Obtaining and study of monosaccharide derivatives of low-molecular-weight chitosan

The possibility of obtaining monosaccharide derivatives of low-molecular-weight chitosan with the use of the Maillard reaction was studied. Chitosan derivatives (molecular weight, 24 and 5 kDa) obtained with glucosamine, N-acetyl galactosamine, galactose, and mannose with a substitution degree of 4–14% and a yield of 60–80% were obtained. Some physicochemical and biological properties of these derivatives were studied. We showed that monosaccharide derivatives of low-molecular-weight chitosan exhibited antibacterial activity. Chitosan at a concentration of 0.01% caused 100% death of bacteria B. subtilis and E. coli. The strongest antibacterial effect was exhibited by 24-kDa derivatives: only 0.02–0.08% of cells survived. These derivatives were two orders of magnitude more effective than the 5-kDa chitosan modified with galactose.     

<Emphasis Type="Italic">Camellia sinensis</Emphasis> increased apoptosis on U2OS osteosarcoma cells and wound healing potential on NIH3T3 fibroblast cells

Camellia sinensis (Cs) is a plant which is rich in polyphenols and has antioxidant, antiinflammatory, antimutagenic, anticarcinogenic and antibacterial activities. In this study, two different methanol extracts (Cs-I and Cs-II) were prepared from the leaf of C. sinensis in order to investigate the wound healing and anticancer activities. Total phenolic content and antioxidant activity of the extracts were determined. Wound healing effects of Cs extracts were evaluated by using Masson’s Trichrome Tecnique on NIH3T3 fibroblast cells. Cytotoxic and apoptotic effects of the extracts were determined by MTT and AnnexinV-PI assays on U2OS osteosarcoma cells. Total phenolic contents and antioxidant activities of the extracts were almost the same. The highest concentration (60 µg/mL) of the extracts showed significant cytotoxic and apoptotic effects on U2OS cells. Especially, the highest apoptotic effect was determined with 60 µg/mL Cs-I extract. Significant wound healing potential on NIH3T3 fibroblast cells were determined especially with low extract concentrations (0.5, 1 and 5 µg/mL), while high extract concentrations showed significant anticancer effects. As a result, two Cs leaf extracts exhibited important apoptotic properties and both have wound healing potential. However, the Cs-I extract was found more effective on apoptotic osteosarcoma cell death and has an increased wound healing potential than the Cs-II extract.     

Topical Niosome Gel Containing an Anthocyanin Complex: a Potential Oral Wound Healing in Rats

Anthocyanins from dietary sources showing potential benefits as anti-inflammatory in oral lesions were developed as an anthocyanin complex (AC), comprised of extracts of Zea mays (CC) and Clitoria ternatea (CT), and formulated into a niosome gel to prove its topical oral wound healing in vitro and in vivo investigations. The AC formed nano-sized clusters of crystalline-like aggregates, occurring through both intra- and inter-molecular interactions, resulting in delivery depots of anthocyanins, following encapsulation in niosomes and incorporation into a mucoadhesive gel. In vitro permeation of anthocyanins was improved by complexation and further enhanced by encapsulation in niosomes. Collagen production in human gingival fibroblasts was promoted by AC and AC niosomes, but not CC or CT. The in vivo wound healing properties of AC gel (1 and 10%), AC niosome gel (1 and 10%), fluocinolone acetonide gel, and placebo gel were investigated for incisional wounds in the buccal cavities of Wistar rats. AC gel and AC niosome gel both reduced wound sizes after 3 days. AC niosome gel (10%) gave the highest reduction in wound sizes after day 3 (compared to fluocinolone acetonide gel, p?<?0.05), and resulted in 100% wound healing by day 5. Histological observations of cross-sectioned wound tissues revealed the adverse effects of fluocinolone gel and wound healing potential of AC niosome gel. Topical application of AC niosome gel exhibited an anti-inflammatory effect and promoted oral wound closure in rats, possibly due to the improved mucosal permeability and presence of delivery depots of AC in the niosome gel.     

