Enzyme Production Ability Research Articles

Screening of intestinal probiotics and the effects of feeding probiotics on the growth, immune, digestive enzyme activity and intestinal flora of Litopenaeus vannamei

In this experiment, 426 strains were isolated from the intestinal tract of Litopenaeus vannamei, and 11 strains showed strong digestive enzyme production activity and antagonistic effect against common bacterial pathogens of shrimp. After hemolysis activity test and drug sensitivity test, 2 candidate probiotics with good bacteriostatic activity, strong enzyme production ability and relatively sensitive to antibiotics were screened out, and were identified by 16s rDNA molecular identification and Biolog-System as Enterobacter hominis (E3) and lactobacillus (L3). First, the biological characteristics of 2 candidate probiotics were studied. The optimum growth conditions of E3: temperature, 30 °C; pH, 8.0; NaCl, 2.5%; bovine bile salt, 0.15%; and the optimum growth conditions of L3: temperature, 40 °C; pH, 6.0; NaCl, 0.5%; bovine bile salt, 0.0015%. Secondly, a 28-day feeding experiment was conducted using probiotic concentration of 107 CFU g−1 to determine the changes of the activities of blood related immune enzymes (SOD, PPO, ACP, POD, CAT, LZM) and intestinal digestive enzymes (NP, AL, LPS) during the feeding process of shrimp, the results showed that during the course of feeding, the activities of immune enzyme and digestive enzyme of shrimp fed with probiotics showed an increasing trend, and the growth rate of body weight of shrimp was higher than that of control group. After feeding, the cumulative mortality of probiotics groups were significantly lower than that of the control group after WSSV infection. And the mid-gut of L. vannamei was observed by electron microscope, the results showed that the intestinal mucosa was tight and the epithelium cells showed an active secretory state in probiotics group. Finally, the intestinal microbial communities of shrimp were compared and analyzed by using Biolog-ECO method in the later period of feeding, the results showed: compared with the control group, the average color change rate of the experimental group fed with probiotics increased significantly, indicating that probiotics enhanced the intestinal microorganism activity; The ability of intestinal microorganism to utilize carbon source was significantly enhanced in the experimental group, which indicated that the digestive enzyme secreted by probiotics could improve the digestion and absorption rate of prawn feed, thus promoting the rapid growth of shrimp; The Shannon index, Simpson index and McIntosh index of probiotics groups showed significant difference in 1st and 5th days, but tended to be the same in the 10th day, the results showed that probiotics could maintain in L. vannamei intestines at least 5 days.

Effects of temperature on extracellular hydrolase enzymes from soil microfungi

Soil microbes play important roles in global carbon and nutrient cycling. Soil microfungi are generally amongst the most important contributors. They produce various extracellular hydrolase enzymes that break down the complex organic molecules in the soil into simpler form. In this study, we investigated patterns of amylase and cellulase (which are responsible for breaking down starch and cellulose, respectively) relative activity (RA) on solid media at different culture temperatures in fungal strains from Arctic, Antarctic and tropical soils. Fungal isolates from all three regions were inoculated onto R2A media supplemented with starch for amylase and carboxymethylcellulose and trypan blue for cellulase screening. The isolates were then incubated at 4, 10, 15, 20, 25, 30, 35 and 40 °C and examined for activity after 5 and 10 days, for tropical and polar isolates, respectively. The data obtained indicate that the polar fungal strains exhibited similar patterns of amylase and cellulase RA. Both Arctic and Antarctic fungi showed highest RA for amylase and cellulase at 35 °C, while colony growth was maximised at 15 °C. Colony growth and RA of the polar isolates were negatively correlated suggesting that, as temperatures increase, the cells become stressed and have fewer resources available to invest in growth. Unlike polar isolates, tropical isolates did not exhibit any trend of colony growth with temperature, rather having idiosyncratic patterns in each isolate. The low enzyme production and RA levels in the tropical strains may suggest both a low ability to respond to temperature variation in their natural thermally stable tropical habitats, as well as a level of thermal stress limiting their enzyme production ability.

