Morphological study of electrospun chitosan/poly(vinyl alcohol)/glycerol nanofibres for skin care applications.

This study aimed to evaluate the influence of formulation and procedure parameters in obtaining thick and continuous chitosan/PVA/glycerol nanofibres to be applied in skin care. For that, the polymers were characterized by nuclear magnetic resonance, Fourier-transform infrared spectroscopy, and size-exclusion chromatography. After this, 96 chitosan/PVA/glycerol nanofibre scaffolds were prepared by electrospinning method, using factorial designs. The independent variables were crude and pure chitosan, 2 brands of PVA, 2 needle gauges, high and low polymer concentration, high and low glycerol concentration, and final solution with and without ultrafiltration. Morphological analysis was performed by scanning electron microscopy, atomic force microscopy, and confocal microscopy. The best sample (NF67) presented an average thickness of 268.3 nm, uniform distribution, and high yield. It was obtained at a 1:3.5 (crude chitosan: PVA with lower molecular weight, but more hydrolysed) ratio and lower glycerol concentration, suggesting that the degree of hydrolysis of the PVA is more important than its molecular weight for obtaining better quality nanofibres and that the glycerol also makes the electrospinning process difficult. Thus, it was possible to choose parameters that provide scaffolds that could be applied as a matrix extracellular-like material in wound healing.

Chilean crab (Aegla cholchol) as a new source of chitin and chitosan with antifungal properties against Candida spp.

In the present work, the chemical composition of the Chilean freshwater crab Aegla cholchol exoskeleton was studied for the first time. α-Chitin was isolated from three main body parts (pincers, legs, carapace), and its content ranged from 9.0-10.4% (w/w). Moreover, chitosan was extracted by alkaline treatment at different temperature and time regimes. Aegla cholchol exoskeleton, chitin and chitosans were characterized by FTIR, TGA, and SEM. Chemical structure of chitin and chitosan was confirmed by NMR spectroscopy. Chitosan molecular weight determinations were carried out by GPC. The obtained chitosan samples had a degree of N-acetylation (DA) between 4 and 15% and molecular weight (Mw) in the range of 65-201 kDa. The antifungal activity of the chitosan samples and the chitooligomer were tested toward twenty isolated clinical strains of Candida yeast. Chitosan with lower DA (4%) and higher molecular weight showed the lowest minimum inhibitory concentration (MIC) values, which was evidenced by the highest antifungal activity toward Candida albicans, Candida tropicalis, Candida glabrata, and Candida parapsilosis. Results suggest that Aegla cholchol is an excellent natural source for production of bioactive materials with potential applications in the health system, to prevent infections associated with Candida strains.

Application of biosurfactants and chitosan in toothpaste formulation.

The aim of the present study was to formulate toothpastes containing biosurfactants and either fungal chitosan or sodium fluoride and evaluate the cytotoxicity, antimicrobial action and inhibition potential against biofilm formed by Streptococcus mutans. Chitosan was extracted from the biomass of the fungus Mucorales. We tested biosurfactants produced by Pseudomonas aeruginosa UCP 0992 (PB), Bacillus metylotrophicus UCP 1616 (BB) and Candida bombicola URM 3718 (CB). Fractional inhibitory concentration analysis was performed to determine the type of interaction between the compounds. Six toothpaste were prepared, the active ingredients of which were the biosurfactants, chitosan or sodium fluoride. The cytotoxicity tests were performed using the 3-[4,5-dimethyl-2-thiazolyl]-2,5-diphenyl-2-H-tetrazolium bromide (MTT) assay for the L929 (mouse fibroblast) and RAW 264.7 (mouse macrophage) cell lines. The toothpastes were tested with regard to pH, consistency and foaming capacity. The inhibition of biofilm was investigated by applying the toothpaste to biofilm formed in modified artificial saliva for 24 h at 37 °C in anaerobiosis. All substances had a minimum inhibitory concentration (MIC) for S. mutans. The combinations of CB and PB with chitosan had an additive effect against S. mutans, whereas BB combined with chitosan had an indifferent effect. The toothpastes were non-toxic. The formulations had pH around 9, spreading capacity between 8 and 17 mm and foaming capacity between 63 and 95%. All formulations inhibited the cellular viability of S. mutans in the biofilm, with similar results compared to the commercial toothpaste tested. The present results show that the formulations suggested are promising when compared to a commercial tooth paste.

