High Amylase Production by a Novel Strain of Bacillus amyloliquefaciens M37 Isolated from Can Gio Mangrove Forest, Vietnam

Amylases are one of the most important industrial enzymes and find applications in many areas such as textiles, chemicals, food, and pharmaceuticals. Most of the amylases are derived from microbes. The objective of the present study was to evaluate amylase production by a bacterium isolated from the Can Gio mangrove forest. The bacterium was identified as a species of genus Bacillus based on morphological and biochemical characteristics. The analysis of 16S rRNA sequences was then confirmed that this strain belonged to Bacillus amyloliquefaciens species (100% similarity). The effect of culture conditions such as temperature, pH, and carbon sources on amylase production through shake-flask culture was investigated. Maximum amylase activity of 904 IU/mL was obtained after 24 h of cultivation in LB medium containing 1% soluble starch at 35oC and pH 7.0. The highest enzyme activity of 1279 IU/mL was achieved in the bioreactor after 30 h of cultivation at optimum conditions. In addition, B. amyloliquefaciens M37 can grow on soybean meal medium. The high bacterial cell number of 456 × 109 CFU/g and amylase activity of 1039 IU/g were obtained after 36 h of cultivation. This newly isolated B. amyloliquefaciens M37 could be a potential producer for industrial amylase production and probiotics with commercial implications.

Amylase Production by Aspergillus niger and Penicillium Species by Solid-State and Submerged Cultivation Using Two Food Industrial Wastes

Amylase enzymes are starch degrading enzymes and have received a great deal of attention due to their perceived technology importance and economic benefit. Amylase enzymes are considered important enzymes used in starch processing industries for the hydrolysis of polysaccharides like starch into simple sugar constituents. This enzyme is also involved in the commercial production of glucose. Solid-state cultivation and submerged cultivation have tremendous potentials for enzyme amylase production by using different solid substrates like rice bran, wheat bran, coconut oil cake, and groundnut oil cake which are rich in starch. These agro-industrial wastes are considered cheap raw materials for the production of amylase. Wastewater from the industry like brewery can also be used as a liquid substrate for submerged cultivation. It may have the possibility of depurination of wastewater. In the present study, Aspergillus niger and Penicillium species were isolated and their amylase activity was determined by the starch hydrolysis method. Enzyme production was done by using coconut oil cake as a substrate for solid-state fermentation and brewery wastewater as a substrate for submerged fermentation. The enzyme produced by the organisms was extracted and enzyme assay was done by the Dinitrisalicilic method (DNS method). The protein estimation was done by Lowry Folin’s method. The qualitative assay was carried out by performing Gas Chromatography-Mass Spectroscopy (GC-MS).

Optimization of a-Amylase Production from Bacillus Amyloliquefaciens using Taguchi method

α-amylase, an enzyme of industrial importance is used extensively in food, pharmaceutical, textile and detergent industries. Since, a substantial quantity of α-amylase isderived from microbial sources, manipulation of bacterial strain, fermentation conditions and media composition has a major effect on yield of enzyme. Bacillus amyloliqifaciens, obtained from MTCC culture collection was used to study the enhancement of α-amylase production using media concentration manipulation. Taguchi’s orthogonal array was designed for maximization of α-amylase output. The different media components selected as parameters to be optimized were calcium chloride, starch, tryptone, ammonium sulphate and glucose. The concentration of starch and tryptone demonstrated to have maximum effect on amylase production. The optimization strategy was successful in obtaining substantial increase in amylase production of about 2 folds as compared to the unoptimized medium.

PRODUCTION OF EXTRACELLULAR HALOPHILIC AMYLASE BY MARINE CYANOBACTERIUM OSCILLATORIA ACUTISSIMA

This study aimed at the amylase production using seawater instead of fresh water. Amylase is one of the most important enzymes and is very important for biotechnology. Versatile applications of amylase in many industries make optimization of the manufacturing process to achieve maximum yield is in need. Amylase is a widely used and sought-after industrial enzyme. The current research paper describes the production conditions of extracellular amylase from Oscillatoria acutissima. The marine cyanobacterium Oscillatoria acutissima culture was statistically optimized using Plackett-Burman and Box-Behnken designs, under submerged fermentation conditions, for optimum amylase production and activity conditions. The verified Plackett-Burman and Box-Behnken designs results showed that the starch degradation and incubation period improved significantly from 53% to 99% and from 18th to 12th days incubation period, respectively, with pH = 7 at temperature 35°C. SDS-PAGE resolved the molecular weight of partially purified amylase at 72 kilodalton. This research work suggested that marine cyanobacterium Oscillatoria acutissima could be a potential source of halophilic amylase enzyme needed for various industrial processes. To the best of our knowledge, this is one of a few studies on cyanobacteria since while and the first report on amylase production by marine cyanobacterium Oscillatoria acutissima.

