Production of fungal phytases in solid state fermentation and potential biotechnological applications.

Phytases are important enzymes used for eliminating the anti-nutritional properties of phytic acid in food and feed ingredients. Phytic acid is major form of organic phosphorus stored during seed setting. Monogastric animals cannot utilize this phytate-phosphorus due to lack of necessary enzymes. Therefore, phytic acid excretion is responsible for mineral deficiency and phosphorus pollution. Phytases have been reported from diverse microorganisms, however, fungal phytases are preferred due to their unique properties. Aspergillus species are the predominant producers of phytases and have been explored widely as compared to other fungi. Solid-state fermentation has been studied as an economical process for the production of phytases to utilize various agro-industrial residues. Mixed substrate fermentation has also been reported for the production of phytases. Physical and chemical parameters including pH, temperature, and concentrations of media components have significantly affected the production of phytases in solid state fermentation. Fungi produced high levels of phytases in solid state fermentation utilizing economical substrates. Optimization of culture conditions using different approaches has significantly improved the production of phytases. Fungal phytases are histidine acid phosphatases exhibiting broad substrate specificity, are relatively thermostable and protease-resistant. These phytases have been found effective in dephytinization of food and feed samples with concomitant liberation of minerals, sugars and soluble proteins. Additionally, they have improved the growth of plants by increasing the availability of phosphorus and other minerals. Furthermore, phytases from fungi have played an important roles in bread making, semi-synthesis of peroxidase, biofuel production, production of myo-inositol phosphates and management of environmental pollution. This review article describes the production of fungal phytases in solid state fermentation and their biotechnological applications.

Green synthesis of iron nanoparticles: Sources and multifarious biotechnological applications.

Green synthesis of iron nanoparticles is a highly fascinating research area and has gained importance due to reliable, sustainable and ecofriendly protocol for synthesizing nanoparticles, along with the easy availability of plant materials and their pharmacological significance. As an alternate to physical and chemical synthesis, the biological materials, like microorganisms and plants are considered to be less costly and environment-friendly. Iron nanoparticles with diverse morphology and size have been synthesized using biological extracts. Microbial (bacteria, fungi, algae etc.) and plant extracts have been employed in green synthesis of iron nanoparticles due to the presence of various metabolites and biomolecules. Physical and biochemical properties of biologically synthesized iron nanoparticles are superior to that are synthesized using physical and chemical agents. Iron nanoparticles have magnetic property with thermal and electrical conductivity. Iron nanoparticles below a certain size (generally 10-20 nm), can exhibit a unique form of magnetism called superparamagnetism. They are non-toxic and highly dispersible with targeted delivery, which are suitable for efficient drug delivery to the target. Green synthesized iron nanoparticles have been explored for multifarious biotechnological applications. These iron nanoparticles exhibited antimicrobial and anticancerous properties. Iron nanoparticles adversely affect the cell viability, division and metabolic activity. Iron nanoparticles have been used in the purification and immobilization of various enzymes/proteins. Iron nanoparticles have shown potential in bioremediation of various organic and inorganic pollutants. This review describes various biological sources used in the green synthesis of iron nanoparticles and their potential applications in biotechnology, diagnostics and mitigation of environmental pollutants.

Enhanced production and immobilization of phytase from Aspergillus oryzae: a safe and ideal food supplement for improving nutrition.

Microbial phytases are potentially excellent candidates for eliminating anti-nutrient i.e. phytic acid, due to hydrolysis of phospho-monoester linkages present in the phytic acid. An average 2.29-fold increase in phytase production was obtained after statistical optimization in solid-state fermentation. Aspergillus oryzae SBS50 phytase was immobilized on a Ca-alginate matrix with an effectiveness of 53%. Immobilized-phytase retained > 50% activity after recycling for five cycles and also displayed more stability in the presence of organic solvents, metal ions, and detergents as compared to free enzyme. Values of Km and Vmax of immobilized phytase were recorded as 0.66 mM and 666.6 nmol/sec, respectively. Immobilized phytase efficiently hydrolyzed the phytate contents in wheat and pearl millet flours, exhibiting > 70% catalytic activity even after three cycles. Phytase supplementation resulted in the improved nutritional quality of these flours. Furthermore, the safety assessment of the treated and untreated samples reveals the absence of any aflatoxin in the phytase produced by the mould. The results revealed the improved stability of phytase after immobilization and as a safe and significant additive for application in the food industry.

