Recovery of filter paperase from mouldy rice husk in solid state fermentation by Aspergillus protuberus.

Recovery of enzymes such as FPase (filter paperase) or exoglucanase from fermented substrate is a sustainable approach in enzyme production; however, there is a scarcity of optimization studies in this field. The present study was aimed to standardize number of parameters (selection of solvent, solvent volume, soaking time, leaching conditions and number of washes) to extract maximum amount of FPase from fermented rice husk by Aspergillus protuberus. Novel Aspergillus protuberus was first report from our lab on cellulases production in solid state fermentation (SSF). Among the tested solvents, citrate phosphate buffer (0.02 M, pH 5.0) proved best solvent for maximum recovery of FPase. Consequent experimental parameters were further optimized with citrate phosphate buffer. Two washes with citrate phosphate buffer each by shaking (60 min) in a ratio of 1 g of rice husk: 5 ml of citrate phosphate buffer together attained higher recovery efficiency (88 %) of FPase from the fermented rice husk.

Leaching Approach for ß-Glucosidase Extraction from Fermented Rice Husk in Solid State Cultivation by Aspergillus protuberus.

Environmental problems are caused by the disposal of agrowastes in developing countries. It is imperative to convert such wastes into useful products, which require enzymes such as ß-glucosidase. ß-Glucosidase has variety of applications in biotechnology including food, textile, detergents, pulp and paper, pharmaceutical and biofuel industries. ß-Glucosidase production was performed using the locally isolated Aspergillus protuberus using best growth circumstances on rice husk in solid-state fermentation (SSF). Leaching of ß-glucosidase from fermented rice husk with number of solvents to evaluate their extraction efficacy. Among the different solvents examined, acetate buffer (0.02 M, pH 5.0) proved to be the best solvent. The subsequent parameters were optimized with acetate buffer. Two washes with acetate buffer each by shaking (30 min) in a ratio of 1 g of rice husk: 5 ml of acetate buffer together attained maximum recovery of ß-glucosidase with 41.95 U/g of rice husk.

Immunodiagnostic potential of Wuchereria bancrofti L1 antigen-based filarial immunoglobulin G4 detection assay.

Background: After mass drug administration to eliminate human lymphatic filariasis, there is a need for surveillance to detect the measurable endpoint of the program. Methods: An immunodominant seroreactive clone, WbL1, was identified through immunoscreening of a Wuchereria bancrofti L3 complementary DNA expression library. Recombinant WbL1 (rWbL1) was analysed with sera from W. bancrofti patients. Diagnostic evaluation was carried out by developing an enzyme-linked immunosorbent assay (ELISA) to detect the filarial-specific antibodies in various categories of filarial sera samples against recombinant WbL1 (rWbL1) protein. Results: Performance parameters of the test in terms of immunoglobulin G (IgG) and IgG4 detection displayed significant sensitivity and specificity values up to 77% and 100%, respectively. Our results showed filarial antibodies against rWbL1 to be highly reactive with microfilaremic and clinical filarial sera samples compared with the endemic and non-endemic control sera samples. Reasonably satisfactory performance of the test was also confirmed from the multicentric evaluation of an anti-WbL1 IgG4 detection ELISA. This test was found to be minimally reactive with other nematode parasites and protozoan infections. Conclusions: The anti-WbL1 IgG4 detection test can be considered as a field test for initial screening and epidemiological monitoring of filarial infections in filariasis-endemic areas.

Using sulfite chemistry for robust bioconversion of Douglas-fir forest residue to bioethanol at high titer and lignosulfonate: a pilot-scale evaluation.

This study demonstrated at the pilot-scale (50 kg) use of Douglas-fir forest harvest residue, an underutilized forest biomass, for the production of high titer and high yield bioethanol using sulfite chemistry without solid-liquor separation and detoxification. Sulfite Pretreatment to Overcome the Recalcitrance of Lignocelluloses (SPORL) was directly applied to the ground forest harvest residue with no further mechanical size reduction, at a low temperature of 145°C and calcium bisulfite or total SO2 loadings of only 6.5 or 6.6 wt% on oven dry forest residue, respectively. The low temperature pretreatment facilitated high solids fermentation of the un-detoxified pretreated whole slurry. An ethanol yield of 282 L/tonne, equivalent to 70% theoretical, with a titer of 42 g/L was achieved. SPORL solubilized approximately 45% of the wood lignin as directly marketable lignosulfonate with properties equivalent to or better than a commercial lignosulfonate, important to improve the economics of biofuel production.