Amylase Production by Aspergillus niger immobilized in Microporous Calcium Alginate Gel Beads

Microporous calcium alginate gel beads were investigated as potential solutions to mass transfer limitations in immobilized cultures. The beads were prepared by gelling mixtures of sodium alginate and fillers (starch or granulated sugars) in calcium chloride solution. The resulting beads were cured in the same solution, during which the fillers leached out of the beads thereby creating micro-pores in the beads (microporous beads). The effectiveness of the microporous beads in improving amylase production by Aspergillus niger LC 269109 was investigated. Spores of A. niger were immobilized in the microporous beads and used for batch alpha amylase and gluco amylase production. Amylase production by the A. niger immobilized in the microporous beads were significantly higher (p<0.01) than the values obtained with the conventional calcium alginate gel beads. Under all the conditions investigated, gluco-amylase activities were significantly (p<0.01) higher than the alpha-amylase activities. Under the optimum conditions of inoculum concentration (1.0 × 105 spores/ml), pH (6), temperature (35°C), bead diameter (3 mm) and calcium chloride concentration (1.5%), the gluco-amylase and alpha amylase activities were 11.98 U/ml and 6.7 U/ml respectively, which were significantly higher (p<0.05) than the 7.8 U/ml and 3.2 U/ml obtained with the conventional gel beads.

Environmental Friendly Production of Amylase from Aspergillus niger EFRL-FC-024 Using Corn Waste as Carbon Source

Amylase is an indispensable and industrially important enzyme that hydrolyzes carbohydrates particularly starch into simple sugars. Amylase enzymes have been isolated from various sources such as microbes, animals and plants. However, microorganisms are highly preferred as compared to plants and animal sources. Amylases of fungal origin are highly stable compared to amylases produced by bacterial species. The aim of this study was to investigate the production of extracellular amylase enzyme from Aspergillus niger EFRL-FC-024 using sugarcane bagasse and corn waste as an energy source under submerge fermentation conditions. Primarily, the fungal strain was grown for 6 days using sugarcane bagasse and corn waste, respectively. Mainly, the growth of a microorganism was also evaluated using different pH, temperature and incubation periods. The results revealed maximum amylase production of 1.64 U/mL when A. niger was cultured for 96 h using corn waste. Moreover, addition of different nitrogen sources showed the highest amylase production when peptone was supplemented as a nitrogen source. Finally, the effect of pH indicated maximal concentration of amylase enzyme at pH 6.0. The present study will highly be beneficial to explore the role of fungal strain A. niger in amylase production at Industrial levels.

The yeast eIF2 kinase Gcn2 facilitates H 2 O 2 -mediated feedback inhibition of both protein synthesis and ER oxidative folding during recombinant protein production

Recombinant protein production is a known source of oxidative stress. Knowledge of which ROS are involved or the specific growth phase in which stress occurs however remains lacking. Using modern, hypersensitive genetic H 2 O 2 -specific probes, micro-cultivation and continuous measurements in batch culture, we observed H 2 O 2 accumulation during and following the diauxic shift in engineered Saccharomyces cerevisiae , correlating with peak α-amylase production. In agreement with previous studies supporting a role of the translation initiation factor kinase Gcn2 in the response to H 2 O 2 , we find Gcn2-dependent phosphorylation of eIF2α to increase alongside translational attenuation in strains engineered to produce large amounts of α-amylase. Gcn2 removal significantly improved α-amylase production in two previously optimized high-producing strains, but not in the wild-type. Gcn2-deficiency furthermore reduced intracellular H 2 O 2 levels and the Hac1 splicing ratio whilst expression of antioxidants and the ER disulfide isomerase PDI1 increased. These results suggest protein synthesis and ER oxidative folding to be coupled and subject to feedback inhibition by H 2 O 2 . Importance Recombinant protein production is a multi-billion dollar industry. Optimizing the productivity of host cells is, therefore, of large interest. In several hosts oxidants are produced as an unwanted side product of recombinant protein production. The buildup of oxidants can result in intracellular stress responses which could compromise the productivity of the host cell. Here we document a novel protein synthesis inhibitory mechanism that is activated by the buildup of a specific oxidant (H 2 O 2 ) in the cytosol of yeast cells upon the production of recombinant proteins. At the center of this inhibitory mechanism lies the protein kinase Gcn2. By removing Gcn2 we observed a doubling of recombinant protein productivity in addition to reduced H 2 O 2 levels in the cytosol. By this study we want to raise awareness of this inhibitory mechanism in eukaryotic cells to further improve protein production and contribute to the development of novel protein-based therapeutic strategies.