Amylase from Bacillus sp. Produced by Solid State Fermentation Using Cassava Bagasse as Starch Source

Abstract Amylases are enzymes involved in starch hydrolysis, generating the most diverse products, such as maltose, glucose and dextrins. This work aimed the study of the production of amylolytic enzymes via solid-state fermentation (SSF) using "crueira", an essentially starchy cassava residue, as substrate-support and Bacillus sp. as microorganism. For the implementation of the experimental part, a Central Composite Design (CCD) with three variables (initial moisture, pH and temperature) was made. Each test was examined at 24, 48 and 72 hours by the method of starch dextrinizing activity. The optimum production conditions were 60% initial moisture, pH 6 and 37 °C. The maximum yield was 437.76 U/g in 72 hours of fermentation. The optimum temperature of enzyme performance was 65 °C. The pH optimum range was 4 to 6. The Co2 +, Ca2 + and K+ ions positively influenced the activity of enzymes and the Fe2+ ion had no effect on enzymatic activity. On the other hand, the ions Hg2+, Zn2+, Cu2+, Mn2+ and Mg2+ adversely influenced enzymatic activity. Therefore, producing amylases from Bacillus sp. and using crueira as a substrate is possible.

Simultaneous production of amylases and proteases by Bacillus subtilis in brewery wastes

ABSTRACT The simultaneous production of amylase (AA) and protease (PA) activity by Bacillus subtilis UO-01 in brewery wastes was studied by combining the response surface methodology with the kinetic study of the process. The optimum conditions (T = 36.0 °C and pH = 6.8) for high biomass production (0.92 g/L) were similar to the conditions (T = 36.8 °C and pH = 6.6) for high AA synthesis (9.26 EU/mL). However, the maximum PA level (9.77 EU/mL) was obtained at pH 7.1 and 37.8 °C. Under these conditions, a considerably high reduction (between 69.9 and 77.8%) of the initial chemical oxygen demand of the waste was achieved. In verification experiments under the optimized conditions for production of each enzyme, the AA and PA obtained after 15 h of incubation were, respectively, 9.35 and 9.87 EU/mL. By using the Luedeking and Piret model, both enzymes were classified as growth-associated metabolites. Protease production delay seemed to be related to the consumption of non-protein and protein nitrogen. These results indicate that the brewery waste could be successfully used for a high scale production of amylases and proteases at a low cost.

Isolation, Identification and Optimization of Culture Conditions of Bacillus sp. Strain PM1 for Alkalo-thermostable Amylase Production.

Aims: To isolate and optimize the culture conditions for thermo stable and alkaline amylase production from bacteria. Study Design: Optimization of different physiological and nutritional parameters for amylase production and kinetic studies of amylase. Place and Duration of Study: Soil Samples: Herbal garden of Amity University Haryana, Gurgaon (Manesar), India, between April 2012 and September 2012. Methodology: Amylolytic isolates were selected by flooding the nutrient agar plates containing 2% starch with Lugol solution. Isolates were selected on the basis of higher ratio of clear zone to colony size and grown in nutrient broth containing 2% starch. The level of amylase was detected in the culture filtrate. The selected isolate showing maximum amylase production was identified on the basis of 16S rDNA amplification. Results: An Alkalo-thermostable amylase producing bacterial isolate from soil was identified as Bacillus sp. strain PM1 on the basis of 16S rRNA. It yielded 3.5 U/ml of amylase in medium containing (%) starch 2.0, beef extract 0.5, NaCl 0.5 at 50ºC, pH 7.0 at 180 rpm after 72 h. The optimum pH and temperature for amylase activity was 8.0 and 50°C, respectively. The enzyme exhibited 67% activity after 60 minute incubation at 50ºC. At pH 8.0, the enzyme retained 78% activity after 4 h. Conclusion: The properties of the isolated enzyme are adequate for its use in starch processing and baking industry.

