Proteomic Analysis of Intra- and Extracellular Proteins of Aspergillus niveus during Submerged Bioprocess Culturing under Different pH Conditions

Background: Proteomics facilitates understanding of the complexity of molecular and physiological mechanisms involved in the metabolic and biological fungal adaptations to pH changes. Proteomics enables the identification of enzymes and fungal proteins involved in these adaptations. This approach may be used to investigate such fungi as Aspergillus niveus, whose proteome has not yet been analyzed, changes the intra- and extracellular protein profiles in response to extracellular pH. Objective: In the current study, we used two-dimensional gel electrophoresis (2DE) and mass spectrometry to evaluate the response of A. niveus to grow at pH 5, 6, 7, and 8 for 96 hours submerged bioprocess culturing. Methods: This study evaluated the response of A. niveus to grow at pH 5, 6, 7, and 8 for 96 h submerged bioprocess culturing, by analysis of two-dimensional gel electrophoresis (2DE), of the intracellular proteomes and the secretome, protein spots of interest were submitted to tryptic digestion and analyzed by matrix-assisted laser desorption/ionization time-offlight tandem mass spectrometry (MALDI-TOF/TOF-MS). Results: This approach revealed substantial differences between the functions of intra- and extracellular proteins of A. niveus. The data suggested that pH-modulated global proteins are involved in important, mainly metabolic, processes, in the pentose phosphate pathway, protein regulation, cell wall maintenance, and others. Moreover, the change in extracellular pH could have altered the availability of nutrients, and induced the production of enzymes that respond to oxidative and other stresses. Conclusion: Proteomic facilitates understanding of the complexity of molecular and physiological mechanisms involved in the metabolic and biological adaptations of fungi to pH changes.

Production of L-asparaginase by Aspergillus niveus under solid-state fermentation using agroindustrial byproducts

<p class="abstract"><strong>Background:</strong> L-asparaginase, produced mainly by microrganisms, cleaves L-asparagine to aspartic acid and ammonia as products. This enzyme has been applied in the treatment of the leukemia and in food preparation preventing the acrylamide formation.</p><p class="abstract"><strong>Methods:</strong> <em>Aspergillus niveus</em> was grown in different solid substrates (agroindustrial byproducts) moistened with different agents (tap water, distilled water and several salt solutions) for different periods (24-240 h) at 30ºC. The enzyme extract was obtained with the addition of cold distilled water, agitation at 50 rpm for 30 min and filtration. The filtrate was used to determine the L-asparaginase activity through the hydroximate aspartic methodology using L-asparagine as substrate. The influence of temperature (30-75ºC), pH (3-9) and chemical compounds on the enzyme activity was analyzed. </p><p class="abstract"><strong>Results:</strong> The highest level of enzyme production was obtained using the M1 mixture (wheat bran, crushed soybean, orange peel; 1:1:1, w/w/w) as substrate humidified with Czapeck Dox salt solution (1:0.5, m/v) for 48-120 h, at 30ºC. The best temperature and pH for the enzyme activity were 35ºC and 5.0, respectively. The enzyme activity was increased in the presence of NaCl and some organic solvents (acetonitrile, butanol ethanol, isopropanol and methanol).</p><p class="abstract"><strong>Conclusions:</strong> <em>A. niveus</em> produced L-asparaginase under SSF using a mixture of agroindustrial byproducts as solid substrate in the absence of L-asparagine as inducer. The temperature and pH of activity, as well as the NaCl tolerance, indicate its potential to be applied for different purposes. <em>A. niveus</em> can be an interesting source of L-asparaginase gene to be investigated targeting future application.</p><p class="abstract"> </p>

Extraction and preliminary study of antibacterial compounds of three species of Aspergillus genus