In vitro evaluation of candidate Bacillus spp. for animal feed.

The aim of this study was to select aerobic spore-formers for animal feed based on their in vitro probiotic potential, including their enzyme-producing ability and safety assessment. Seven isolates out of 187 spore-forming bacteria were selected for their ability to produce cellulase (89.21-1668.32 U/ml), xylanase (1399.68-4351.10 U/ml), and phytase (2.72-28.70 U/ml). Among seven isolates, five had activities towards a broad range of p-nitrophenyl esters with acyl chain lengths from C2 to C12. The probiotic properties of all selected isolates varied with respect to their acid and bile salt tolerance under simulated gastrointestinal tract (GIT) conditions, and their adherence ability to human intestinal cell lines (Caco-2 and HT-29). The safety assessment revealed that the isolate CM40 was not cytotoxic to Caco-2 and HT-29, did not exhibit hemolytic activity, carried no enterotoxin or emetic toxin genes, and was susceptible to ten antibiotics, including six key antibiotics (chloramphenicol, erythromycin, gentamicin, tetracycline, streptomycin, and kanamycin) as recommended by the European Food Safety Authority (EFSA). Co-incubation of isolate CM40 with enteric bacteria (Salmonella Typhi, Salmonella Enteritidis 1781, and Escherichia coli) demonstrated that CM40 significantly decreased the number of pathogens (about 30-48%) adhering to Caco-2 and HT-29 (P < 0.05). Analysis of gene encoding 16S rRNA, gyrase A (gyrA) and the cheA histidine kinase revealed that CM40 belongs to Bacillus subtilis. On the basis of probiotic properties and basic safety aspects, the B. subtilis strain CM40 was found to possess desirable in vitro probiotic properties, and may be a potential candidate for supplementation of animal feed.

Identification and Efficiency of a Nitrogen-fixing Endophytic Actinobacterial Strain from Sugarcane

Sugarcane production in China mostly uses a high amount of chemical nitrogen (N) as compared to the average use of other countries. A microbe-based alternative may help to manage this problem in ecofriendly manner. Therefore, the present study focused on the characterization of a sugarcane growth-promoting nitrogen-fixing endophytic actinobacteria. The actinobacterial strain WZS021 was identified on the basis of morphology, biology and sequencing of 16S rRNA, gyrB and acdS gene amplification. The nitrogenase activity was determined with acetylene reduction assay and effects of different nutritional conditions were evaluated, and 15N isotope dilution method was used to determine the N-fixing capability under sugarcane seedlings in the pot. The strain was identified as Streptomyces chartreusis, by three genes (16S rRNA, gyrB and acdS), but nifH gene showed similarity with Bacillus and these results showed nitrogen fixation ability and 1-aminocyclopropane-1-carboxylate deaminase enzyme production ability at the molecular level. Moreover, strain WZS021 showed maximum nitrogenase enzyme production under pH 8 and sustained at higher temperature. Different carbon sources have less influence on nitrogenase activity. However, small amounts of N source can stimulate the nitrogenase activity, but after certain concentration it starts to inhibit the enzyme activity. Next to this, in the 15N isotope pot assay strain WZS021 significantly enhanced the dry weight of shoot and root by 20 and 75%, and N content increased by 30 and 36%, respectively. The N-fixation percentage of strain WZS021 under sugarcane seedlings was 9.16%. Overall, N-fixing ability of Streptomyces chartreusis strain WZS021 was characterized, and sugarcane biomass enhancement and substantial %Ndfa indicated that the promising strain could be applied in sugarcane as a growth promoter.

