Potential of Aspergillus oryzae as a biosynthetic platform for indigoidine, a non-ribosomal peptide pigment with antioxidant activity.

The growing demand for natural pigments in the industrial sector is a significant driving force in the development of production processes. The production of natural blue pigments, which have wide industrial applications, using microbial systems has been gaining significant attention. In this study, we used Aspergillus oryzae as a platform cell factory to produce the blue pigment indigoidine (InK), by genetic manipulation of its non-ribosomal peptide synthetase system to overexpress the indigoidine synthetase gene (AoinK). Phenotypic analysis showed that InK production from the engineered strain was growth associated, owing to the constitutive control of gene expression. Furthermore, the initial pH, temperature, and glutamine and MgSO4 concentrations were key factors affecting InK production by the engineered strain. The pigment secretion was enhanced by addition of 1% Tween 80 solution to the culture medium. The maximum titer of total InK was 1409.22 ± 95.33 mg/L, and the maximum productivity was 265.09 ± 14.74 mg/L·d. Moreover, the recombinant InK produced by the engineered strain exhibited antioxidant activity. These results indicate that A. oryzae has the potential to be used as a fungal platform for overproduction of extracellular non-ribosomal peptide pigments.

Biochemical characterization and synergism of cellulolytic enzyme system from Chaetomium globosum on rice straw saccharification.

BACKGROUND: Efficient hydrolysis of lignocellulosic materials to sugars for conversion to biofuels and chemicals is a key step in biorefinery. Designing an active saccharifying enzyme system with synergy among their components is considered a promising approach. RESULTS: In this study, a lignocellulose-degrading enzyme system of Chaetomium globosum BCC5776 (CG-Cel) was characterized for its activity and proteomic profiles, and synergism with accessory enzymes. The highest cellulase productivity of 0.40 FPU/mL was found for CG-Cel under the optimized submerged fermentation conditions on 1% (w/v) EPFB (empty palm fruit bunch), 2% microcrystalline cellulose (Avicel®) and 1% soybean meal (SBM) at 30 °C, pH 5.8 for 6 d. CG-Cel worked optimally at 50-60 °C in an acidic pH range. Proteomics analysis by LC/MS/MS revealed a complex enzyme system composed of core cellulases and accessory hydrolytic/non-hydrolytic enzymes attacking plant biopolymers. A synergistic enzyme system comprising the CG-Cel, a ß-glucosidase (Novozyme® 188) and a hemicellulase Accellerase® XY was optimized on saccharification of alkaline-pretreated rice straw by a mixture design approach. Applying a full cubic model, the optimal ratio of ternary enzyme mixture containing CG-Cel: Novozyme® 188: Accellerase® XY of 44.4:20.6:35.0 showed synergistic enhancement on reducing sugar yield with a glucose releasing efficiency of 256.4 mg/FPU, equivalent to a 2.9 times compared with that from CG-Cel alone. CONCLUSIONS: The work showed an approach for developing an active synergistic enzyme system based on the newly characterized C. globosum for lignocellulose saccharification and modification in bio-industries.

ß-Mannanase production by Aspergillus niger BCC4525 and its efficacy on broiler performance.

BACKGROUND: Mannan is a hemicellulose constituent commonly found in plant-derived feed ingredients. The gum-like property of mannan can obstruct digestive enzymes and bile acids, resulting in impaired nutrient utilisation. In this study, ß-mannanase production by Aspergillus niger strain BCC4525 was investigated using several agricultural residues under solid state condition. The biochemical properties of the target enzyme and the effects of enzyme supplementation on broiler performance and energy utilisation were assessed. RESULTS: Among five carbon sources tested, copra meal was found to be the best carbon source for ß-mannanase production with the maximum yield of 1837.5 U g(-1) . The crude ß-mannanase exhibited maximum activity at 80 °C within a broad range of pH from 2 to 6. In vitro digestibility assay, which simulates the gastrointestinal tract system of broilers, showed that ß-mannanase could liberate reducing sugars from corn/soybean diet. Surprisingly, ß-mannanase supplementation had no significant effect on broiler feed intake, feed conversion rate or energy utilisation. CONCLUSION: A high level of ß-mannanase was produced by A. niger BCC4525 under solid state condition using copra meal as carbon source. Although the enzyme has the desired properties of an enzyme additive for improving broiler performance, it does not appear to be beneficial.