In the interest of discovering new antibiotic molecules, the antibacterial activity of three fungal strains namely: Aspergillus quadrilineatus, Aspergillus niveus, and Aspergillus wentii isolated from particular ecosystems was sought against six bacterial strains including three with Gram-positive staining (Staphylococcus aureus, Bacillus subtilis, Enterococcus faecalis) and three with Gram-negative staining (Escherichia coli, Pseudomonas aeruginosa, Klebsiella pneumoniae). The results of the agar cylinder technique highlighted that the three fungal strains showed a considerable antibacterial activity. In order to optimize the extraction conditions of the bioactive molecules, five solvents in different polarities were tested, of which chloroform turned out to be the best one. After the selection of this solvent, four culture media of different compositions were used in order to determine the most adequate medium for the production of antibacterial substances. The results revealed that Czapek-dox medium supplemented with yeast extract turned out to be the most favorable one for the production of bioactive molecules from both strains: A. quadrilineatus and A. niveus, while the most suitable medium for the A. wentii strain was Sabouraud. In addition, a study of the antibacterial effect of organic extracts by the Biolog micro-culture system was performed using a range of concentrations. The obtained results revealed that the extracts of the three fungal strains presented a remarkable activity with different concentrations and this, against all the tested bacterial strains. It was recorded only for the three used fungal species, the antibacterial activity was studied for the first time by the Biolog system.

Extraction and preliminary study of antibacterial compounds of three species of Aspergillus genus

In the interest of discovering new antibiotic molecules, the antibacterial activity of three fungal strains namely: Aspergillus quadrilineatus, Aspergillus niveus, and Aspergillus wentii isolated from particular ecosystems was sought against six bacterial strains including three with Gram-positive staining (Staphylococcus aureus, Bacillus subtilis, Enterococcus faecalis) and three with Gram-negative staining (Escherichia coli, Pseudomonas aeruginosa, Klebsiella pneumoniae). The results of the agar cylinder technique highlighted that the three fungal strains showed a considerable antibacterial activity. In order to optimize the extraction conditions of the bioactive molecules, five solvents in different polarities were tested, of which chloroform turned out to be the best one. After the selection of this solvent, four culture media of different compositions were used in order to determine the most adequate medium for the production of antibacterial substances. The results revealed that Czapek-dox medium supplemented with yeast extract turned out to be the most favorable one for the production of bioactive molecules from both strains: A. quadrilineatus and A. niveus, while the most suitable medium for the A. wentii strain was Sabouraud. In addition, a study of the antibacterial effect of organic extracts by the Biolog micro-culture system was performed using a range of concentrations. The obtained results revealed that the extracts of the three fungal strains presented a remarkable activity with different concentrations and this, against all the tested bacterial strains. It was recorded only for the three used fungal species, the antibacterial activity was studied for the first time by the Biolog system.

Itaconic Acid Production by Novel Aspergillus Niveus in Solid State Fermentation Using Agrowastes

Itaconic acid (IA) is an organic compound. This dicarboxylicacid is a white solid that is soluble in water, ethanol, and acetone. Historically, itaconic acid was obtained by the distillation of citricacid, but currently it is produced by fermentation. Itaconic acid is essentially utilized as a co-monomer in the generation of polymers, for example, tar, plastic, elastic, paints, surfactant with applications in the paper and structural covering industry.. The fermentative production of Itaconic acid was probably achieved by the filamentous fungus Aspergillus terrus, Aspergillus itaconicus and mainly by Aspergillus niger. In the present work, an attempt was made to produce IA by Aspergillus niveus employing Solid State Fermentation (SSF) from various agro wastes like rice bran, rice husk,tamarind seed,wheat stuff and sugarcane bagasse as carbon substrates, which was pretreated in order to soften it.10 g of each substrate was taken in a 500 ml conical flasks separately and supplemented with 40 mL nutrient solution containing glucose, at pH 3.5. One milliliter inoculum containing 1×107 spores was added and moisture was maintained at 70%. After incubation at 32°C for 14 days, the acid production was estimated by spectrophotometric method and by HPLC analysis. Interestingly, the yield of itaconic acid was promising with all the above substrates, where tamarind seed, sugarcane bagasse and rice bran supported higher yields.