Effect of Different Mutation Methods on Xylanase Production Ability by Aspergillus Niger

Based on the Aspergillus niger AS3.350, the mutants were brewed through inducing five treatments.The effect of different mutation methods on enzyme production ability of Xylanase was disscused and obtainedmutant strain of Xylanase production high ability. The results indicate that: the mutagenesis of UV irradiation and fast neutron irradiation was sensitive to increasing Xylanase activity. UV irradiation seted at 256 nm wavelength,the vertical distance was 20cm,exposure time was 90s, obtained mutant UV256-90 of the emzyme activity was 170.79U/ml,it was 2.18 times of the original strain.The rate 0.0025Gy / s of fast neutron irradiation and vertical distance 12cm,the optimal dose is 0.75Gy, screened out the mutant Fn5-2, the activity is 134.77U/ml,its 1.72 times of the original strain. Mutagenesis by diethyl sulfate, sodium nitrite, and nitrosoguanidine, enzyme activityof the optimal mutant strains were recently starting strain of 1.51 times, 1.46 times, and 1.38 times.Two physical mutagenic methods include ultraviolet mutagenesis and fast neutron irradiation were more sensitive on producing xylanase based on aspergillus Niger.

Studies on an Indigenous Probiotic (&lt;i&gt;Shewanella algae&lt;/i&gt;) Isolated from Healthy Shrimp Intestine

An indigenous probiotic ( Shewanella algae ) previously isolated from healthy shrimp ( Penaeus monodon ) intestine and found to protect Clarias gariepinus from Aeromonas hydrophila infection was screened for antibiotic susceptibility, enzymatic activities and abiotic stress tolerance. Optimization of the cultivation conditions and identification of its bioactive compounds were also carried out. The antibiotic susceptibility was performed by disk diffusion method. A total of fifteen antibiotic discs were employed. Qualitative screening for extracellular enzyme-producing ability was done using skim milk agar, starch agar, egg yolk agar, gelatin agar and cellulose agar for protease, amylase, lipase, gelatinase and cellulase activities respectively. The optimal conditions for growth of the isolate were assessed by growing on nutrient broth at various pH, temperature and salinity levels. Cell growth was estimated by standard plate count. The probiotic was screened for antimicrobial activity by agar well diffusion assay against the pathogen ( Aeromonas hydrophila ) using cell-free supernatant (crude extract) and methanolic extract. The methanolic extract was analyzed using gas chromatography- mass spectrometry (GC-MS). The isolate showed a wide range of environmental tolerance with pH (4-11), temperature (28-40℃) and salinity (0-100 ppt). The optimum conditions for cell growth were pH 6-8, temperature 37℃ and salinity 5-15 ppt. The isolate was susceptible to all tested antibiotics which supported the ideal probiotic characteristic. The isolate was capable of producing extracellular enzymes such as protease, amylase, lipase and gelatinase which could improve feed digestibility and feed utilization. The crude and methanolic extracts were active against the pathogen with inhibition zones of 13.0 ± 0.02 mm and 18.0 ± 0.03 mm respectively. Two bioactive compounds (Tromethamine and Pyrrolo [1, 2-a] pyrazine-1, 4-dione, hexahydro-) were identified. These results suggest that the Shewanella algae could probably be used as a potential probiotic in aquafeed formulation and a good source of novel drug development.

Autochthonous Gut Bacteria in Two Indian Air-breathing Fish, Climbing Perch (Anabas testudineus) and Walking Catfish (Clarias batrachus): Mode of Association, Identification and Enzyme Producing Ability.

Scanning electron microscopy (SEM) was used to define the location of epithelium-associated bacteria in the gastrointestinal (GI) tract of two Indian air-breathing fish, the climbing perch (Anabas testudineus) and walking catfish (Clarias batrachus). The SEM examination revealed substantial numbers of rod shaped bacterial cells associated with the microvillus brush borders of enterocytes in proximal (PI) and distal regions (DI) of the GI tract of both the fish species. Ten (two each from the PI and DI of climbing perch and three each from the PI and DI of walking catfish) isolated bacterial strains were evaluated for extracellular protease, amylase and cellulase production quantitatively. All the bacterial strains exhibited high cellulolytic activity compared to amylolytic and proteolytic activites. Only two strains, CBH6 and CBH7, isolated from the DI of walking catfish exhibited high proteolytic activity. Maximum cellulase activity was exhibited by the strain, CBF2, isolated from the PI of climbing perch. Six most promising enzyme-producing adherent bacterial strains were identified by 16S rDNA gene sequence analysis. The strain ATH1 (isolated from climbing perch) showed high similarity fo Bacillus amyloliquefaciens whereas, the remaining five strains (isolated from walking catfish) were most closely related to Bacillus licheniformis.