Immobilized phytases: an overview of different strategies, support material, and their applications in improving food and feed nutrition.

Phytases are the most widely used food and feed enzymes, which aid in nutritional improvement by reducing anti-nutritional factor. Despite the benefits, enzymes usage in the industry is restricted by several factors such as their short life-span and poor reusability, which result in high costs for large-scale utilization at commercial scale. Furthermore, under pelleting conditions such as high temperatures, pH, and other factors, the enzyme becomes inactive due to lesser stability. Immobilization of phytases has been suggested as a way to overcome these limitations with improved performance. Matrices used to immobilize phytases include inorganic (Hydroxypatite, zeolite, and silica), organic (Polyacrylamide, epoxy resins, alginate, chitosan, and starch agar), soluble matrix (Polyvinyl alcohol), and nanomaterials including nanoparticles, nanofibers, nanotubes. Several surface analysis methods, including thermal gravimetric analysis (TGA), differential scanning calorimetry (DSC), and FTIR analysis, have been used to characterize immobilized phytase. Immobilized phytases have been used in a broad range of biotechnological applications such as animal feed, biodegradation of food phytates, preparations of myo-inositol phosphates, and sulfoxidation by vanadate-substituted peroxidase. This article provides information on different matrices used for phytase immobilization from the last two decades, including the process of immobilization and support material, surface analysis techniques, and multifarious biotechnological applications of the immobilized phytases.

Biochemical properties of cellulolytic and xylanolytic enzymes from Sporotrichum thermophile and their utility in bioethanol production using rice straw.

Production of cellulolytic and xylanolytic enzymes by Sporotrichum thermophile was enhanced using response surface methodology in solid-state fermentation (SSF) using wheat straw and cotton oil cake. Cellulolytic and xylanolytic enzymes were partially purified by ammonium sulfate precipitation followed by ion exchange and gel filtration chromatographic techniques. Xylanase of S. thermophile is neutral xylanase displaying optimal activity at 60 °C with Km and Vmax values of 0.2 mg/mL and 238.05 µmole/min, respectively. All cellulases produced by the thermophilic mold showed optimal activity at pH 5.0 and 60 °C with Km values of 0.312 mg/mL, 0.113 mg/mL, and 0.285 mM for carboxymethyl cellulase (CMCase), filter paper cellulase (FPase), and ß-glucosidase, respectively and while Vmax values were 181.81, 138.88, and 66.67 µmole/min, respectively. The presence of various metal ions (Ca2+ and Co2+), chemical reagent (glutaraldehyde), and surfactants (Tween 80 and Triton X-100) significantly improved the activities of all enzymes. All the enzymes showed high storage stability under low temperature (-20 and 4 °C) conditions. Cellulolytic and xylanolytic enzymes resulted in enhanced liberation of reducing sugars (356.34 mg/g) by hydrolyzing both cellulosic and hemicellulosic fractions of ammonia-pretreated rice straw as compared to other pretreatment methods used in the study. Fermentation of enzymatic hydrolysate resulted in the formation of 28.88 and 27.18 g/L of bioethanol in separate hydrolysis and fermentation (SHF) process by Saccharomyces cerevisiae and Pichia stipitis, respectively. Therefore, cellulolytic and xylanolytic enzymes of S. thermophile exhibited ideal properties of biocatalysts useful in the saccharification of cellulosic and hemicellulosic fractions of rice straw for the production of bioethanol.

Enhanced Phytase Production by Bacillus subtilis subsp. subtilis in Solid State Fermentation and its Utility in Improving Food Nutrition.