Chitosan oligosaccharide (COS): An overview.

The frequently studied polysaccharide, chitosan oligosaccharide/chitooligosaccharide (COS) is the major degradation product of chitosan/chitin via chemical hydrolysis or enzymatic degradation involving deacetylation and depolymerization processes. Innumerable studies have revealed in the recent decade that COS has various promising biomedical implications in the past analysis, current developments and potential applications in a biomedical, pharmaceutical and agricultural sector. Innovations into COS derivatization has broadened its application in cosmeceutical and nutraceutical productions as well as in water treatment and environmental safety. In relation to its parent biomaterials and other available polysaccharides, COS has low molecular weight (Mw), higher degree of deacetylation (DD), higher degree of polymerization (DP), less viscous and complete water solubility, which endowed it with significant biological properties like antimicrobial, antioxidant, anti-inflammatory and antihypertensive, as well as drug/DNA delivery ability. In addition, it is also revealed to exhibit antidiabetic, anti-obesity, anti-HIV-1, anti-Alzheimer's disease, hypocholesterolemic, calcium absorption and hemostatic effects. Furthermore, COS is shown to have higher cellular transduction and completely absorbable via intestinal epithelium due to its cationic sphere exposed on the more exposed shorter N-glucosamine (N-Glc) units. This paper narrates the recent developments in COS biomedical applications while paying considerable attention to its physicochemical properties and its chemical composition. Its pharmacokinetic aspects are also briefly discussed while highlighting potential overdose or lethal dosing. In addition, due to its multiple NGlc unit composition and vulnerability to degradation, its safety is given significant attention. Finally, a suggestion is made for extensive study on COS anti-HIV effects with well-refined batches.

Otimização e padronização de processos para obtenção de quitosana purificada para uso farmacêutico e alimentício / Optimization and standardization of processes to obtain purified chitosan for pharmaceutical and food use

Quitosana é um biopolímero encontrado principalmente na parede celular de crustáceos e é obtida pela desacetilação da quitina. Como biopolímero a quitosana é utilizada como excipiente para medicamentos e composição de alimentos. No entanto a quitosana devidamente purificada para uso farmacêutico ou alimentício tem custo financeiro elevado. Outro fator que contribui para o uso limitado é a falta de procedimento padronizado para desacetilação, o que resulta em materiais com diferentes graus de qualidade, dificultando suas aplicações e controle de qualidade de matéria prima e produto. Este trabalho tem como principal objetivo estabelecer procedimento reprodutível para a extração da quitina e da quitosana, por meio da aplicação dos conceitos de Quality by Design e planejamento de experimentos. A quitosana foi obtida pela desacetilação da quitina de crustáceos pelas etapas de desmineralização, desproteinização e despigmentação. O procedimento técnico para purificação da quitosana foi definido a partir de planejamento fatorial com ponto central para as etapas otimizadas, por meio da aplicação dos conceitos de Quality by Design e planejamento de experimentos. O projeto definiu um procedimento padronizado para purificação da quitosana que pode ser empregado em escala industrial, e financeiramente vantajoso para produção de medicamentos ou alimentos

Chitosan is a biopolymer found mainly in the cell wall of crustaceans and is obtained by the deacetylation of chitin. As biopolymer chitosan is used as excipient for medicaments and food composition. However, chitosan duly purified for pharmaceutical or food use has a high financial cost. Another factor that contributes to the limited use is the lack of standardized procedure for deacetylation, which results in materials with different grades of quality, hindering their applications and quality control of raw material and product. This work has as main objective to establish a reproducible procedure for the extraction of chitin and chitosan, through the application of the concepts of Quality by Design and planning of experiments. Chitosan was obtained by the deacetylation of chitin from crustaceans through the demineralization, deproteinization and depigmentation stages. The technical procedure for purification of chitosan was defined from a factorial planning with a central point for the optimized steps, through the application of the concepts of Quality by Design and planning of experiments. The project defined a standard procedure for the purification of chitosan that can be used on industrial scale and financially advantageous for the production of medicines or foods

Synthesis and Application of Scaffolds of Chitosan-Graphene Oxide by the Freeze-Drying Method for Tissue Regeneration.

