a comprehensive manner. The review also includes the historical trajectory of rubber nanocomposites and a comprehensive account on the factors affecting the properties of the NR nanocomposites.

Integrative measurement analysis via machine learning descriptor selection for investigating physical properties of biopolymers in hairs.

Integrative measurement analysis of complex subjects, such as polymers is a major challenge to obtain comprehensive understanding of the properties. In this study, we describe analytical strategies to extract and selectively associate compositional information measured by multiple analytical techniques, aiming to reveal their relationships with physical properties of biopolymers derived from hair. Hair samples were analyzed by multiple techniques, including solid-state nuclear magnetic resonance (NMR), time-domain NMR, Fourier transform infrared spectroscopy, and thermogravimetric and differential thermal analysis. The measured data were processed by different processing techniques, such as spectral differentiation and deconvolution, and then converted into a variety of "measurement descriptors" with different compositional information. The descriptors were associated with the mechanical properties of hair by constructing prediction models using machine learning algorithms. Herein, the stepwise model refinement via selection of adopted descriptors based on importance evaluation identified the most contributive descriptors, which provided an integrative interpretation about the compositional factors, such as α-helix keratins in cortex; and bounded water and thermal resistant components in cuticle. These results demonstrated the efficacy of the present strategy to generate and select descriptors from manifold measured data for investigating the nature of sophisticated subjects, such as hair.

A Landscape of Lignocellulosic Biopolymer Transformations into Valuable Molecules by Heterogeneous Catalysis in C'Durable Team at IRCELYON.

This review article highlights part of the research activity of the C'Durable team at IRCELYON in the field of sustainable chemistry. This review presents a landscape of the work performed on the valorization of lignocellulosic biopolymers. These studies intend to transform cellulose, hemicellulose and lignin into valuable molecules. The methodology usually consists in evaluating the behavior of the biopolymers in the absence of catalyst under various conditions (solvent, temperature), and then to assess the influence of a catalyst, most often a heterogeneous catalyst, on the reactivity. The most significant results obtained on the upgrading of cellulose and lignin, which have been mainly investigated in the team, will be presented with an opening on studies involving raw lignocellulose.

Development and Characterization of Novel Biopolymer Derived from Abelmoschus esculentus L. Extract and Its Antidiabetic Potential.

Abelmoschus esculentus (Okra) is an important vegetable crop, widely cultivated around the world due to its high nutritional significance along with several health benefits. Different parts of okra including its mucilage have been currently studied for its role in various therapeutic applications. Therefore, we aimed to develop and characterize the okra mucilage biopolymer (OMB) for its physicochemical properties as well as to evaluate its in vitro antidiabetic activity. The characterization of OMB using Fourier-transform infrared spectroscopy (FT-IR) revealed that okra mucilage containing polysaccharides lies in the bandwidth of 3279 and 1030 cm-1, which constitutes the fingerprint region of the spectrum. In addition, physicochemical parameters such as percentage yield, percentage solubility, and swelling index were found to be 2.66%, 96.9%, and 5, respectively. A mineral analysis of newly developed biopolymers showed a substantial amount of calcium (412 mg/100 g), potassium (418 mg/100 g), phosphorus (60 mg/100 g), iron (47 mg/100 g), zinc (16 mg/100 g), and sodium (9 mg/100 g). The significant antidiabetic potential of OMB was demonstrated using α-amylase and α-glucosidase enzyme inhibitory assay. Further investigations are required to explore the newly developed biopolymer for its toxicity, efficacy, and its possible utilization in food, nutraceutical, as well as pharmaceutical industries.

Solvents, CO2 and biopolymers: Structure formation in chitosan aerogel.