Combined land cover changes and habitat occupancy to understand corridor status of Laljhadi-Mohana wildlife corridor,Nepal

Corridor design is a centripetal conservation tool to facilitate movement between fragmented patches. Increases in anthropogenic activity have caused degradation in forest connectivity, influencing animal movement to a small degree. Laljhadi-Mohana wildlife corridor (LMWC), a corridor between Shuklaphanta National Park (Nepal) and Dudhwa National Park (India) created to be used by Panthera tigris and Elephas maximus in western Nepal, is under pressure of anthropogenic change. Using current knowledge, we analyzed land cover changes (LCC) of LMWC between 2002 and 2012. We used ERDAS IMAGINE 9.2 and Arc GIS 9.2 to process satellite images, and occupancy survey to assess status of corridor. We classified land cover into dense forest, sparse forest, cultivation, water bodies, grassland, expose surfaces, and sand bank as structural attributes of the corridor. Our analysis found dense forest area was reduced by 18.35% in a decade while cultivation and sparse forest increased by 10.15% and 8.89%, respectively. Illegal forest encroachment, resource extraction, grazing pressure, invasive species, and flood were major drivers of forest change. The null occupancy model estimated the highest detection probability of Elephas maximus (0.48 ± 0.08) and the lowest of Axis axis (0.20 ± 0.08). Incorporating site covariates improved occupancy estimates of Sus scrofa (0.82), Axis axis (0.76), Elephas maximus (0.76), Boselaphus tragocamelus (0.66), and Panthera pardus (0.55). Distance to cultivation was the most influential covariate, supported by the expansion of cultivated land in the corridor. LMWC is a functional wildlife corridor despite a decline in forest cover. This decline influenced the number and detection rates of large mammals, instigating crop raiding and conflict. Mitigation measures on LCC drivers, particularly forest encroachment, can improve the functional status of LMWC and raise detection rates of large mammals in future studies.     

In vivo imaging of epithelial wound healing in the cnidarian <Emphasis Type="Italic">Clytia hemisphaerica</Emphasis> demonstrates early evolution of purse string and cell crawling closure mechanisms

Background

All animals have mechanisms for healing damage to the epithelial sheets that cover the body and line internal cavities. Epithelial wounds heal either by cells crawling over the wound gap, by contraction of a super-cellular actin cable (“purse string”) that surrounds the wound, or some combination of the two mechanisms. Both cell crawling and purse string closure of epithelial wounds are widely observed across vertebrates and invertebrates, suggesting early evolution of these mechanisms. Cnidarians evolved ~600 million years ago and are considered a sister group to the Bilateria. They have been much studied for their tremendous regenerative potential, but epithelial wound healing has not been characterized in detail. Conserved elements of wound healing in bilaterians and cnidarians would suggest an evolutionary origin in a common ancestor. Here we test this idea by characterizing epithelial wound healing in live medusae of Clytia hemisphaerica.

Results

We identified cell crawling and purse string-mediated mechanisms of healing in Clytia epithelium that appear highly analogous of those seen in higher animals, suggesting that these mechanisms may have emerged in a common ancestor. Interestingly, we found that epithelial wound healing in Clytia is 75 to >600 times faster than in cultured cells or embryos of other animals previously studied, suggesting that Clytia may provide valuable clues about optimized healing efficiency. Finally, in Clytia, we show that damage to the basement membrane in a wound gap causes a rapid shift between the cell crawling and purse string mechanisms for wound closure. This is consistent with work in other systems showing that cells marginal to a wound choose between a super-cellular actin cable or lamellipodia formation to close wounds, and suggests a mechanism underlying this decision.

Conclusions

1. Cell crawling and purse string mechanisms of epithelial wound healing likely evolved before the divergence of Cnidaria from the bilaterian lineage ~ 600mya 2. In Clytia, the choice between cell crawling and purse string mechanisms of wound healing depends on interactions between the epithelial cells and the basement membrane.