Several biomaterials, including natural polymers, are used to increase cellular interactions as an effective way to treat bone injuries. Chitosan (CS) is one of the most studied biocompatible natural polymers. Graphene oxide (GO) is a carbon-based nanomaterial capable of imparting desired properties to the scaffolds. In the present study, CS and GO were used for scaffold preparation. CS was extracted from the mycelium of the fungus Aspergillus niger. On the other hand, GO was synthesized using an improved Hummers-Offemann method and was characterized by Fourier transform infrared spectroscopy (FTIR), Raman spectroscopy, atomic force microscopy (AFM), X-ray diffraction (XRD), and dynamic light scattering (DLS). Subsequently, three formulations (GO 0%, 0.5%, and 1%) were used to prepare the scaffolds by the freeze-drying technique. The scaffolds were characterized by FTIR, thermogravimetric analysis (TGA), and scanning electron microscopy (SEM), to determine their thermal stability and pore size, demonstrating that their stability increased with the increase of GO amount. Finally, the scaffolds were implanted, recollected 30 days later, and studied with an optical microscope, which evidenced the recovery of the tissue architecture and excellent biocompatibility. Hence, these results strongly suggested the inherent nature of chitosan/graphene oxide (CS/GO) scaffolds for their application in bone tissue regeneration.

Lactoferrin, chitosan and Melaleuca alternifolia-natural products that show promise in candidiasis treatment

Abstract The evolution of microorganisms resistant to many medicines has become a major challenge for the scientific community around the world. Motivated by the gravity of such a situation, the World Health Organization released a report in 2014 with the aim of providing updated information on this critical scenario. Among the most worrying microorganisms, species from the genus Candida have exhibited a high rate of resistance to antifungal drugs. Therefore, the objective of this review is to show that the use of natural products (extracts or isolated biomolecules), along with conventional antifungal therapy, can be a very promising strategy to overcome microbial multiresistance. Some promising alternatives are essential oils of Melaleuca alternifolia (mainly composed of terpinen-4-ol, a type of monoterpene), lactoferrin (a peptide isolated from milk) and chitosan (a copolymer from chitin). Such products have great potential to increase antifungal therapy efficacy, mitigate side effects and provide a wide range of action in antifungal therapy.

Extraction of crude chitosans from squid (Illex argentinus) pen by a compressional puffing-pretreatment process and evaluation of their antibacterial activity.

Chitosan is produced by thermochemical alkaline deacetylation of chitin, but the process is usually environmentally problematic. In the present study, Illex argentinus squid pen chitin, after de-proteinization and demineralization, was pretreated with a compressional-puffing (CP) process under various puffing pressures. The CP process facilitated the increase of the crystalline index and degree of deacetylation of chitins. The CP-treated chitins were subjected to further extraction of chitosan, and four chitosan isolates (CI1-CI4) were obtained. The CP process was found to have beneficial effects in terms of increased extraction yield and increased antibacterial activity of the extracted chitosans. Moreover, the antibacterial property of the extracted chitosans seemed to be negatively related to their molecular weight (MW). Our findings showed that CI4 exhibited the highest extraction yield and the greatest antibacterial activity, and thus we recommend it as a safe and potent antibacterial agent for food, biomedicine, and other industrial usages.

Lactoferrin, chitosan and Melaleuca alternifolia-natural products that show promise in candidiasis treatment.

The evolution of microorganisms resistant to many medicines has become a major challenge for the scientific community around the world. Motivated by the gravity of such a situation, the World Health Organization released a report in 2014 with the aim of providing updated information on this critical scenario. Among the most worrying microorganisms, species from the genus Candida have exhibited a high rate of resistance to antifungal drugs. Therefore, the objective of this review is to show that the use of natural products (extracts or isolated biomolecules), along with conventional antifungal therapy, can be a very promising strategy to overcome microbial multiresistance. Some promising alternatives are essential oils of Melaleuca alternifolia (mainly composed of terpinen-4-ol, a type of monoterpene), lactoferrin (a peptide isolated from milk) and chitosan (a copolymer from chitin). Such products have great potential to increase antifungal therapy efficacy, mitigate side effects and provide a wide range of action in antifungal therapy.

Structure, morphology and properties of genipin-crosslinked carboxymethylchitosan porous membranes.

Highly porous genipin cross-linked membranes of carboxymethylchitosan exhibiting different crosslinking degree (3%

Chitin and chitosan from Brazilian Atlantic Coast: Isolation, characterization and antibacterial activity.

