Amylase synthesis inAspergillus flavus andAspergillus niger grown on cassava peel

Summary Aspergillus flavus andAspergillus niger produce extracellular amylase into the culture medium when grown on basal medium containing 2% (w/v) soluble starch or cassava peel as the sole carbon source. On soluble tarch the highest amylase activities were 1.6 and 5.2 mg of starch hydrolyzed/min per mg protein forA. flavus andA. niger, respectively. When grown on cassava peel, the highest amylase activity in the culture filtrate ofA. flavus was 170-times higher than that on soluble starch, while that ofA. niger was 16-times higher. The mycelial dry weight for both organisms was not significantly affected by the carbon sources. Maximum enzyme activity was obtained at the growth temperature of 29.0±1°C and pH 7 for both organisms. It is concluded that cassava peel might be a better substrate for the production of amylase byA. flavus andA. niger than commercial soluble starch.     

The posibility of soil micromycetes produced the abscisic acid

The typical soil micromycetes Aspergillus niger and Cladosporium cladosporioides from the family moniliaceae were investigated with emphasis on production of ABA into the culture medium. The both fungi were cultivated in a static liquid Czapek — Dox medium and agar Czapek — Dox medium. Aspergillus niger and Cladosporium cladosporioides showed ability to produce ABA. Analytical detection of ABA from the culture medium was performed by TLC combinated with biotest and HPLC with spectroscopy.     

Comparative study of the production of extracellular β-glucosidase by four different strains of Aspergillus using submerged fermentation

Four strains of Aspergillus (Aspergillus niger CDBB-H-176, A. niger CDBB-H-175, A. niger ATCC 9642, and Aspergillus terreus CDBB-H-194) were used to produce extracellular β-glucosidase. Using an orthogonal experimental design (L₉), we optimized the parameters of culture medium to maximize the activity of β-glucosidase. The optimal conditions (same for the four strains) were as follows: temperature, 30°C; pH, 6.0; orbital agitation, 200?rpm; concentration of sucrose, 0.5% (w/v). The most productive strain was A. niger CDBB-H-175, with a yield of 701.2?U/mL. In a second stage, we optimized (L₁₈) the concentration of nutrients in the culture medium to determine whether this modification would increase the production of β-glucosidase. The optimal conditions for A. niger CDBB-H-175 were as follows (%, w/v): NaNO₃, 0.3; KCl, 0.3; KH₂PO₄, 0.15; NH₄NO₃, 0.1; NH₄H₂PO₄, 0.1; MgSO₄?·?7H₂O, 0.05; yeast extract, 0.1. The production of β-glucosidase under these conditions was 1207.9?U/mL. Enzymatic assays were used to characterize the enzyme; the optimum temperature and pH of β-glucosidase produced by the four selected micro-organisms were found to be 65°C and 5.0, respectively. We determined the Michaelis–Menten constants (K_m) only for A. niger CDBB-H-175 and CDBB-H-176; the values were 2.7 and 2.2?mM, respectively.     

Effects of Octadecanoylsucrose Derivatives on the Production of Inulinase by Apergillus niger SL-09

Summary The effect of the addition of octadecanoylsucrose esters to the growth medium on the production of inulinase by Aspergillus niger SL-09 was studied in batch culture using shake flasks. The activities of inulinase in vitro and in vivo formed by Aspergillus niger SL-09 was enhanced dramatically by the addition of sucrose ester S-770 to the medium, and it was confirmed that sucrose ester acted as a very efficient inducer for inulinase production. As a result, with the addition of 6 g sucrose ester l⁻¹ at the beginning of the culture, the enzyme activities were enhanced near 7-fold higher than that obtained in the basal medium.     

Microbial biomass production by continuous fermentation of bark hydrolysate

Summary The fungus Aspergillus niger, an unidentified filamentous fungus (strain no. PDDCC8239) and the yeast Candida tropicalis were grown in continuous culture in stirred tank reactors at dilution retes varying between 0.02–0.1 h^-1 on a bark extract medium made by dilute acid hydrolysis of Pinus radiata bark. Maximum yields were 5.1, 18.7, and 61.5 mg biomass·g^-1 bark for the unidentified fungus, A. niger and C. tropicalis respectively. Culturing in a tower fermenter under otherwise identical environmental conditions increased the yield of A. niger to 27.3 mg biomass·g^-1 bark. The yield of C. tropicalis represents a productivity of 0.26g biomass·l^-1·h^-1 which exceeds other reported values of bark fermentations. Analysis of the medium and spent broth revealed significant breakdown of tannin material by C. tropicalis during growth. This ability may be of value in the treatment of tannin-based industrial effluents.     