     

Bacterial cellulose synthesized by <Emphasis Type="Italic">Gluconacetobacter hansenii</Emphasis> for medical applications

The properties of a polymer synthesized by the Gluconacetobacter hansenii strain GH-1/2008 were investigated. The studied bacterial cellulose polymer films are characterized by a mesh nanostructure composed of micro- and macrofibrils, a high water absorption capacity 556 ± 16.8%, and high strength and elasticity. Analysis of the spectrum recorded by ¹³С CP/MAS NMR spectroscopy showed that the bacterial cellulose synthesized by G. hansenii GH-1/2008 is a pure compound composed of Iα (65–70%) and Iβ (30–35%) allomorphs without any other impurities. It was found that the bacterial cellulose films with adsorbed antibiotics such as amoxiclav and fluconazole can be used as antibacterial and antifungal wound healing materials.     

A new 2-(4<Emphasis Type="Italic">H</Emphasis>-1,3-benzoxazin-4-on-2-yl)-1,3-tropolone: Synthesis,structure, and antibacterial properties

2-(4H-1,3-Benzoxazin-4-on-2-yl)-4,5,6-trichloro-1,3-tropolone, structural properties of which were studied using ¹H NMR, IR-spectroscopy, mass spectrometry, and quantum chemistry has been obtained for the first time using an acid-catalyzed condensation reaction of 3,4,5,6-tetrachloro-1,2-benzoquinone with 2-methyl-4H-3,1-benzoxazin-4-one. It has been shown that the new tropolone possesses antibacterial activity against hospital-acquired strains of gram-negative (Escherichia coli, Salmonella enterica sv. Enteritidis, Acinetobacter baumannii, and Pseudomonas aeruginosa) and Gram-positive (Staphylococcus aureus) bacteria. The obtained substance is suggested for development of a new antibacterial drug.     

Textural and Rheological Properties of Soy Protein Isolate Tofu-Type Emulsion Gels: Influence of Soybean Variety and Coagulant Type

The contribution of soybean variety and coagulant type to the textural and rheological properties of soy protein isolate (SPI) tofu-type emulsion gels was studied. SPIs from eight soybean varieties were subjected to amino acid and sodium dodecyl sulfate–polyacrylamide gel electrophoresis (SDS-PAGE) analysis, and results showed that the 11S fraction proteins (r?=?0.833, p?<?0.05) and the ratio of 11S to 7S (r =?0.920, p <?0.01) were positively correlated with the hardness of CaSO₄-induced emulsion gels and glucono-δ-lactone (GDL)-induced gels, with the correlation coefficients of 0.827 (p <?0.05) and 0.893 (p <?0.01), respectively. In the case of microbial transglutaminase (MTGase), strong relations between the content of glutamate (r =?0.886, p?<?0.01) and lysine (r =?0.810, p <?0.05) and gel hardness were found. Rheological data demonstrated that CaSO₄-induced emulsion gel was stiffer with high rigidity but gel induced by MTGase performed better elasticity. The findings of this study are of great importance to further understand the gelation mechanisms of different coagulants and provide useful information for the development of SPI-based filled tofu.     

Synthesis and antibacterial activity of some new benzo[5,6]chromeno[2,3-<Emphasis Type="Italic">d</Emphasis>]pyrimidines

The synthesis and antibacterial activity of some new benzo[5,6]chromeno[2,3-d]pyrimidine derivatives are described. The title compounds were obtained by the reaction of 1H-benzo[f]chromenes with aliphatic and aromatic amines. The structures of all newly synthesized compounds were confirmed by IR, ¹HNMR, ¹³C NMR, and NOESY experiments. The compounds exhibited potent antibacterial activity against gram-positive and gram-negative bacterial species. 10-Methyl-12-(4-hydroxyphenyl)-10,12-dihydro-11H-benzo[5,6]chromeno[2,3-d] pyrimidin-11-imine displayed greater antibacterial activity against gramnegative bacterial species than did ciprofloxacinandamoxicillin.     

Antibiotic activity of fungi isolated from soil samples from Indonesia

Two hundred and thirteen fungal cultures were recovered from 88 soil samples from different parts of Indonesia; 39.4% belonged to the genusPenicillium, 19.7% to the genusAspergillus, 9.9% to the genusFusarium and the rest to different systematic groups. One hundred and fifty two cultures were antibiotically active; 80% of these were antagonists ofBacillus subtilis, 55% ofEscherichia coli, 20% ofSaccharomyces cerevisiae and 37% ofCandida pseudotropicalis. In agreement with previous observations it was found that the activity spectrum of antagonists was related to the altitude above sea level at which they were found. As the altitude increased, the incidence of antagonists with both antibacterial and antifungal activity decreased, but the incidence of antagonists with only antibacterial or only antifungal activity increased. Fungi of the generaPenicillium andAspergillus were the most frequent antibiotic producers. The incidence of penicillin producers was much lower than in collections of fungi isolated in higher latitudes (China, Bulgaria, Slovakia).     