Viabilidade, confirmação taxonômica e detecção enzimática de espécies de Acremonium preservadas sob óleo mineral na Coleção de Culturas University Recife Mycology / Viability, taxonomic confirmation and enzymatic detection of Acremonium species preserved under mineral oil in the URM Culture Collection

Enzimas hidrolíticas secretadas por fungos têm um papel importante na patogenicidade das infecções. Objetivando avaliar a atividade enzimática foram testados 31 isolados de Acremonium mantidos na Coleção de Culturas University Recife Mycology. Fragmentos das culturas foram transferidos para caldo glicosado para reativação e posterior crescimento em meio ágar batata dextrose, para verificar viabilidade, pureza e confirmação taxonômica pela observação das características macroscópicas e microscópicas. Para detecção enzimática foram utilizados substratos de caseína do leite e gelatina para protease, amido para amilase e lecitina de soja para fosfolipase. Das 31 culturas, 26 (83,9 por cento) mantiveram-se viáveis e 24 (92,3 por cento) foram confirmadas taxonomicamente. Das 24 culturas, 12 (50 por cento) apresentaram atividade proteásica, duas (16,7 por cento) em caseína do leite, uma (8,3 por cento) em gelatina e nove (75 por cento) em ambos os substratos; 16 (66,7 por cento) degradaram amido. Nenhuma cultura apresentou atividade fosfolipásica. Conclui-se que espécies de Acremonium são capazes de produzir enzimas envolvidas na patogenicidade das infecções fúngicas.

Hydrolytic enzymes secreted by fungi play an important role in the pathogenesis of infection. With the aim of evaluating the enzymatic activity, 31 isolates of Acremonium stored in the University of Recife Mycology (URM) Culture Collection were tested. Culture fragments were transferred to glycoside broth for reactivation and further growth in potato dextrose agar medium in order to investigate viability and purity and to confirm the taxonomy through observing the macroscopic and microscopic characteristics. To detect enzymes, milk casein and gelatin were used as substrates for proteinase, starch for amylase and soy lecithin for phospholipase. Among the 31 cultures, 26 (83.9 percent) remained viable and 24 (92.3 percent) were confirmed taxonomically. Out of these 24 cultures, 12 (50 percent) presented proteinase activity, of which two (16.7 percent) were on milk casein, one (8.3 percent) on gelatin and nine (75 percent) on both substrates; 16 (66.7 percent) degraded starch. None of the cultures presented phospholipase activity. It was concluded that Acremonium species are able to produce enzymes that are involved in the pathogenicity of fungal infections.

Growth and amylase production by Aspergillus oryzae during solid state fermentation using banana waste as substrate.

Aspergillus oryzae, isolated from the starch rich litter soil, produced amylase when banana fruit stalk was used as substrate in a solid state fermentation system. The effects of incubation period, pH, temperature and various carbon sources on the production of amylase were studied. A maximum yield of 380U/mg was recorded on 96th hour of incubation. The amylase is tolerant to wide range of initial culture pH values (3 to 8) and temperature (25 to 35 degrees C), with an optimum pH of 5 and temperature of 35 degrees C. In SSF addition of starch (1%) increased the amylase production, when compared with other carbon sources used.

Amylase induction during seed germination in isoproturon susceptible and resistant biotypes of Phalaris minor Retz.

Isoproturon resistant biotype of P. minor germinates early, shows higher germination percentage and faster rate of growth as compared to the susceptible biotype. Higher amylase activity is observed in the initial hours of imbibition in the resistant biotype. In the susceptible biotype it is activated at a much later stage.

Production of industrially important enzymes by some actinomycetes producing antifungal compounds.

Seventeen strains of actinomycetes antagonistic to yeast and moulds have been tested for their ability to produce amylase, lipase, gelatinase and caseinase using solid media containing starch, Tween-20, gelatin and skimmed milk, respectively, Enzyme producing potential of test strains is expressed in ternis of relative enzyme activity (REA). Actinomycetes strain Streptomyces somaliensis GS 1242 and Streptomyces sampsonii GS 1322 showed higher amylase production (REA 6.5) while maximum lipase activity was noted in Streptomyces strain SAP 1089 (REA 7.0). Gelatinase activity was noted higher is S. sampsonii GS 1322 (REA 9.6) and S. somaliensis GS 1242 (REA 8.8). Enzyme producing potential of these strains has been discussed in terms of their industrial significance.

Production of amylases and proteases by wild-type and mutant strains of Metarhizium anisopliae Var. anisopliae

Linhagens parentais, mutantes auxotróficos e morfológicos do fungo entomopatogênico Metarhizium anisopliae var. anisopliae foram analisados para secreçäo de amilases e proteases, em meio sólido. Näo foram observadas diferenças na secreçäo de proteases e amilases entre colônias monospóricas, originadas da mesma linhagem. Algumas das linhagens com marcas auxotróficas e morfológicas secretaram mais proteases e amilases, quando comparadas aos seus respectivos parentais, demonstrando efeito pleiotrópico para uma ou mais mutaçöes nestas linhagens. Nos dois meios usados para atividade de amilases, um contendo somente amido e outro contendo amido e glicose, linhagens parentais demonstraram menor secreçäo de amilases em presença de glicose em comparaçäo com as linhagens mutantes usadas

Production of extracellular proteases and amylases by some acidophilic and alkalophilic bacteria.