Metabolic characteristics and effects on &lt;i&gt;Microcystis&lt;/i&gt; growth of attached bacteria of &lt;i&gt;Microcystis &lt;/i&gt;blooms

为研究附生细菌对蓝藻水华细胞微环境的影响,从微囊藻水华中分离出34株藻附生细菌,研究其产酶能力. 对氮代谢相关的酶活性测定结果显示:其中13株菌具有蛋白酶活性,5株菌具有较强的氨氮脱除能力,1株菌具有强烈的硝酸还原酶活性;4株菌具有较高的碱性磷酸酶活性;11株菌具有产脂酶的活性. 通过菌和藻共同培养的方法,观察菌对微囊藻生长的影响,结果显示,有11株菌表现出较明显的促藻生长作用,占总筛选菌株总数的32%;有3株菌表现出较显著的抑制藻生长的作用,占总筛选菌株总数的9.9%. 附生菌产酶能力与生长相关性的分析显示,促进微囊藻生长的附生菌都具有蛋白酶或脂酶活性.;To study the effects of attached bacteria on the microenvironment of Microcystis cells of blooms, thirty-four attached bacterial strains were isolated from Microcystis colonies sampled from 4 different bloom samples, and enzyme-producing abilities of the isolated strains were measured. Measuring of nitrogen metabolism enzymes showed that 13 strains could hydrolyze casein, 5 strains could effectively remove ammonia, and 1 strain had a strong nitrate reductase activity. Measuring of phosphorus metabolism enzymes displayed 4 strains with phosphates' activity. The measurement also showed 11 strains with esterase activity. To evaluate the effects of the bacterial strains on the growth of Microcystis, each isolated strain was co-cultured with Microcystis. The results showed that 11 strains could obviously promote Microcystis growth,took up 32% of the total isolates; three strains obviously inhibited the growth, took up 9.9% of the total isolates. Relationship of enzyme production ability and effects on Microcystis growth revealed that the growth-promoting strains were case in aseor esterase-producing strains.

Characterization of Cell Wall Degrading Enzymes of &lt;i&gt;Rhizoctonia solani&lt;/i&gt; AG-3 from Tobacco Target Spot

Rhizoctonia solaniKühn is a causal pathogen responsible for many types of plant disease worldwide and a major soilborne fungal pathogen that severely impairs yield and quality of tobacco worldwide. Activities, pathogenicity of the cell wall-degrading enzymes produced by theRhizoctoniasolanifrom tobacco target spot disease both in liquid medium and in tobacco tissue were studied. The result showed thatR.solanifrom tobacco can produce pectinase and cellulase both in vitro and vivo, and the activity of PG and PMG was the highest in vitro. The activity of Cx and β-glucosidase was the highest in vivo, and enzyme production ability of strong pathogenicity strains is stronger than the weak pathogenicity strains in vitro.

Optimization and strain improvement by mutation for enhanced cellulase production by Bacillus sp. (MTCC10046) isolated from cow dung

The goal was to investigate the cellulase enzyme production ability of bacterial strain C1 isolated from cow dung and identified as Bacillus sp. on the basis of 16 S rDNA sequence homology. The effects of different carbon sources like Carboxymethyl cellulose (CMC), avicel, starch, maltose, sucrose, glucose, fructose, galactose and lactose on cellulase production at varying environmental parameters of incubation period (2–10days), temperature (35–55°C), and pH (6.0–8.5) were examined. The CMC was the best carbon source for cellulase production followed by lactose in this bacterial strain. The maximum enzyme production was achieved at a temperature of 50°C by Bacillus sp. with pH of 7.0 on the 8th day of growth. The nitrogen source NH4NO3 at 0.175% was optimum for cellulase production by this bacterium. A putative mutant (C1M26) was screened from wild C1 strain after mutagenesis with N-methyl-N′-nitro-N-nitrosoguanidine (NTG) as a mutagenic agent. The mutant C1M26 produced a larger amount of cellulase in comparison to wild type C1 strain.