On the nature of the domination of oligomeric (dA:dT)<Subscript><Emphasis Type="Italic">n</Emphasis></Subscript> tracts in the structure of eukaryotic genomes

The comparative analysis of the frequency of oligomeric W- and S-tracts was conducted in the genomes of different eukaryotic species that differ in their GC-composition. The domination of mononucleotide and mixed (A/T) _n -sequences compared with (G/C) _n -sequences was detected in the studied eukaryotic genomes, including Dictyostelium discoideum (GC ~ 25.7%), Caenorhabditis elegans (GC ~ 36.9%), Arabidopsis thaliana (GC ~ 38.0%), Drosophila melanogaster (GC ~ 38.8%), Homo sapiens (GC ~ 40.0%), Gallus gallus (GC ~ 50.0%), Leishmania major (GC ~ 59.1%). Using the results of quantum–chemical calculations, a theoretical rationale of the important role of existing differences in the initial hidden structural polymorphism of the hydrogen bonding of the Watson–Crick AT and GC pairs in the development of this phenomenon is given. It is concluded that a decreased structural polymorphism of AT pairs compared with the high fourfold polymorphism of the main state of GC pairs may be the most probable source of the preference and reliability of the “use” of AT pairs by nature in genomic DNA of many species.     

Isolation of hydroquinone (benzene-1,4-diol) metabolite from halotolerant <Emphasis Type="Italic">Bacillus methylotrophicus</Emphasis> MHC10 and its inhibitory activity towards bacterial pathogens

A halotolerant bacterial isolate-MHC10 with broad spectrum antibacterial activity against clinical pathogens was isolated from saltpans located in Tuticorin and Chennai (India). 16S rRNA gene analysis of MHC10 revealed close similarity to that of Bacillus methylotrophicus. The culture conditions of B. methylotrophicus MHC10 strain were optimized for antibacterial production using different carbon and nitrogen sources, as well as varying temperature, pH, sodium chloride (NaCl) concentrations and incubation periods. The maximum antibacterial activity of B. methylotrophicus MHC10 was attained when ZMB was optimized with 1 % (w/v) glucose, 0.1 % (w/v) soybean meal which corresponded to a C/N ratio of 38.83, temperature at 37 °C, pH 7.0 and 8 % NaCl. The activity remained stable between 72 and 96 h and then drastically decreased after 96 h. Solvent extraction followed by chromatographic purification steps led to the isolation of hydroquinone (benzene-1,4-diol). The structure of the purified compound was elucidated based on FTIR, ¹H NMR, and ¹³C NMR spectroscopy. The compound exhibited efficient antibacterial activity against both Gram-positive and Gram-negative bacterial pathogens. The minimum inhibitory concentration (MIC) for Gram-positive pathogens ranged from 15.625 to 62.5 µg/mL^?1, while it was between 7.81 and 250 µg/mL^?1 for Gram-negative bacterial pathogens. This is the first report of hydroquinone produced by halotolerant B. methylotrophicus exhibiting promising antibacterial activity.     

Bacterial Strain <Emphasis Type="Italic">Alcaligenes denitrificans</Emphasis> C-32 Containing Two Nitrilases with Different Substrate Specificities

Two genes encoding nitrilases with different properties have been found in an Alcaligenes denitrificans C-32 strain with high nitrilase activity that is currently used as a biocatalyst for commercial ammonium acrylate production. Both genes were expressed in E. coli, and the properties of the recombinant nitrilases were studied. One of these genes, which is designated as nitC1, controlled the formation of nitrilase that preferred aliphatic nitriles (acrylonitrile and butyronitrile) as best substrates. The nucleotide sequence of the gene nitC1 was almost (99%) identical to the gene sequence of an aliphatic nitrilase from Acidovorax facilis 72W (DQ4444267). In turn, nitC2 had a high level of homology (85%) with the arylacetonitrilase gene from Alcaligenes faecalis JM3 (D13419). Benzyl cyanide was shown to be the best substrate for nitC2-encoded nitrilase. In light of the results of DNA homology and differences in substrate specificity, the NitC2 and NitC1 nitrilases from Alcaligenes denitrificans C-32 were allocated to the groups of aliphatic nitrilases and arylacetonitrilases, respectively.     