Sr Isotope Tracing of Differences in the Effects of Microorganisms on Weathering of Calcite and Apatite

To study differences in the effects of microorganisms on weathering of calcite and apatite, one strain of Aspergillus niger (A. niger) and one strain of Penicillium glaucum (P. glaucum), which respectively contain the mixture of calcite and apatite were cultivated for 24 days in the sucrose-potato culture medium, supernatant was taken every three days from the culture medium, followed by the determination of Ca²⁺ and Sr²⁺ contents and Sr isotopic ratios. The results of measurement showed that the Sr isotope ratios in the supernatant from the culture medium are intermediate between those of the end-member constituents calcite and apatite (0.70721-0.70861). Results of isotope mixing equations to calculation showed that in the first 15 days A. niger played a dominant role in weathering of calcite in the apatite/calcite mixture. The contribution rate of apatite for Ca²⁺ in the solution increased from 39.0% on the 18th day to 61.6% on the 24th day; P. glaucum played a key role in weathering of apatite in the first 3 days. Ions dissolved from apatite account for 73.9% of the total. It is known from the results of Sr isotope tracing that in the prior period of fungus cultivation A. niger plays a key role in weathering of calcite while P. glaucum plays a key role in weathering of apatite. The ability of P. glaucum to weather calcite tends to intensify progressively over time. Therefore, Sr isotope tracing can be used to accurately recognize differences in the effects of microorganisms on weathering of minerals.     

Effects of different carbon sources on the synthesis of pectinase by Aspergillus niger in submerged and solid state fermentations

A study was made to compare the production of pectinase by Aspergillus niger CH4 in solid-state (SSF) and submerged (SmF) fermentations. Production of endo- (endo-p) and exo-pectinase (exo-p) by SSF was not reduced when glucose, sucrose or galacturonic acid (up to 10%) were added to a culture medium containing pectin. Moreover, both activities increased when concentrations of the carbon sources were also increased. In SmF, these activities were strongly decreased when glucose or sucrose (3%) was added to culture medium containing pectin. The addition of galacturonic acid affected endo-p activity production to a lesser extend than exo-p. Final endo-p and exo-p activities in SSF were three and 11 times higher, respectively, than those obtained in SmF. The overall productivities of SSF were 18.8 and 4.9 times higher for endo-p and exo-p, respectively, than those in SmF. These results indicate that regulatory phenomena, such as induction-repression or activation-inhibition, related to pectinase synthesis by A. niger CH4 are different in the two types of fermentation. Correspondence to: E. Favela-Torres     

Production of tannase from wood-degrading fungus using as substrate plant residues: purification and characterization

In the present study Lenzites elegans, Schizophyllum commune, Ganoderma applanatum and Pycnoporus sanguineus (wood-degrading fungi) were assayed for their tannase producing potential in culture media containing plant residues or/and tannic acid as carbon source. Aspergillus niger was used as positive control for tannase production. We also carried out the isolation, purification and characterization of the enzyme from the fungi selected as the major productor. The highest fungal growth was observed in A. niger and L. elegans in the media containing tannic acid + glucose + plant residues (Fabiana densa). A. niger and L. elegans reached the highest extracellular tannase production in a medium containing tannic acid + F. densa and in a medium supplemented with glucose + tannic acid + F. densa. The produced enzyme by L. elegans was purified by DEAE-Sepharose. Km value was 5.5 mM and relative molecular mass was about 163,000. Tannase was stable at a pH range 3.0–6.0 and its optimum pH was 5.5. The enzyme showed an optimum temperature of 60°C and was stable between 40 and 60°C. This paper is the first communication of tannase production by wood-degrading fungi. Fermentation technology to produce tannase using plant residues and xylophagous fungi could be very important in order to take advantage of plant industrial waste.     

Heterologous protein secretion from Aspergillus niger in phosphate-buffered batch culture

Summary Aspergillus niger was grown in batch culture containing various initial concentrations of sodium phosphate buffer (pH 6.5). A wild-type strain of A. niger and a transformed strain producing hen egg-white lysozyme were studied. The maximum cell yield was attained in medium not supplemented with phosphate. In those cultures acidification of the medium resulted in a minimum of pH 2.0 before reverting to near neutrality. Increasing the initial levels of phosphate buffer reduced the fall in pH but lowered cell yields. Secreted levels of lysozyme were maximal in the 50–100 mm range of added phosphate buffer although mycelial yields were reduced by one third of mycelial yields in medium unsupplemented with phosphate buffer. Offprint requests to: D. B. Archer     

A newly isolated fungal strain used as whole‐cell biocatalyst for biodiesel production from palm oil