Chitin and chitosan were obtained by chemical treatments of shrimp shells. Different particle sizes (50-1000 µm) of the raw material were used to study their effect on size distribution, demineralization, deproteinization and deacetylation of chitin and chitosan isolation process. The particle size in the range of 800-1000 µm was selected to isolate chitin, which was achieved by measuring nitrogen, protein, ash, and yield %. Hydrochloric acid (5%, v/v) was optimized in demineralization step to remove the minerals from the starting material. Aqueous solution of sodium hydroxide (5%, w/v) at 90 °C for (20 h) was used in deproteinization step to remove the protein. Pure chitin was consequently impregnated into high concentration of sodium hydroxide (50%) for 3.5 h at 90 °C to remove the acetyl groups in order to form high pure chitosan. The degree of deacetylation (DDA) of chitosan was controlled and evaluated by different analytical tools. The chemical structure of chitin and chitosan was confirmed by elemental analysis, ATR-FTIR, H/C NMR, XRD, SEM, UV-Vis spectroscopy, TGA, and acid-base titration. The isolated chitin and chitosan from shrimp shell showed excellent antibacterial activity against Gram (-ve) bacteria (Escherichia coli) comparing with commercial biopolymers.

Isolation of chitosan from Ganoderma lucidum mushroom for biomedical applications.

Chitin biopolymer production and its by-product chitosan show great potential. These biomaterials have great applicability in various fields because they are non-toxic, biodegradable, biocompatible, and have antimicrobial effects. The most common source of chitin and chitosan is the crustaceous shell; however, mushrooms are an alternative source for isolating these biopolymers because their cellular wall has a high content of chitin, which may be transformed into chitosan through a deacetylation reaction. The main objective of this research was to obtain chitosan through the deacetylation of chitin isolated from the Ganoderma lucidum basidiomycetes mushroom, which is obtained through biotechnological culture. The material characterization was performed using X-ray diffraction, Fourier transform infrared spectroscopy, thermogravimetric analysis, and an evaluation of cytotoxicity comparing the results obtained with results for commercial chitosan. Protocol results showed that chitosan obtained from this mushroom had a significant similitude with commercial chitosan, yet the one obtained using P2 protocol was the one that rendered the best results: including diffractogram peaks, characteristic infrared analysis bands, and an 80.29 % degree of deacetylation. Cytotoxicity in vitro testing showed that the material was non-toxic; furthermore, it rendered very promising information regarding the evaluation of future applications of this biomaterial in the field of biomedicine.

Enhancement of Cunninghamella elegans UCP/WFCC 0542 biomass and chitosan with amino acid supply.

Studies were carried out with Cunninghamella elegans UCP/WFCC 0542 to evaluate the effects of an abundant supply of amino acids, asparagine and corn steep liquor associated with sucrose on the production of biomass and chitosan by submerged fermentation. The concentrations of the components of the culture medium which were determined by a 2³ full factorial design evaluated the interactions and effects of the independent variables of the sucrose, asparagine and corn steep liquor in relation to carbon and nitrogen sources, on the production of chitosan regarding biomass. The best results were observed at the central point [asparagine 0.025%, sucrose 0.15% and 0.45% of corn steep liquor, ratio C:N=2:6], and produced maximum yields of 16.95 g/L biomass and 2.14 g/L chitosan, after 96 h of submerged fermentation. However, the lowest level of sucrose, asparagine and corn steep liquor produced a low amount of biomass (10.83 g/L) and chitosan (0.60 g/L). The infrared spectrum absorption of the chitosan produced by C. elegans showed bands regarding OH-axial stretching between 3406 and 3432 cm⁻¹, superimposed on the NH stretching band with axial deformation of the amide C=O group at about 1639 cm⁻¹, NH angular deformation at approximately 1560 cm⁻¹; axial deformation of amide-CN at around 1421 cm⁻¹, symmetrical angular deformation in CH3 at 1379 cm⁻¹, -CN axial deformation of amino groups from 1125 to 1250 cm⁻¹ and polysaccharide structure bands in the range of between 890-1150 cm⁻¹. The crystallinity index of chitosan was 60.92%, and its degree of deacetylation was 75.25%. A low percentage of a supply of sucrose and asparagine with corn steep liquor offered higher yields of biomass and chitosan production at low cost.