Enzymatic Activities of Intestinal Bacteria Isolated from Farmed &lt;i&gt;Clarias gariepinus&lt;/i&gt;

The culturable bacteria associated with the digestive tract of a freshwater cultured fish, Clarias gariepinus , and their degradative abilities were established. The spread plate method was employed for bacterial isolation. The bacterial isolates were qualitatively screened for extracellular enzyme-producing ability using milk agar, starch agar, egg yolk agar and cellulose agar for protease, amylase, lipase and cellulase activities respectively. A total of 18 bacterial isolates were identified. Bacteria of the genera Bacillus , Staphylococcus , Vibrio , Aeromonas , Pseudomonas , Lactobacillus , Escherichia , Salmonella , Enterobacter , Micrococcus and Flavobacterium were isolated from fish digestive tract at different frequencies with Bacillus predominating. Enzymatic studies indicated that the bacterial isolates possess the ability to degrade proteins, starch, lipids and cellulose. The percentage composition of enzyme-producing bacteria are - protease producing strains (72.2%), lipase producing strains (61.1%), amylase producing strains (55.6%) and cellulose producing strains (38.9%). All the isolates possessed multienzyme activity. An isolate ( Bacillus sp. B 1 ) showed activity for protease, amylase, lipase and cellulase enzymes. Therefore, the isolated indigenous multiple enzyme-producing strains can be effectively exploited for use as probiotics while formulating aquafeeds.

Biodegradation of crude oil from saline waste water using white rot fungus Phanerochaete chrysosporium

In this research, the effect of salinity on fungal growth and enzyme production ability of Phanerochaete chrysosporium was investigated. The ability of this fungus in the biodegradation of crude oil pollution in saline waste waters like Bilge water was also studied. Brine with salinities of 0, 10, 20, 30 and 40g/l were selected for cultivation of the fungus. The statistical analysis of data were also provided using F-test with significance level of 0.05 to specify whether the results are significant or not significant. The p-values revealed that salinity had not any effect on the fungal growth i.e. there was not a significant difference in fungal growths in different salinities. Also, production of MnP enzyme was not affected up to the salinity of 20g/l, but it decreased in 30 and 40g/l salinities. The effect of salinity on the LiP enzyme activity was more noticeable, and the activity of this enzyme was inhibited completely (100%) in the salinity of 40g/l. Furthermore, the effect of two surfactants, namely NAR-111-2 and Tween80, was investigated on removal efficiency of crude oil by fungus. The results revealed the positive effect of these surfactants on crude oil removal efficiency. Based on the results of this study the P. chrysoisporium is able to degrade crude oil from saline waste water adequately as a result of the appropriate enzymes activity.

Isolation and Application of Thermophilic and Psychrophilic Microorganisms in the Composting Process

Because temperature is a limited factor in compost process, we attempted to isolate and screen efficient themophilic and psychrophilic strains from composts and soils adapting to high and low temperature, respectively. Firstly, two cellulose-effectively-decomposing thermophilic bacteria, HNS39 and HNC41, were screened from 13 thermophilic compost samples and one soil sample from tropical rain forest. By addition of microbial agents with single HNS39, HNC41, or their mixture (1:1 v/v), the progress of composing was more effectively improved with HNS39. According to 16S rDNA gene sequencing and its growth characteristics, HNS39 was identified as Geobacillus sp. Meanwhile, the psychrotrophs strains of B6-38, B10-40, B10-44 isolated, could rapidly reproduce and effectively degrade organic compounds with highest enzyme activity in low temperature from 10 samples collected from sewage sludge, poultry manure and paddy soil. It was indicated that cold-adapted bacteria had a better effect on the raise of composting temperature, especially B6-38 with the highest protease activity of 46.08 U/ml. It grew fast and had good ability of enzyme production and strong organic degradation capabilities at 15 °C. By identification, Strain B6-38 was associated with Aeromonasa sp. Therefore, the research was fully shown that the isolated thermophilic and psychrophilic microorganisms had high potential of application in the composting under high and low temperature.