BACKGROUND: Phytic acid acts as anti-nutritional factor in food and feed ingredients for monogastric animals as they lack phytases. OBJECTIVE: Phytase production by Bacillus subtilis subsp. subtilis JJBS250 was studied in solid-state fermentation and its applicability in dephytinization of food. METHODS: Bacterial culture was grown in solid state fermentation using wheat bran and various culture conditions were optimized using 'One variable at a time' (OVAT) approach. Effects of different substrates (wheat bran, wheat straw, sugarcane bagasse), incubation time (24, 48, 72 and 96 h), incubation temperatures (25, 30, 35 and 40°C), pH (4.0, 5.0, 6.0, 7.0 and 8.0) and moisture content (1:1.5, 1:2.0, 1:2.5 and 1:3) were studied on phytase production. Bacterial phytase was used in dephytinization of food samples. RESULTS: Optimization of phytase production was studied in solid state fermentation (SSF) using 'One variable at a time' (OVAT) approach. Bacillus subtilis subsp. subtilis JJBS250 grew well in various agroresidues in SSF and secreted high enzyme titres using wheat bran at 30°C and pH 5.0 after incubation time of 48 h with substrate to moisture ratio of 1:3. Glucose and ammonium sulphate supplementation to wheat bran further enhanced phytase production in SSF. Optimization of phytase production resulted in 2.4-fold improvement in phytase production in solid state fermentation. The enzyme resulted in dephytinization of wheat and rice flours with concomitant release of inorganic phosphate, reducing sugar and soluble protein. CONCLUSION: Optimization resulted in 2.34-fold enhancement in phytase production by bacterial culture that showed dephytinization of food ingredients with concomitant release of nutritional components. Therefore, phytase of B. subtilis subsp. subtilis JJBS250 could find application in improving nutritional quality of food and feed of monogastric animals.

Utility of acidic xylanase of Bacillus subtilis subsp. subtilis JJBS250 in improving the nutritional value of poultry feed.

Cane molasses has been employed as a cost-effective medium for enhanced xylanase production in submerged fermentation. Bacillus subtilis subsp. subtilis JJBS250 produced xylanase (15.16 U/ml) at pH 4.0, 35 °C and 200 rpm after 54 h using optimized basal medium by 'one variable at a time approach'. Addition of Tween 80 and PEG 4000 also enhanced xylanase production in cane molasses medium. Combined effect of yeast extract, incubation time and PEG 4000 using statistical optimization enhanced xylanase production to 38.60 U/ml, which was 2.54-fold higher than the 'one variable at a time approach'. The efficacy of xylanase from Bacillus subtilis subsp. subtilis JJBS 250 was evaluated in the improvement of poultry feed nutrition. Xylanase addition (10 IU/g feed) enhanced liberation of reducing sugars (95.540 mg/g substrate) after 48 h at 60 °C. Optimization has resulted in enhanced production of xylanase that improved the nutritional quality of poultry feed.

Purification and kinetics of a protease-resistant, neutral, and thermostable phytase from Bacillus subtilis subsp. subtilis JJBS250 ameliorating food nutrition.

A novel protease-resistant and thermostable phytase from Bacillus subtilis subsp. subtilis JJBS250 was purified 36-fold to homogeneity with a combination of ammonium sulfate precipitation followed by Q-Sepharose and Sephadex G-50 chromatographic techniques. The estimated molecular mass of the purified phytase was 46 kDa by electrophoresis with optimal activity at pH 7.0 and 70 °C. About 19% of original activity was maintained at 80 °C for 10 min. Phytase activity was stimulated in presence of surfactants like Tween-20, Tween-80, and Triton X-100 and metal ions like Ca+2, K+, and Co+2 and it was inhibited by SDS and Mg+2, Al+2, and Fe+2. Purified enzyme showed specificity to different salts of phytic acid and values of Km and Vmax were 0.293 mM and 11.49 nmoles s-1, respectively for sodium phytate. The purified enzyme was resistant to proteases (trypsin and pepsin) that resulted in amelioration of food nutrition with simultaneous release of inorganic phosphate, reducing sugars, and soluble protein.