The functionality of biopolymer aerogels is inherently linked to its microstructure, which in turn depends on the synthesis protocol. Detailed investigations on the macroscopic size change and nanostructure formation during chitosan aerogel synthesis reveal a new aspect of biopolymer aerogels that increases process flexibility. Formaldehyde-cross-linked chitosan gels retain a significant fraction of their original volume after solvent exchange into methanol (50.3 %), ethanol (47.1 %) or isopropanol (26.7 %), but shrink dramatically during subsequent supercritical CO2 processing (down to 4.9 %, 3.5 % and 3.7 %, respectively). In contrast, chitosan gels shrink more strongly upon exchange into n-heptane (7.2 %), a low affinity solvent, and retain this volume during CO2 processing. Small-angle X-ray scattering confirms that the occurrence of the volumetric changes correlates with mesoporous network formation through physical coagulation in CO2 or n-heptane. The structure formation step can be controlled by solvent-polymer and polymer-drying interactions, which would be a new tool to tailor the aerogel structure.

Analysis of Plant Cell Walls Using High-Throughput Profiling Techniques with Multivariate Methods.

Plant cell walls are composed of a number of coextensive polysaccharide-rich networks (i.e., pectin, hemicellulose, protein). Polysaccharide-rich cell walls are important in a number of biological processes including fruit ripening, plant-pathogen interactions (e.g., pathogenic fungi), fermentations (e.g., winemaking), and tissue differentiation (e.g., secondary cell walls). Applying appropriate methods is necessary to assess biological roles as for example in putative plant gene functional characterization (e.g., experimental evaluation of transgenic plants). Obtaining datasets is relatively easy, using for example gas chromatography-mass spectrometry (GC-MS) methods for monosaccharide composition, Fourier transform infrared spectroscopy (FT-IR) and comprehensive microarray polymer profiling (CoMPP); however, analyzing the data requires implementing statistical tools for large-scale datasets. We have validated and implemented a range of multivariate data analysis methods on datasets from tobacco, grapevine, and wine polysaccharide studies. Here we present the workflow from processing samples to acquiring data to performing data analysis (particularly principal component analysis (PCA) and orthogonal projection to latent structure (OPLS) methods).

Design of Turn-On Near-Infrared Fluorescent Probes for Highly Sensitive and Selective Monitoring of Biopolymers.

Simple, sensitive, and selective detection of specific biopolymers is critical in a broad range of biomedical and technological areas. We present a design of turn-on near-infrared (NIR) fluorescent probes with intrinsically high signal-to-background ratio. The fluorescent signal generation mechanism is based on the aggregation/de-aggregation of phthalocyanine chromophores controlled by selective binding of small-molecule "anchor" groups to a specific binding site of a target biopolymer. As a proof-of-concept, we demonstrate a design of a sensor for EGFR tyrosine kinase-an important target in cancer research. The universality of the fluorescent signal generation mechanism, as well as the dependence of the response selectivity on the choice of the small-molecule "anchor" group, make it possible to use this approach to design reliable turn-on NIR fluorescent sensors for detecting specific protein targets present in the low-nanomolar concentration range.

Production and Characterization of Bioplastic by Polyhydroxybutyrate Accumulating Erythrobacter aquimaris Isolated from Mangrove Rhizosphere.

The synthesis of bioplastic from marine microbes has a great attendance in the realm of biotechnological applications for sustainable eco-management. This study aims to isolate novel strains of poly-ß-hydroxybutyrate (PHB)-producing bacteria from the mangrove rhizosphere, Red Sea, Saudi Arabia, and to characterize the extracted polymer. The efficient marine bacterial isolates were identified by the phylogenetic analysis of the 16S rRNA genes as Tamlana crocina, Bacillus aquimaris, Erythrobacter aquimaris, and Halomonas halophila. The optimization of PHB accumulation by E. aquimaris was achieved at 120 h, pH 8.0, 35 °C, and 2% NaCl, using glucose and peptone as the best carbon and nitrogen sources at a C:N ratio of 9.2:1. The characterization of the extracted biopolymer by Fourier-transform infrared spectroscopy (FTIR), Nuclear magnetic resonance (NMR), and Gas chromatography-mass spectrometry (GC-MS) proves the presence of hydroxyl, methyl, methylene, methine, and ester carbonyl groups, as well as derivative products of butanoic acid, that confirmed the structure of the polymer as PHB. This is the first report on E. aquimaris as a PHB producer, which promoted the hypothesis that marine rhizospheric bacteria were a new area of research for the production of biopolymers of commercial value.