Enhancement of chitosanase production by cell immobilization of Gongronella sp. JG

Chitosanase production of Gongronella sp. JG cells immobilized in calcium alginate gel and polyurethane foam was compared with that of the free cells, there was a 60% increase in the enzyme yield (2429 U/L) compared to the highest yield obtained from free cells (1513 U/L). The optimal immobilization parameters (concentrations of sodium alginate, calcium chloride, bead inoculums, bead diameter, etc) for the enhanced production of chitosanase were determined as: sodium alginate 2% (w/v), 0.1 M calcium chloride, inoculum 10 mL beads to 100 mL production media and 2.7 mm bead diameter. Maximum chitosanase production was achieved with initial pH of 5.5 and temperature of 30 °C. The alginate beads had well stability, retained 85% ability of enzyme production even after 7 cycles of repeated batch fermentation. These results showed the immobilization technique was a feasible and economical method for chitosansase production by Gongronella sp. JG.

Study on the Use of Agricultural Wastes for Cellulase Production by Using Aspergillus Niger

It was the goal to investigate the Cellulase enzyme production ability of fungal strains such as Aspergillus niger against the lignocellulosic bio-waste like Rice husks, Millet husks, Maize cobs, Wheat straws and leaves at varying environmental parameters of pH, substrate concentration and dry weight and wet weight. The enzyme production was analyzed individually by Miller's modified method of Dinitrosalicylic acid ( DNSA ). In this study the high level of enzyme production was achieved at pH 3 by using Rice husks, Millet husks, Maize cobs, Wheat straws and leaves and pH 4 by using Maize cobs at varying substrate concentrations. Out of all the five used substrates rice husks gave the higher production of the cellulose enzyme. The importance of cellulase enzyme in industries cannot be over emphasized.

Carbohydrate degrading enzyme production by plant pathogenic mycelia and microsclerotia isolates of Macrophomina phaseolina through koji fermentation

The ability of hydrolytic enzyme production by two different isolates of Macrophomina phaseolina was studied on apple pomace as a substrate for solid state fermentation (SSF). Initial moisture level, temperature and fermentation period was optimized so as to achieve higher output. Among the two different isolates, microsclerotial (MphP) and mycelial (MphM), MphP was observed as a potential source of different hydrolytic enzymes as compared to MphM. MphP gave higher enzyme activities (IU/gram dry substrate (gds): filter paper cellulase (FPase) activity 196.21±16.3 (120h), carboxymethyl cellulase (CMCase) 279.34±28.25 (72h), β-glucosidase (BGL) 129.82±12.41 (96h), xylanase 2527.88±46.15 (120h), and amylase 2780.72±38.13 (96h), respectively at 70% (v/w) IML. The incubation temperature was also found to have impact on the enzyme production ability of Macrophomina strains. The higher enzyme activities were achieved (IU/gds) as follows FPase 276.13±25.02 (40°C, 120h), CMCase 278.11±24.47 (35°C, 144h), BGL 189.47±15.05 (30°C, 144h), xylanase 3845.77±43.38 (35°C, 144h) and amylase activity of 3309.45±29.22 (35°C, 120h), respectively using MphP at 70% (v/w) IML. This study reports for the first time the potential of carbohydrate degrading enzyme bioproduction by different isolates of M. phaseolina.

Characterization and Identification of Enzyme-producing Bacteria Isolated from the Digestive Tract of Bata, Labeo bata

Isolation and enumeration of aerobic bacterial flora in the foregut and hindgut regions of the gastrointestinal tract of bata, Labeo bata, were carried out on selected nutrient media following the enrichment culture technique. Four bacterial isolates were qualitatively screened on the basis of their extracellular enzyme-producing ability. The selected strains were further quantitatively assayed for amylase, cellulase, and protease activities. Amylolytic strains were present in higher densities in the foregut region, whereas the cellulolytic and proteolytic populations exhibited maximum densities in the hindgut region. Peak amylase, cellulase, and protease activities were exhibited by bacterial strains BF2 and BH4, isolated from the foregut and hindgut regions, respectively. Both the strains are Gram-positive rods, motile, and capable of forming endospores. They can tolerate a wide range of temperatures (25–42 C and 25–55 C in cases of BF2 and BH4, respectively) and pH (5–11). On the basis of phenotypic characteristics and 16S rDNA sequence analysis, the strains were identified as Bacillus licheniformis (BF2, GenBank Accession Number: EF635428) and Bacillus subtilis (BH4, GenBank Accession Number: EF032683). The study provides scope for further research on the use of the enzyme-producing fish gut bacteria in aquafeeds, especially for the larval stages.