A strain of Aspergillus niger isolated from atmospherically exposed bread and Jatropha curcas seed was utilized as a whole‐cell biocatalyst for palm oil methanolysis to produce fatty acid methyl esters (FAME), or biodiesel. The A. niger strain had a lipase activity of 212.58 mU mL^?1 after 144 h incubation at 25 °C with an initial pH value of 6.5, using 7% polypeptone (w/w on basal medium) as the nitrogen source and 3% olive oil (w/w on basal medium) as a carbon source. The A. niger cells spontaneously immobilized within polyurethane biomass support particles (BSPs) during submerged fermentation. Thereafter, the methanolysis of palm oil was achieved via a three‐step addition of methanol in the presence of BSPs‐immobilized with A. niger cells. The influence of water content, reaction temperature and enzyme concentration on reaction rate was investigated. An 8% water content and a temperature of 40 °C in the presence of 30 immobilized BSPs, resulted in an 87% FAME yield after 72 h.     

Bioleaching of nickel from equilibrium fluid catalytic cracking catalysts

Summary This study investigates the possibility of reusing metal-contaminated equilibrium fluid catalytic cracking (FCC) catalyst after bioleaching. Leaching with Aspergillus niger culture was found to be more effective in the mobilization of nickel from the catalyst particles compared to chemical leaching with citric acid. Bioleaching achieved 32% nickel removal whereas chemical leaching achieved only 21% nickel removal from catalyst particles. The enhanced nickel removal from the catalysts in the presence of A. niger culture was attributed to the biosorption ability of the fungal mycelium and to the higher local concentration of citric acid on the catalyst surface. It was found that 9% of solubilized nickel in the liquid medium was biosorbed to fungal biomass. After nickel leaching with A. niger culture, the hydrogen-to-methane molar ratio and coke yield, which are the measures of dehydrogenation reactions catalysed by nickel during cracking reactions, decreased significantly.     

Ram horn peptone as a source of citric acid production by <Emphasis Type="Italic">Aspergillus niger</Emphasis>, with a process

The present study deals with the production of citric acid from a ram horn peptone (RHP) by Aspergillus niger NRRL 330. A medium from RHP and a control medium (CM) were compared for citric acid production using A. niger in a batch culture. For this purpose, first, RHP was produced. Ram horns were hydrolyzed by treatment with acids (6 N H₂SO₄, 6 N HCl) and neutralizing solutions. The amounts of protein, nitrogen, ash, some minerals, total sugars, total lipids and amino acids of the RHP were determined. RHP was compared with peptones with a bacto-tryptone from casein and other peptones. The results from RHP were similar to those of standard peptones. The optimal concentration of RHP for the production of citric acid was found to be 4% (w/w). A medium prepared from 4% RHP was termed ram horn peptone medium (RHPM). In comparison with CM, the content of citric acid in RHPM broth (84 g/l) over 6 days was 35% higher than that in CM broth (62 g/l). These results show that citric acid can be produced efficiently by A. niger from ram horn.     

Production of tannase by <Emphasis Type="Italic">Aspergillus niger</Emphasis> HA37 growing on tannic acid and Olive Mill Waste Waters

Production of tannase (tannin acyl hydrolase, EC 3.1.1.20) by Aspergillus nigerHA37 on a synthetic culture medium containing tannic acid at different concentrations has been studied. Maximal enzymatic activity increased according to the initial concentration of tannic acid; respectively 0.6, 0.9 and 1.5 enzyme activity units (EU) ml⁻¹ medium in the presence of 0.2%, 0.5% and 1% of tannic acid. Tannase production by A. niger HA37 on fourfold diluted olive mill waste waters (OMWW) as substrate, was between 0.37 and 0.65 EU ml⁻¹. Enzyme production on the diluted OMWW remained globally stable during more than 30 h. Growth of A. niger HA37 on OMWW was correlated with about 70% degradation of phenolic compounds present in the waste. This strain has therefore the capacity to degrade complex wastewaters which cause environmental damage to aquatic streams.     

Development of a serial bioreactor system for direct ethanol production from starch using<Emphasis Type="Italic">Aspergillus niger</Emphasis> and<Emphasis Type="Italic">Saccharomyces cerevisiae</Emphasis>

Aspergillus niger hyphae were found to grow with unliquefied potato starch under aerobic conditions, but did not grow under anaerobic conditions. The raw culture ofA. niger catalyzed saccharification of potato starch to glucose, producing approximately 12 g glucose/L/day/ The extracellular enzyme activity was decreased in proportion to incubation time, and approximately 64% of initial activity was maintained after 3 days. At 50°C,A. niger hyphae growth stopped, while the extracellular enzyme activity peaked. On the basis of theA. niger growth property and enzyme activity, we designed a serial bioreactor system composed of four different reactors. Fungal hyphae were cultivated in reactor I at 30°C, uniquefied starch was saccharified to glycose by a fungal hyphae culture in reactors II and III at 50°C, and glucose was fermented to ethanol bySaccharomyces cerevisiae in reactor IV. The total glucose produced by fungal hyphae in reactor I and saccharification in reactor II was about 42 g/L/day. Ethanol production in reactor IV was approximately 22 g/L/day, which corresponds to about 79% of the theoretical maximum produced from 55 g starch/L/day.     