Improvement of recombinant endoglucanase produced in Pichia pastoris KM71 through the use of synthetic medium for inoculum and pH control of proteolysis.

The long lag time in basal salts medium (BSM) and an occurrence of proteolysis are major problems for recombinant protein production in Pichia pastoris KM71. In this study, optimal conditions were explored for fed-batch cultivation of recombinant fungal endoglucanase in P. pastoris KM71. It was found that lag and process times were much reduced when the synthetic FM22 medium was used for the inoculum compared with enriched buffered glycerol complex (BMGY) medium. The highest endoglucanase activity was obtained at 30°C which was more than 10 fold higher than that produced from shake flask. At 30°C, the specific endoglucanase activity was dependent on culture pH and a higher specific activity was observed at pH 5.0 than at pH 6.0. The higher activity was likely due to lower rate of proteolysis, since a truncated protein species was apparent at pH 6.0, but not pH 5.0. Thus, production of endoglucanase at 30°C and pH 5.0 is the optimal condition suitable for economical production in large scale. The combination of using synthetic FM22 medium for inoculum and proteolysis control by growth at lower pH could be applied for production of other recombinant proteins in P. pastoris.

Production of multi-fiber modifying enzyme from Mamillisphaeria sp. for refining of recycled paper pulp.

Enzymatic modification of pulp is receiving increasing interest for energy reduction at the refining step of the paper-making process. In this study, the production of a multi-fiber modifying enzyme from Mamillisphaeria sp. BCC8893 was optimized in submerged fermentation using a response-surface methodology. Maximal production was obtained in a complex medium comprising wheat bran, soybean, and rice bran supplemented with yeast extract at pH 6.0 and a harvest time of 7 d, resulting in 9.2 IU/mL of carboxymethyl cellulase (CMCase), 14.9 IU/mL of filter paper activity (FPase), and 242.7 IU/mL of xylanase. Treatment of old corrugated container pulp at 0.2-0.3 IU of CMCase/g of pulp led to reductions in refining energy of 8.5-14.8%. The major physical properties were retained, including tensile and compression strength. Proteomic analysis showed that the enzyme was a complex composite of endo-glucanases, cellobiohydrolases, beta-1,4-xylanases, and beta-glucanases belonging to various glycosyl hydrolase families, suggestive of cooperative enzyme action in fiber modification, providing the basis for refining efficiency.

Optimization of xylanase production from Aspergillus niger for biobleaching of eucalyptus pulp.

A crude endo-xylanase produced by Aspergillus niger BCC14405 was investigated for its potential in pre-bleaching of chemical pulp from eucalyptus. The optimal fermentation conditions on the basis of optimization using response surface methodology included cultivation in a complex medium comprising wheat bran, rice bran, and soybean meal supplemented with yeast extract, glucose, peptone, and lactose with a starting pH of 6.0 for 7 d. This resulted in production of 89.5 IU/mL of xylanase with minor cellulase activity. Proteomic analysis using LC/MS/MS revealed that the crude enzyme was a composite of hemicellulolytic enzymes, including endo-ß-1,4-xylanase and other hemicellulolytic enzymes attacking arabinoxylan and mannan. Pretreatment of the pulp at a xylanase dosage of 10 IU/g increased the brightness ceiling after the C-Eop-H bleaching step up to 3.0% using a chlorine charge with a C-factor of 0.16-0.20. Xylanase treatment also led to reduction in chlorine charge of at least 20%, with an acceptable brightness level. The enzyme pretreatment resulted in a slight increase in pulp viscosity, suggesting an increase in relative cellulose content. The crude enzyme was potent in the enzyme-aided bleaching of chemical pulp in an environmentally friendly pulping process.

Menisporopsin B, a new polyester from the seed fungus Menisporopsis theobromae BCC 4162.

A new linear polyester, menisporopsin B, along with the known macrocyclic polyester, menisporopsin A, was isolated from the seed fungus Menisporopsis theobromae BCC 4162. The structure of menisposopsin B was addressed primarily by spectroscopic analyses, and the stereochemistry was established by chemical correlation. Menisporopsin B exhibited antimalarial activity with an IC(50) value of 1.0 microg/ml.