Identification and Characterization of α-Amylase and Endoxylanase, Produced by Aspergillus Mutant Strains

ABSTRACTFilamentous fungi are widely used for the production of homologous and heterologous proteins. Recently, there has been increasing interest in Aspergillus oryzae PP and Aspergillus awamori K-1 because of its ability to produce heterologous proteins in submerged (liquid) cultures. The goal of this investigation was to determine the α-amylase and endoxylanase enzyme production ability and molecular characteristics of fungal strains Aspergillus oryzae PP and Aspergillus awamori K-1 and its mutant strains R5 and A45. The strains were cultivated in liquid culture and maximum enzyme production of parent and mutant strains was determined after 72–96 h of cultivation. Extracellulars α-amylase and endoxylanase were partially purified from the culture filtrates, using molecular sieve chromatography with Gel permeation chromatography. The molecular weight of the partial purified enzymes has been estimated to be 57 kDa for α-amylase and 31 kDa for endoxylanase on SDS-polyacrylamide gel electrophoresis. The temperature optimum of the enzyme α-amylase was 30°C, respectively for endoxylanase was 40°C and the pH optimum was 4.7 and 4.0.

Enhanced production of a thermostable mannanase by immobilized cells of Bacillus subtilis on various membranes

Cells of the thermophilic Bacillus subtilis WY34 were immobilized on various formaldehyde-activated polymer membranes and the immobilized cells were used for the production of thermostable mannanase in flasks. The results showed that polyethersulfone membranes (PES) and nylon-6 membranes were the most suitable supports for cell immobilization to produce the mannanase. Moreover, PES and nylon-6 membranes immobilized cells provided 1.78- and 1.74-fold higher mannanase activity compared to the control after 4 days of cultivation, respectively. The immobilized cells on PES and nylon-6 membranes had good stability and retained 131.5 and 114.3% of ability of enzyme production even after six cycles of repeated batch fermentation, respectively. Active cell growth was observed by scanning electron microscopy (SEM) after 16 days (four cycles) repeated batch cultivation. Therefore, the membrane-immobilized cells of B. subtilis WY34 can be proposed as an effective biocatalyst for repeated usage for production of the thermostable mannanase.

Characterization of Enzyme Endoxylanase Produced by Mutants Strains of Aspergillus Awamori K-1

ABSTRACTXylanolytic enzymes of microbial origin have received great attention due to their biotechnological utility and potential applications in a range of industrial processes but the observed effects vary depending on xylanase specificities.The goal of this investigation was to determine the endoxylanase enzyme production ability and molecular characteristics of fungal strains Aspergillus awamori K-1 and its mutant strains A59, A50, A45 and A60. The strains were cultivated in liquid culture with 1% wheat bran and 2% ground corn-cobs as inducer. Maximum enzyme production of parent strain Aspergillus awamori K-1 of 42,35 IU/ml and of mutant strains A59 - 96,91 IU/ml, A50 - 71,67 IU/ml, A45 - 74,04 IU/ml and A60 - 62,59 IU/ml was determined after 72–96 h of cultivation.Extracellular endoxylanase was partially purified from the culture filtrates of five strains, using molecular sieve chromatography with two type of Sephadex—G 50 and G100. The purification conditions were optimized and the main endoxylanase activity was determined in second fraction in case of Sephadex G100. The partial purified enzyme fractions showed the same band for all strains on SDS polyacrylamide gel electrophoresis (SDS-PAGE) with an apparent molecular weight of 32 kDa.