Efficacy of mechanically modified electrospun poly(<Emphasis Type="SmallCaps">l</Emphasis>-lactide-co-<Emphasis Type="Italic">ε</Emphasis>-caprolactone)/gelatin membrane on full-thickness wound healing in rats

Bioengineered skin substitute offers new opportunities for treating various skin ailments. To compensate the structural integrity problems of scaffolds prepared from natural components, we mechanically developed highly modified electrospun nanofibrous membranes, incorporating poly(l-lactide-co-ε-caprolactone) (PLCL) into gelatin [poly(l-lactide-co-ε-caprolactone)/ gelatin membrane, (P/G (3/7)]. Subsequent to our previous in vitro study, our goal was to evaluate the in vivo performance of PLCL, gelatin, and P/G (3/7) membranes, and investigate the feasibility of the newly developed P/G (3/7) membrane for wound healing. Histological analysis using the mathematical model of wound healing and contraction, revealed the association between stiffness of skin substitute with cytokeratin production and wound contraction rate, and the defect site covered with the stiffer membrane showed lower cytokeratin production, and inversely, higher wound contraction rate. Overall, the P/G (3/7) membrane induced a satisfactory wound healing outcome. However, lower cytokeratin production rate with the mechanically modified P/G membrane involves the importance of the conditional blending of PLCL. Conversely, the condition of PLCL showed some incompatibility and hindrance of skin regeneration, consistent with previous in vitro results. With proper mechanical strength and cell viability, the P/G (3/7) membrane could successfully be used as a suitable skin substitute scaffold.     

In Vitro Selection and Identification of Potential Probiotics Isolated from the Gastrointestinal Tract of Nile Tilapia, <Emphasis Type="Italic">Oreochromis niloticus</Emphasis>

Fish gut bacteria can be used as probiotics for aquaculture. The aim of this study is to screen and identify beneficial probiotic bacteria from the gut of Nile tilapia, Oreochromis niloticus. Nine out of one hundred thirty-five isolates were non-pathogenic through intraperitoneal injection and had antibacterial activities with at least a strain from the five isolated fish pathogens, Aeromonas sobria, Aeromonas hydrophila, Pseudomonas aeruginosa, Pseudomonas putida, and Staphylococcus aureus. Further tests showed that such isolates can survive in the presence of high bile concentration (10%) and at different acidic pH values. A strains (14HT) was sensitive to all selected antibiotics, two strains were (9HT and 11HT) resistant to streptomycin and three strains (9HT, 11HT and 38HT) had resistance to two antibiotics. Four isolates (11HT, 33HT, 38HT and 41HT) had an amylase and a protease activities and one strain (47HT) showed only amylase activity. Based on 16S rRNA gene analysis, the isolated strains were identified as follows: Lactococcus lactis (8HT, 9HT, 11HT and 33HT); Enterococcus faecalis (14HT), Lysinibacillus sp. (38HT) and Citrobacter freundii (39HT, 41HT and 47HT).     