Mainly Dimers and Trimers of Chinese Bayberry Leaves Proanthocyanidins (BLPs) are Utilized by Gut Microbiota: In Vitro Digestion and Fermentation Coupled with Caco-2 Transportation.

Chinese bayberry leaf proanthocyanidins (BLPs) are Epigallocatechin gallate (EGCG) oligomers or polymers, which have a lot of health-promoting activity. The activity is closely related to their behavior during in vitro digestion, which remains unknown and hinders further investigations. To clarify the changes of BLPs during gastrointestinal digestion, further research is required. For in vitro digestion, including gastric-intestinal digestion, colon fermentation was applied. Caco-2 monolayer transportation was also applied to investigate the behavior of different BLPs with different degrees of polymerization. The trimers and the tetramers were significantly decreased during in vitro gastric-intestinal digestion resulting in a significant increase in the content of dimers. The dimers and trimers were the main compounds utilized by gut microbiota and they were assumed not to degrade through cleavage of the inflavan bond. The monomers and dimers were able to transport through the Caco-2 monolayer at a rate of 10.45% and 6.4%, respectively.

Diseño innovador para la obtención y caracterización de un bioplástico utilizando como materia base la fibra de la cáscara de coco y papaya / Innovative design for obtaining and characterizing a bioplastic using as a base material the fiber of the coconut shell and papaya fiber

Los plásticos se producen a partir del petróleo. Estos polímeros perduran en la naturaleza por largos períodos de tiempo y, por tanto, se acumulan, generando así grandes cantidades de residuos sólidos. El objetivo principal de esta investigación es producir, a escala de laboratorio, un bioplástico a partir de la fibra contenida en la cáscara de coco, que pueda servir de materia prima para la elaboración de productos biodegradables.

Plastics are produced from oil. These polymers persist in nature for long periods of time and, therefore, they are accumulated in large amounts of solid waste. The main objective of this research is to produce, on a laboratory scale, a bioplastic from the fiber contained in the coconut shell, which can serve as raw material for the production of biodegradable products.

Understanding the physics of hydrocolloids interaction using rheological, thermodynamic and functional properties: A case study on xanthan gum-cress seed gum blend.

Some rheological, thermodynamic and functional properties of selected hydrocolloids (xanthan gum-cress seed gum (XG-CSG)) blends at different ratios (1-0, 3-1, 1-1, 1-3, 0-1) were characterized to understand physically the biopolymers interaction and networks. XG showed a greater rigidity (elastic modulus, G'LVE = 58.60 Pa), total structural strength (complex modulus, G*LVE = 70.69 Pa), yield stress (limiting value of stress, τL = 7.58 Pa), emulsion capacity (EC = 6.78%) and foam stability (FC = 18.92%) than CSG (G'LVE = 7.05 Pa, G*LVE = 8.53 Pa, τL = 1.44 Pa, EC = 86.48% and FC = 14.98%), respectively. Among blends, 3-1 XG-CSG showed the highest G*LVE, foaming stability (FS) and the extent of recovery (Rr%). The results were summarized using the clustering technique and principal component analyses. The coefficient of the interaction of some parameters, Cole-Cole plots and Gibbs free energy changes (ΔG) of predisturbed and intact networks were investigated. In samples with an intact network, greater compatibility directly related to the extent of synergistic interaction, while in predisturbed samples, the lower compatibility directly related to the extent of synergistic interaction. Although all blends were highly incompatible with antagonistic behavior, 1-1 XG-CSG showed the lowest incompatibility (ΔG = 8028.60 J/mol) among samples with intact structure, while at disrupted state, 1-3 XG-CSG showed the lowest incompatibility (ΔG = 158.6 J/mol).

Analysis of Product Quality of Complex Polymeric IgM Produced by CHO Cells.

