Isolation and Identification of Glucose Oxidase Hyper Producing Strain of Aspergillus niger.

Aims: Glucose oxidase is an enzyme with large scale applications in various industries. It is also used in several diagnostic kits which makes it medically important as well. Our aim was to isolate indigenous glucose oxidase hyper producing strain of Aspergillus niger from different soil samples of Punjab, Pakistan. Study Design: An experimental study. Place and Duration of Study: Institute of Industrial Biotechnology, GC University, Lahore from March 2011 to July 2012. Methodology: Two hundred and seventy nine fungal strains were isolated from soil of different localities of Punjab. Isolates were screened for glucose oxidase production using submerged fermentation. Glucose oxidase hyper producer isolate was identified using morphological and molecular techniques i.e. 18S rDNA. DNA was isolated and amplified using PCR. Gene sequencing was done and homology analysis was studied. Rate of glucose oxidase production was also analysed. Results: Glucose oxidase hyper producing isolate was identified as A. niger A247 strain. This strain gave best reproducible results (145.22 ±0.034 U/g of cell mass) after 72 hrs of fermentation at 30ºC and at a medium pH of 7.2. Conclusion: Our results indicate the natural ability of A. niger to produce Glucose oxidase in large quantity instead of using genetic manipulation techniques.

Use of Aspergillus niger in the bioleaching of colemanite for the production of boric acid

Colemanite is one of the most important underground riches of Turkey, having approximately 60 percent of the world boron deposits, and it has a large portion in the deposits. In this study, chemical leaching and biological leaching methods were used for production of boric acid from colemanite (2CaO · 3B3O3 · 5H2O) (Emet-Kütahya, Turkey). Oxalic acid concentration, temperature, stirring time and solid-to-liquid ratio were taken as parameters in the chemical leaching process. It was found that the dissolution rate increases with increasing oxalic acid concentration and temperature but it decreases at higher solid-to-liquid ratios in the chemical leaching process. Using optimum conditions (d100 = 0.075 mm; 5 percent solids by weight; 0.55 M oxalic acid; 80 +/- 2 ºC leaching temperature; 150 rpm stirring speed; 90 min leaching time) for colemanite sample (28.05 percent B2O3) on chemical leaching with oxalic acid experiments, the calculated boric acid extraction efficiency from colemanite ore was 97.89 percent. Optimum conditions on bioleaching of Emet-Kütahya, Turkey colemanite ores using the fungus Aspergillus niger were found to be as follows: reaction temperature 25 +/- 2ºC; solid-to-liquid ratio 5 percent solids by weight; d100 = 0.075 mm; stirring speed 150 rpm; initial the fungus populations in the inocula about 3 x 10(7) cells/ml and reaction time 21 days. The calculated boric acid extraction efficiency from colemanite ore was 90.18 percent under the optimum conditions. Bioleachate contained 12.95 g/l B2O3, 6.60 g/l Ca and 0.087 g/l Mg. Compared with chemical leaching at 5 percent pulp density, the fungus was less efficient in the extraction of B2O3 from colemanite but the difference in the extraction yields between the two processes was less than 10 percent. Although bioleaching generally requires a longer period of operation compared to chemical leaching, these results suggest that bioleaching by A...

Influencia de la concentración de inóculo en la producción de celulasa y xilanasa por aspergillus niger / Influence of inoculum concentration on the cellulase and xylanase production by aspergillus niger

Existe un gran interés por el uso de enzimas lignocelulolíticas en varias industrias, y en la biodegradación de biomasa para la producción de biocombustibles y otras aplicaciones. Entre las fuentes microbianas de enzimas, Aspergillus niger es uno de los microorganismos más utilizados en la producción de enzimas industriales, debido a sus niveles altos de secreción de proteína y a su condición GRAS (generally regarded as safe). El objetivo del presente estudio fue evaluar la influencia de la concentración de inóculo en la morfología y producción de celulasas y xilanasas con A. niger en cultivo sumergido. Para ello, fueron inoculados matraces de 250 mL con 40 mL de medio con 3% (v/v) de una suspensión de 104 o 108 esporas por mililitro e incubados a 28 ºC y 175 rpm durante 120 horas. Se utilizaron 10 g*L-1 de lactosa como fuente de carbono. En cada caso se determinó la cantidad de biomasa, la proteína extracelular soluble, lactosa residual, actividad celulasa total y xilanasa cada 24 horas. Aunque no hubo un efecto notorio en la morfología de crecimiento, salvo en el color y el diámetro de pellets obtenidos, sí se afectó la µmax (0,06 y 0,03 h-1 para 104 y 108 esporas*mL-1, respectivamente) y la concentración máxima de biomasa. Además, mientras que las productividades volumétricas de celulasa (ΓFPA) (8,2 y 8,0 UI.*L-1*h-1 para 104 y 108 esporas*mL-1, respectivamente) fueron similares para ambos inóculos, la productividad de xilanasa (ΓXIL) fue mayor para el inóculo más concentrado (29,7 y 33,4 UI¨*L-1*h-1 para 104 y 108 esporas*mL-1, respectivamente). Los resultados indican que la productividad de celulasas y xilanasas está estrechamente relacionada con la concentración de inóculo.