Fungal infections of ear with special reference to chronic suppurative otitis media

Fungus were found to take important role in ear infections of the 344 patients (CSOM 286, Otomycosis 44, Otitis externa 14), significant fungal infections (with positive smear and culture) were detected on 49%, 79.5%, 66.6% patients respectively. 84.8% patients were detected both by smear and culture, 14.1% patients by culture and 0.1% patients in smear preparation only. In CSOM patients, age predominated in 20–27 yrs group, sex in male below 30 yrs, and Aspergillus flavus, A. niger, Penicillium, A. fumigatus in mycelial fungus, Candida albicans, C. parapsillosis in yeast. But in 18 post antibiotic fungus infected patients Penicillium and A. niger were the important isolates. In otomycosis and otitis externa patients A. niger took the main role.     

Metabolism of dimethylterephthalate byAspergillus niger

Aspergillus niger (AG-1) metabolized dimethylterephthalate through monomethylterephthalate, terephthalate and protocatechuate. Degradation of dimethylterephthalate was followed by extraction of residual dimethylterephthalate from the spent medium. The quantitative UV analysis showed that 58% of the dimethylterephthalate supplement was taken up in 144 h. The metabolites were isolated from resting cell cultures. Thin layer chromatography analysis of the extract revealed the presence of two intermediates, monomethylterephthalate and terephthalate. Use of an inhibitor in resting cell culture experiment demonstrated the accumulation of protocatechuate. The time course of protocatechuate accumulation was also studied. Metabolites were identified by employing various physicochemical methods. Enzyme studies using cell-free extracts exhibited dimethylterephthalate esterase and protocatechuate dioxygenase activities. Protocatechuate was oxidized by themeta cleavage pathway. A tentative pathway for the degradation of DMTP has been proposed inA. niger.Abbreviations A. niger Aspergillus niger (AG1) - DMSO dimethyl sulfoxide - DMTP dimethylterephthalate - MMTP monomethylterephthalate - MS mass spectra - NMR nuclear magnetic resonance spectra - PCA protocatechuate - TLC thin layer chromatography - TP terephthalate - UV ultra violet spectra     

Tryptophan auxotrophic mutants in Aspergillus niger: inactivation of the trpC gene by cotransformation mutagenesis

Summary Aspergillus niger tryptophan auxotrophic mutants have been isolated after UV irradiation of conidiospores. The mutants belong to two different complementation groups, trpA and trpB, which complement each other in heterokaryons. Neither of the mutations could be complemented with the cloned A. niger trpC gene. To obtain A. niger trpC mutants in a direct way, gene inactivation by cotransformation was performed. For this purpose an in-frame gene fusion between the A. niger trpC and Escherichia coli lacZ genes was constructed and shown to be functionally expressed after introduction into A. niger by cotransformation with the pyrA gene as selective marker. Among the -galactosidase expressing cotransformants, obtained with either circular or linearized vectors, no trpC mutants were detected, even after enrichment. Such mutants, however, could be obtained by cotransformation of A. niger with specific fragments of the fusion gene. Biochemical analysis of the cotransformants indicated that in nearly all cases the fusion gene had replaced the wild-type trpC gene. Genetic analysis showed that the trpC mutation is not linked to any of the A. niger loci described so far. The trpC mutants can be complemented by the cloned A. niger trpC gene as well as by the A. nidulans trpC gene.     

Biotransformation and improved enzymatic extraction of chlorogenic acid from coffee pulp by filamentous fungi

The highest enzymatic extraction of covalent linked chlorogenic (36.1%) and caffeic (CA) (33%) acids from coffee pulp (CP) was achieved by solid‐state fermentation with a mixture of three enzymatic extracts produced by Aspergillus tamarii, Rhizomucor pusillus, and Trametes sp. Enzyme extracts were produced in a practical inexpensive way. Synergistic effects on the extraction yield were observed when more than one enzyme extract was used. In addition, biotransformation of chlorogenic acid (ChA) by Aspergillus niger C23308 was studied. Equimolar transformation of ChA into CA and quinic acids (QA) was observed during the first 36 h in submerged culture. Subsequently, after 36 h, equimolar transformation of CA into protocatechuic acid was observed; this pathway is being reported for the first time for A. niger. QA was used as a carbon source by A. niger C23308. This study presents the potential of using CP to produce enzymes and compounds such as ChA with biological activities. © 2013 American Institute of Chemical Engineers Biotechnol. Prog., 29: 337–345, 2013