Immunoglobulin M (IgM) antibodies are considered as promising biopharmaceutical drugs in the future despite recombinant production is quite challenging as incomplete polymer formation or nucleic acid adherence can decrease the quality of the IgM preparation. Therefore, we defined densitometric and chromatographic methods as suitable tools to analyze the polymer distribution and the remaining nucleic acid content after initial IgM purification. Additionally, the quality of the glycosylation pattern is an important parameter for biological safety and efficacy.

Detailed characterization of Pinus ponderosa sporopollenin by infrared spectroscopy.

In plant spores and pollen, sporopollenin occurs as a structural polymer with remarkable resistance to chemical degradation. This recalcitrant polymer is well-suited to analysis by non-destructive infrared spectroscopy. However, existing infrared characterization of sporopollenin has been limited in scope and occasionally contradictory. This study provides a comprehensive structural analysis of sporopollenin in the Pinus ponderosa pollen exine using infrared spectroscopy, including detailed band assignments, descriptions of chemical reactivity, and comparison to multiple reference substances. We observe that the infrared spectral characteristics of sporopollenin prepared by enzymatic digestion of the polysaccharide-based intine are largely consistent with a copolymer of aliphatic lipids and trans-4-hydroxycinnamic acid, without distinct contributions from α-pyrone or carotenoid substructures.

Subacute feeding toxicity of low-sodium sausages manufactured with sodium substitutes and biopolymer-encapsulated saltwort (Salicornia herbacea) in a mouse model.

BACKGROUND: Low-sodium sausages were manufactured using sodium substitution and biopolymer encapsulation. A diet comprising 10% treatment sausages (six treatment groups: C (100% NaCl), T1 (55% sodium substitute + 45% saltwort salt), T2 (55% sodium substitute + 45% saltwort salt with chitosan), T3 (55% sodium substitute + 45% saltwort salt with cellulose), T4 (55% sodium substitute + 45% saltwort salt with dextrin), and T5 (55% sodium substitute + 45% saltwort salt with pectin)) was added to a 90% commercial mouse diet for 4 weeks. RESULTS: Subacute toxicity, hematology, liver function, and organ weight tests in low-sodium sausage groups showed results similar to those of the control group, and all toxicity test levels were within normal ranges. CONCLUSIONS: All low-sodium sausage types tested are suggested to be safe in terms of subacute toxicity. Moreover, low-sodium sausages can be manufactured by biopolymer encapsulation of saltwort using pectin, chitosan, cellulose, and dextrin without toxicity. © 2019 Society of Chemical Industry.

Chronic venous ulcers: a review on treatment with fibrin sealant and prognostic advances using proteomic strategies

Venous ulcers are the main causes of chronic lower-limb ulcers. The healing difficulties encourage the research and development of new products in order to achieve better therapeutic results. Fibrin sealant is one of these alternatives. Besides being a validated scaffold and drug delivery system, it possesses excellent healing properties. This review covered the last 25 years of the literature and showed that the fibrin sealant is used in various clinical situations to promote the healing of different types of ulcers, especially chronic ones. These are mostly venous in origin and usually does not respond to conventional treatment. Commercially, only the homologous fibrin sealants obtained from human blood are available, which are highly efficient but very expensive. The heterologous fibrin sealant is a non-commercial experimental low-cost product and easily produced due to the abundance of raw material. The phase I/II clinical trial is already completed and showed that the product is safe and promisingly efficacious for the treatment of chronic venous ulcers. In addition, clinical proteomic strategies to assess disease prognosis have been increasingly used. By analyzing liquid samples from the wounds through proteomic strategies, it is possible to predict before treatment which ulcers will evolve favorably and which ones will be difficult to heal. This prognosis is only possible by evaluating the expression of isolated proteins in exudates and analysis using label-free strategies for shotgun. Multicentric clinical trials will be required to evaluate the efficacy of fibrin sealant to treat chronic ulcers, as well as to validate the proteomic strategies to assess prognosis.(AU)