Twelve bacteria were isolated from two effluent sources of Shaw-Wallace Gelatins, Jabalpur. Six bacteria from dicalcium phosphate plant effluent (pH-5) and six from main drain of the factory (pH-10) were isolated. Two facultatively acidophilic and two facultatively alkalophilic bacteria were selected and tentatively identified as Plesiomonas shigelloides, Aeromonas hydrophilla, Klebsiella pneumoniae and Staphylococcus saprophyticus respectively. Acidic amylases were produced in higher amounts on 4th day of incubation by Plesiomonas shigelloides and on 6th day by Aeromonas hydrophilla. Alkaline amylases were produced in higher amounts on 4th day of incubation by Klebsiella pneumoniae and on 8th day by Staphylococcus saprophyticus in vitro.

Effect of fungicides on the production of amylase by Rhizopus oryzae.

Seven fungicides (Brassicol), captan, Dithane M-45, Fytolan, Parasan, Sulfex and Thiram) were tested for their effect on the production of amylase enzyme by Rhizopus oryzae. The activity of amylase was determined by cup-plate method. All the fungicides were found inhibitory for the synthesis of amylase. Out of these, Parasan was the most effective causing hundred percent inhibition at 0.025% concentration. No amylase production was recorded when Thiram, Fytolan, Brassicol, Captan and Dithane M-45 were used at 0.5, 1.5, 2.0, 2.0, and 2.0% concentration respectively. Sulfex was found to be less effective, as hundred percent inhibition of amylase production has not been recorded even at 3.0% concentration.

Amylolytic activity of humicola sp

The thermophylic and cellulolytic fungus Humicola sp. secretes amylases in the liquid culture medium. This activity is induced by starch, maltose and cellobiose. Glucose impairs accumlation of amylolitic activity in the culture medium. The enzyme hydrolyzes starch, maltose and pullulan to glucose as the endproduct

Extra cellular hydrolytic enzyme secretion in Bacillus thuringiensis H14 & B. sphaericus & their significance in media design.

Level of extracellular proteolytic and amylolytic enzyme production was determined to assess the ability of these strains in utilizing complex carbon and nitrogen sources. Protease secretion reached maximum at around 12th h of growth in case of all the three B. thuringiensis strains and declined sharply thereafter. In B. sphaericus strains the protease level gradually increased and reached maximum at around 24th h of growth. Amylase activity was not detectable in the culture supernatants of B. sphaericus strains whereas all the three B. thuringiensis strains tested showed significant amount of amylase activity.

Gastric metastasis and ectopic amylase production in bronchogenic carcinoma.

A case of small cell anaplastic carcinoma of bronchus is described. Primary tumour had metastasized in the stomach and there was production of amylase by the tumour evidenced by raised serum amylase and pleural fluid amylase.

Synthesis and contents of pancreatic exportable enzymes: effect of oleic acid intraduodenal administration

Ratas Sprague-Dawley fueron estimuladas intraduadernalmente con ácido oleico y sacrificadas 20, 40, 60 y 80 m luego de la instilación. En todos los grupos se administró, 10m antes del sacrifício, 50 uCi de 3H-fenilalanina (3H-F) intraperitoneal. Se cuantificaron los niveles intrapancreáticos de Am, Chtg, Tg y Li y la incorporación de 3H-F en las proteínas secretoras. A los 40 m de la administración de ácido oleico se registró la máxima estimulación de ácido oleico se registró la máxima estimulación para Chtg (45%), Tg (38%) y Li (23%) por sobre los valores controles, no modificándose la Am. Todos los valores enzimáticos cayeron por debajo de los controles a los 60 y 80 minutos. La incorporación 3H-F fue máxima a los 40 m decayendo a tiempos más prolongados. En el presente trabajo demostramos que la administración intraduodenal de ácido oleico genera un aumento no paralelo en el nivel intrapancreático de algunas enzimas exportables, y que dichos valores caen a partir de los 40 m post-instilación, siendo la síntesis proteica afectada de manera similar