There is a great interest for the use of lignocellulolytic enzymes in several industries and in biomass degradation for production of biofuels and other applications. Among the microbial sources of enzymes, Aspergillus niger is one of the most used microorganisms in the production of industrial enzymes due to its high levels of protein secretion and its GRAS (generally regarded as safe) condition. The aim of the present study was to evaluate the influence of A. niger inoculum concentration in the morphology and production of cellulases and xylanases in submerged cultures. For this, 250 mL flasks containing 40 mL culture medium were inoculated with a 3% (v/v) of either 104 or 108 spores per milliliter suspension and incubated at 28 º C and 175 rpm during 120 hours. Lactose (10 g*L-1) was used as the carbon source. In each case, the amount of biomass, the extracellular soluble protein, residual lactose, total celullase activity and xylanase activity were determined every 24 hours. Even thought there was not a notorious effect on the growth morphology, except in color and diameter of pellets; µmax was affected (0.06 and 0.03 h-1 for 104 and 108 spores*mL-1, respectively) as well as maximum biomass concentration. In addition, while the volumetric productivity of cellulase (8.2 and 8.0 UI*L-1*h-1 for 104 and 108 spores*mL-1, respectively) were similar for both inocula, the productivity of xylanase was greater for the more concentrated inoculum (29.7 and 33.4 UI*L-1*h-1 for 104 and 108 spores*mL-1, respectively).The results show that cellulase and xylanase productivities are closely related to the inoculum concentration.

Biosorption of Cr (VI) ions from electroplating industrial effluent using immobilized Aspergillus niger biomass.

A fungus, Aspergillus niger was chemically treated with 0.1 M H2SO4 and 0.1 N NaOH to form biosorbent and it was immobilized in calcium alginate beads. The biosorption capacity of immobilized biosorbents for Cr (VI) was found to depend on pH, contact time, biosorbent dose and initial concentration of Cr (VI). The maximum uptake of Cr (VI) was 92.5, 95.9 and 98.4 mg respectively at a pH of 1.5 and with an increase in pH up to 10.5 the metal uptake decreased gradually up to 38.75, 50.19 and 65.28 mg respectively for acid treated, untreated and base treated fungal biosorbents. Increase in biosorbent dose up to 1 g of biomass and contact time up to 60 min resulted in an increase in biosorption from 19.6, 15.6 and 26.1 mg at a biosorbent dose of 0.1 g 100 ml(-1) to 92.45, 95.7 and 98.52 mg at a biosorbent dose of 1.0 g 100 ml(-1) and then further increase in adsorbent dose and contact time did not resulted in more Cr (VI) adsorption by per unit weight of biosorbent. The value of Kad (adsorption rate constant) revealed the pseudo-first order nature of biosorption. The percentage metal uptake by the biosorbent was found to decrease upto 62.33, 52.67 and 83.5 percent respectively for acid treated, untreated and base treated fungal biosorbents at the 300 mgl(-1) Cr (VI) ion concentration. The resulted data was found to fit well in Langmuir model of adsorption isotherm with a high value of correlation coefficient. The value of Qmax, b (Langmuir constants), R(L) (separation factor) and delta G (Gibb's free energy) revealed the favourable nature of adsorption. The biosorbed metal was eluted from the biosorbent by using 0.1 M H2SO4 as elutant. Immobilized biosorbent can be reused for five consecutive biosorption/desorption cycles without apparent loss of efficiency after its reconditioning. The biosorbent was found to perform well in the electroplating industrial effluent.

Biochemical characterization of N-methyl N'-nitro-N-nitrosoguanidine-induced cadmium resistant mutants of Aspergillus niger.

Two cadmium resistant mutants (Cd1 and Cd2) of Aspergillus niger, among the six isolated by mutagenization with N-methyl N'-nitro-N-nitrosoguanidine (MNNG) at pH 6.4 were selected for the study. Analysis of lipid composition of the mutants and the wildtype indicated that total lipid as well as individual lipids of the cadmium resistant mutants were changed as compared with that of the wildtype. The increased activities of metal-lothionein and reduced activities of D-xylose isomerase and L-phenylalanine ammonia lyase in cell free extract of the cadmium resistant mutants suggested that mutants could allow high concentration of cadmium salt as compared with that of the wildtype. The respiratory activity and intracellular as well as extracellular Cd2+ concentration of the mutants reflected the high tolerance of the Cd mutants to cadmium ion.

Fungal metabolites from Aspergillus niger AN27 related to plant growth promotion.

Two metabolites have been isolated from Aspergillus niger AN27, a biocontrol agent, and identified as 2-carboxymethyl 3-n-hexyl maleic acid (compound 1) and 2-methylene-3-hexylbutanedioic acid (compound 2). Their biological activities related to crop growth promotion have been assayed. Both the compounds increased germination and improved crop vigour. Compound 1 was more effective for increase in germination and shoot length, whereas compound 2 had relatively greater role in increasing the root length and biomass of cauliflower seedlings.