Enhancement of chitosanase production by cell immobilization of Gongronella sp. JG

Chitosanase production of Gongronella sp. JG cells immobilized in calcium alginate gel and polyurethane foam was compared with that of the free cells, there was a 60% increase in the enzyme yield (2429 U/L) compared to the highest yield obtained from free cells (1513 U/L). The optimal immobilization parameters (concentrations of sodium alginate, calcium chloride, bead inoculums, bead diameter, etc) for the enhanced production of chitosanase were determined as: sodium alginate 2% (w/v), 0.1 M calcium chloride, inoculum 10 mL beads to 100 mL production media and 2.7 mm bead diameter. Maximum chitosanase production was achieved with initial pH of 5.5 and temperature of 30 ºC. The alginate beads had well stability, retained 85% ability of enzyme production even after 7 cycles of repeated batch fermentation. These results showed the immobilization technique was a feasible and economical method for chitosansase production by Gongronella sp. JG.

Effects of spray drying on physicochemical properties of chitosan acid salts.

The effects of spray-drying process and acidic solvent system on physicochemical properties of chitosan salts were investigated. Chitosan used in spray dryings was obtained by deacetylation of chitin from lobster (Panulirus argus) origin. The chitosan acid salts were prepared in a laboratory-scale spray drier, and organic acetic acid, lactic acid, and citric acid were used as solvents in the process. The physicochemical properties of chitosan salts were investigated by means of solid-state CP-MAS (13)C nuclear magnetic resonance (NMR), X-ray powder diffraction (XRPD), differential scanning calorimetry, and Fourier transform infrared spectrometry (FTIR) and near-infrared spectroscopy. The morphology of spray-dried chitosan acid salts showed tendency toward higher sphericity when higher temperatures in a spray-drying process were applied. Analysis by XRPD indicated that all chitosan acid salts studied were amorphous solids. Solid-state (13)C NMR spectra revealed the evidence of the partial conversion of chitosan acetate to chitin and also conversion to acetyl amide form which appears to be dependent on the spray-drying process. The FTIR spectra suggested that the organic acids applied in spray drying may interact with chitosan at the position of amino groups to form chitosan salts. With all three chitosan acid salts, the FTIR bands at 1,597 and 1,615 cm(-1) were diminished suggesting that -NH groups are protonated. The FTIR spectra of all chitosan acid salts exhibited ammonium and carboxylate bands at 1,630 and 1,556 cm(-1), respectively. In conclusion, spray drying is a potential method of preparing acid salts from chitosan obtained by deacetylation of chitin from lobster (P. argus) origin.

Zygomycetes from herbivore dung in the ecological reserve of Dois Irmãos, Northeast Brazil

Thirty-eight taxa of Zygomycetes distributed in 15 genera were recorded from tapir (Tapirus terrestris), camel (Camelus bactrianus), horse (Equus caballus), deer (Cervus elaphus), agouti (Dasyprocta aguti), donkey (Equus asinus), llama (Llama glama) and waterbuck (Kobus ellipsiprymnus) dung collected at the Reserva Ecológica de Dois Irmãos located in Recife, State of Pernambuco, Northeast Brazil. The samples were collected on a monthly basis from June 2005 to May 2006, taken to the laboratory and incubated in moist chambers. Higher number of taxa was observed in the excrements of tapir, followed by deer and donkey. The highest number of species was detected for Mucor, followed by Pilobolus. Statistical analyses showed significant differences in richness of Zygomycetes taxa between the herbivore dung types. Differences of species composition, however, were weak. Seasonality influenced the Zygomycetes species composition but not its richness. Variations in taxa composition between ruminants and non-ruminants dung were non significant.

Characterization and comparison of Serratia marcescens isolated from edible cactus and from silkworm for virulence potential and chitosan susceptibility

Representative strains of Serratia marcescens from an edible cactus plant and silkworms were characterized and a comparison based on their cellular fatty acid composition, 16S rRNA and groE gene sequence analysis as well as silkworm virulence and chitosan susceptibility was carried out. Results from this study indicate that there are no significant differences between the phenotypic and molecular characterization, virulence and chitosan susceptibility of the S. marcescens strains from the cactus plant and silkworms. Silkworms inoculated with S. marcescens from either plant or silkworm resulted in nearly 100 percent mortality. Chitosan solution exhibited strong antibacterial activity against S. marcescens. This activity increased with the increase of chitosan concentration and incubation time regardless of the strain source. Also, the results indicate that the plant associated S. marcescens maybe plays a possible role in the contamination of humans and animals, in particular silkworms, while chitosan showed a potential to control the contamination caused by S. marcescens.