Alysson L. Angelim

Sulfated chitosan as tear substitute with no antimicrobial activity.

Chitosan of high molar mass and with 82% deacetylation was sulfated using two procedures and characterized. In the first method sample chitosan-S1 was produced using chlorosulfonic acid as the sulfating agent and N,N-dimethylformamide as the medium, and in the second method (chitosan-S2) formic acid was also used. The degrees of sulfation were 0.87 (chitosan-S1) and 0.67 (chitosan-S2). FTIR spectra showed bands at 1230, 800 and 580 cm(-1), attributed to sulfation. Moisture content followed the order: chitosan-S-0.87>chitosan-S-0.67>chitosan. Chain depolymerization was verified by GPC. Aqueous solutions showed pseudoplastic behavior and the viscosity at a concentration of 0.3% (w/v) was higher than that of healthy human tears (close to 3 mPas at shear rate 130 s(-1)). Substitutions in the C2NH and in C6OH groups were verified by NMR. Antimicrobial activity against Staphylococcus aureus and Pseudomonas aeruginosa was not observed. Considering that chitosan-S-0.67 had a higher solubility, less chain depolymerization, higher yield and better thermal stability in comparison with chitosan-S-0.87, the derivative with DS 0.67 offered the greatest potential for use in formulations of tear substitutes.

An innovative bioremediation strategy using a bacterial consortium entrapped in chitosan beads

This aim of this work was to develop a bioremediation strategy for oil-contaminated mangrove sediments using chitosan beads containing an immobilised hydrocarbonoclastic bacterial consortium. The consortium composed of 17 isolates was obtained from an enrichment culture. The isolates were identified by 16S rDNA sequencing, which revealed 12 different genera. Thirteen isolates showed resistance to chitosan and were thus able to be trapped in chitosan beads for microcosm evaluation. The data revealed that entrapped consortium grew in the microcosms until day 15, which is when the beads disintegrated and released their biomass into the sediments. Bacterial bioaugmentation within the sediments was confirmed by cell counts; additionally, the dynamics of the bacterial populations were analysed through denaturing gradient gel electrophoresis. The chitosan showed a prebiotic effect on the autochthonous bacterial communities. Therefore, chitosan beads containing selected immobilised bacteria attain two bioremediation purposes, bioaugmentation and biostimulation, and thus represent an emergent approach.

Cytotoxic activity of fungal strains isolated from the ascidian Eudistoma vannamei.

The cytotoxic activity at 50 μg/ml of extracts obtained from eleven fungal strains associated to Eudistoma vannamei, an endemic ascidian from Northeast Brazil, against two cell lines, i.e., the HCT‐8 (colon cancer) and the MDA‐MB‐435 (melanoma) cell lines, was investigated. The most promising extract (EV10) was obtained from a fungus identified as Aspergillus sp. by molecular analysis and was selected for bioassay‐guided isolation of its active principals. Large‐scale fermentation of EV10 in potato‐dextrose broth followed by chromatographic purification of the active extract from the liquid medium allowed the isolation of the isocoumarins mellein, cis‐4‐hydroxymellein, and trans‐4‐hydroxymellein, besides penicillic acid. All isolated compounds were tested for their cytotoxicity against the tumor cell lines MDA‐MB‐435 and HCT‐8 and revealed penicillic acid as the only cytotoxic compound (cell growth inhibitions >95%).

Culturable populations of Acinetobacter can promptly respond to contamination by alkanes in mangrove sediments

This study evaluated the potential of bacterial isolates from mangrove sediments to degrade hexadecane, an paraffin hydrocarbon that is a large constituent of diesel and automobile lubricants. From a total of 18 oil-degrading isolates obtained by an enrichment technique, four isolates showed a great potential to degrade hexadecane. The strain MSIC01, which was identified by 16S rRNA gene sequencing as Acinetobacter sp., showed the best performance in degrading this hydrocarbon, being capable of completely degrading 1% (v/v) hexadecane within 48 h without releasing biosurfactants. Its hydrophobic surface probably justifies its potential to degrade high concentrations of hexadecane. Thus, the sediments from the studied mangrove harbour bacterial communities that are able to use oil as a carbon source, which is a particularly interesting feature due to the risk of oil spills in coastal areas. Moreover, Acinetobacter sp. MSIC01 emerged as a promising candidate for applications in bioremediation of contaminated mangrove sediments.

Bioaugmentation strategy employing a microbial consortium immobilized in chitosan beads for oil degradation in mesocosm scale.