Boric Acid Inhibition of <Emphasis Type="Italic">Trichophyton rubrum</Emphasis> Growth and Conidia Formation

Trichophyton rubrum is a common human dermatophyte that is the causative agent of 80–93% of fungal infections of the skin and nails. While dermatophyte infections in healthy people are easily treatable with over-the-counter medications, such infections pose a higher risk for patients with compromised immune function and impaired regenerative potential. The efficacy of boric acid (BA) for the treatment of vaginal yeast infections prompted an investigation of the effect of BA on growth and morphology of T. rubrum. This is of particular interest since BA facilitates wound healing, raising the possibility that treating athlete’s foot with BA, either alone or in combination with other antifungal drugs, would combine the benefits of antimicrobial activity and tissue regeneration to accelerate healing of infected skin. The data presented here show that BA represses T. rubrum growth at a concentration reported to be beneficial for host tissue regeneration. Oxygen exposure increases BA toxicity, and mycelia growing under BA stress avoid colonizing the surface of the growth surface, which leads to a suppression of aerial mycelium growth and surface conidia formation. BA penetrates into solid agar matrices, but the relative lack of oxygen below the substrate surface limits the effectiveness of BA in suppressing growth of embedded T. rubrum cells.     

A New Broad-Spectrum Peptide Antibiotic Produced by <Emphasis Type="Italic">Bacillus brevis</Emphasis> Strain MH9 Isolated from Margalla Hills of Islamabad,Pakistan

The antimicrobial peptide from a bacterial strain is isolated from soil sample of Margalla Hills of Islamabad, Pakistan. The peptide is found to significantly inhibit the growth of both Gram-positive (Staphylococcus aureus ATCC 6538 and Micrococcus luteus ATCC 10240) and Gram-negative (Escherichia coli ATCC 25922 and Salmonella typhi ATCC 14028) bacteria as compared to gramicidin as standard. The bacterium is identified as Bacillus brevis strain MH9 based on phenotype and phylogenetic analysis. The antibacterial polypeptide was produced optimally at 35 °C after 48 h of growth, precipitated by 50 % ammonium sulphate, and further purified using HPLC. The sequential steps of purification decrease the peptide contents with prominent antibacterial activity. The peptide composed of 11 amino acid was further characterized by FT-IR and NMR. Results suggested that the peptide molecule is a novel antibacterial agent that is effective against both Gram-positive and Gram-negative bacteria. This study may have important implications for new peptide antibiotic that could be a new addition to treat infections.     

Nano-silver-incorporated biomimetic polydopamine coating on a thermoplastic polyurethane porous nanocomposite as an efficient antibacterial wound dressing

Background

Developing an ideal wound dressing that meets the multiple demands of good biocompatibility, an appropriate porous structure, superior mechanical property and excellent antibacterial activity against drug-resistant bacteria is highly desirable for clinical wound care. Biocompatible thermoplastic polyurethane (TPU) membranes are promising candidates as a scaffold; however, their lack of a suitable porous structure and antibacterial activity has limited their application. Antibiotics are generally used for preventing bacterial infections, but the global emergence of drug-resistant bacteria continues to cause social concerns.

Results

Consequently, we prepared a flexible dressing based on a TPU membrane with a specific porous structure and then modified it with a biomimetic polydopamine coating to prepare in situ a nano-silver (NS)-based composite via a facile and eco-friendly approach. SEM images showed that the TPU/NS membranes were characterized by an ideal porous structure (pore size: ~?85 μm, porosity: ~?65%) that was decorated with nano-silver particles. ATR-FITR and XRD spectroscopy further confirmed the stepwise deposition of polydopamine and nano-silver. Water contact angle measurement indicated improved surface hydrophilicity after coating with polydopamine. Tensile testing demonstrated that the TPU/NS membranes had an acceptable mechanical strength and excellent flexibility. Subsequently, bacterial suspension assay, plate counting methods and Live/Dead staining assays demonstrated that the optimized TPU/NS2.5 membranes possessed excellent antibacterial activity against P. aeruginosa, E. coli, S. aureus and MRSA bacteria, while CCK8 testing, SEM observations and cell apoptosis assays demonstrated that they had no measurable cytotoxicity toward mammalian cells. Moreover, a steady and safe silver-releasing profile recorded by ICP-MS confirmed these results. Finally, by using a bacteria-infected (MRSA or P. aeruginosa) murine wound model, we found that TPU/NS2.5 membranes could prevent in vivo bacterial infections and promote wound healing via accelerating the re-epithelialization process, and these membranes had no obvious toxicity toward normal tissues.

Conclusion

Based on these results, the TPU/NS2.5 nanocomposite has great potential for the management of wounds, particularly for wounds caused by drug-resistant bacteria.