Molecular cloning, purification and characterization of thermostable beta-1,3-1,4 glucanase from Bacillus subtilis A8-8.

A gene encoding a beta-1,3-1,4-glucanase (CelA) belonging to family 5 of glycoside hydrolases was cloned and sequenced from the Bacillus subtilis A8-8. The open-reading-frame of celA comprised 1499 base pairs and the enzyme was composed of 500 amino acids with a molecular mass of 55 kDa. The recombinant beta-1,3-1,4 glucanase was purified by GST-fusion purification system. The pH and temperature optima of the enzyme were 8.0 and 60 degrees C, respectively. The enzyme was stable within pH 6.0-9.0. It was stable up to 60 degrees C and retained 30% of its original activity at 70 degrees C for 60 min. It hydrolyzed lichenan, CMC, xylan, laminarin, avicel and pNPC, but was inactive towards cellobiose. The enzyme activity was markedly activated by Co2+ and Mn2+, but was strongly inactivated by Fe3+. The truncated gene, devoid of cellulose-binding domain (CBD) showed 60% of activity and bound to avicel.

Optimization of extraction of ß-endoglucanase from the fermented bran of Aspergillus niger.

A local isolate of Aspergillus niger was cultivated under optimal growth conditions on wheat bran in solid state fermentation. ß-endoglucanase from fermented bran was separately extracted with different solvents to test recovery of enzyme. Among solvents tested, distilled water served the best leachate. Conditions were further optimized with this leachate. Two washes of fermented bran with the leachate for 30 min each under shaking conditions in a ratio of 1 g of wheat bran: 4 ml of distilled water together yielded maximum recovery of 16.7 U/g of wheat bran.

Activities of cellulase and amylase in soils as influenced by insecticide interactions.

Interaction effects of the insecticides monocrotophos and quinalphos (organophosphates), and cypermethrin (pyrethroid), on microbial activities in two agricultural soils-black vertisol soil and red alfinsol soil were tested for 30 days under laboratory conditions. Individual application of the three insecticides at 5, 10 and 25microg g(-1) to the soil distinctly enhanced the activities of cellulase and amylase. Insecticide combinations involving monocrotophos or quinalphos with cypermethrin yielded synergistic, antagonistic and additive interaction effects on both enzymes in the soils. At lower levels, 5 and 10microg g(-1), the insecticides in combination interacted additively or synergistically toward both enzymes. But, both combinations at the highest level of 25microg g(-1) exhibited an antagonistic interaction, with a reduction in enzyme activities to a level lower than that of the control. Interaction effects of insecticides in combinations on two enzyme activities in both soils were related to populations of cellulolytic and amylolytic organisms in soils under the impact of combination of insecticides. These interaction responses were persistent even for 30 days.

Cellulolytic enzymes on lignocellulosic substrates in solid state fermentation by Aspergillus niger.

The production of cellulolytic enzymes by Aspergillus niger on lignocellulosic substrates groundnut fodder, wheat bran, rice bran and sawdust in solid state fermentation in a laboratory scale was compared. Czapek Dox liquid broth amended with cellulose (0.5%) was used to moisten lignocellulosic solid supports for cultivation of Aspergillus niger. The production of filter paperase, carboxymethyl cellulase and -glucosidase were monitored at daily intervals for 5 days. The peak production of the enzymes occurred within 3 days of incubation. Among solid supports used in the study, wheat bran was the best solid matrix followed by groundnut fodder in production of cellulolytic enzymes in solid state fermentation. Groundnut fodder supported significant production of FPase (2.09 FPU/g), CMCase (1.36 U/g) and -glucosidase activity (0.0117 U/g) in solid state fermentation. Considerable secretion of protein (5.10 mg/g) on groundnut fodder at peak time interval 1st day of incubation was recorded.