A bacterial consortium composed by four metagenomic clones and Bacillus subtilis strain CBMAI 707, all derived from petroleum reservoirs, was entrapped in chitosan beads and evaluated regarding hydrocarbon degradation capability. Experiments were carried out in mesocosm scale (3000L) with seawater artificially polluted with crude oil. At different time intervals, mesocosms were sampled and subjected to GC-FID and microbiological analyses, as total and heterotrophic culturable bacterial abundance (DAPI and CFU count), biological oxygen demand (BOD) and taxonomic diversity (massive sequencing of 16S rRNA genes). The results obtained showed that degradation of n-alkane hydrocarbons was similar between both treatments. However, aromatic compound degradation was more efficient in bioaugmentation treatment, with biodegradation percentages reaching up to 99% in 30days. Community dynamics was different between treatments and the consortium used in the bioaugmentation treatment contributed to a significant increase in aromatic hydrocarbon degradation.

Studies on the secondary metabolites of a Pseudoalteromonas sp. isolated from sediments collected at the northeastern coast of Brazil.

Continuing search for anticancer compounds from the marine environment, we have studied microorganisms that inhabit intertidal sediments of the northeastern Brazilian coast. Of the 32 strains isolated, 13 were selected for biological evaluation of their crude extracts. The acetate extract obtained from a Gram‐negative bacterium was strongly active against cancer cell lines with IC50 values that ranged from 0.04 (HL60 leukemia cells) to 0.26 μg/ml (MDA MB‐435 melanoma cells). The bacterium was identified as a Pseudoalteromonas sp. based on 16S rRNA gene sequencing. A bioassay‐guided fractionation of the active extract led to the isolation of prodigiosin, a well‐known tripyrrole red pigment with immunosuppressive and anticancer activities. Further experiments with ErbB‐2 overexpressing cell lines, including HB4a‐C3.6 (moderate overexpression), HB4a‐C5.2 (high overexpression), and the parental HB4a cell line, were performed. Prodigiosin was moderately active toward HB4a cells with an IC50 of 4.6 μg/ml, while it was 115 and 18 times more active toward HB4a‐C3.6 cells (IC50 of 0.04 μg/ml) and HB4a‐C5.2 (IC50 of 0.26 μg/ml) cells, respectively. These data suggest that, in spite of its previously described apoptosis‐inducing properties, prodigiosin can selectively recognize cells overexpressing ErbB‐2, which could be highly appealing in human breast cancer therapy.

Microbial epibionts of the colonial ascidians Didemnum galacteum and Cystodytes sp.

Symbiosis with microorganisms has been well documented for many marine invertebrate taxa. However, knowledge of the diversity of microorganisms associated with ascidians is still limited. This study assessed the microbial epibionts of Didemnum galacteum and Cystodytes sp., two ascidian species collected from the western coast of Ceará state (Brazil), at Dois Coqueiros beach and the port of Pecém, respectively. The microbiota were examined using optical microscopy, followed by subsequent analysis of fingerprinting profiles obtained by denaturing gradient gel electrophoresis (DGGE) and 16S rRNA clone libraries. The microscopy analysis showed for both ascidians a community comprising cyanobacteria, mainly Prochloron-like species, and diatoms. The DGGE results indicated that D. galacteum hosts a more diverse microbiota when compared to Cystodytes sp. The same analysis also suggested that the diversity of the seawater microbiota was higher than the diversity of the ascidian-associated microbiota. The analysis of the 16S rRNA clone libraries revealed the dominance of Proteobacteria symbionts associated with both ascidians, with Alphaproteobacteria as the major component in D. galacteum and Gammaproteobacteria the major component in Cystodytes sp. The analysis of the clone libraries also revealed the presence of other taxa such as Bacteroidetes, Planctomycetes, Actinobacteria, Cyanobacteria, and uncultured bacteria in D. galacteum, but not in Cystodytes sp. Among the bacteria found to be exclusively associated with the ascidians, none were shared by the two studied hosts. The combined results point to a diverse microbiota associated with the external surface of the ascidians, with a mixed composition including organisms typically found in the surrounding seawater, but also a more specific set of taxa.

Qualidade da carne e desempenho de suínos alimentados com dietas contendo farinha de silagem de pescado

The objective of this work was to determine the nutrient digestibility and metabolizable energy (ME) of fish silage, as well as to evaluate the effect of the dietary inclusion of fish silage meal (FSM) in diets on the performance, carcass characteristics, meat quality, sensory analysis of meat and mortadella, and economic viability of growing and finishing pigs. In the digestibility assay, 16 barrows (33.20±4.93 kg) received diets with and without FSM. The fish silage had 39.01% crude protein and 4,032 kcal kg-1 ME. In the performance assay, 32 barrows (26.00±1.68 kg) were fed diets containing different inclusion levels of FSM (0, 25, 50, and 75%). FSM, obtained from the mixture (1:1) of fish silage with corn, showed a quadratic effect on average daily gain, and the best result was obtained with the inclusion level of 25.83%. The results for feed conversion and economic viability indicate that up to 25% FSM, corresponding to 5.87% of fish silage based on dry matter, can be used in the pig growing and finishing phases.