Microbial Chemistry

An alkaline protease was purified from a halophilic and thermotolerant potent alkaline protease-producing strain Streptomyces pseudogrisiolus NRC-15 using ammonium sulphate precipitation and Sephadex G-100 column chromatography. The enzyme was purified to 77.24-folds with a yield of 91.8% and the specific activity was 112 U/mg of protein. The protease showed a single band on SDS-PAGE with its molecular mass at 20 kDa and exhibited a maximum relative activity of 100% using casein as a substrate and. The enzyme had an optimum pH of 9.5 and displayed optimum activity at 50°C. The enzyme activity was completely inhibited by the serine protease inhibitor PMSF, suggesting the presence of serine residue in the active site. The enzyme activity was increased by the metal ions Ca 2+ , Co 2+ , K + and Mg 2+ . The enzyme significantly enhanced the removal of stains when used with wheel detergent, indicating the potential of the enzyme for using as a laundry detergent additive to improve the performance of heavy-duty laundry detergent.

Collections of 13 yeast strains isolated from different sources of marine water (Ras Al Bar, Alexandria and Marsa Matruh) were screened for β-glucosidase production. Rhodotorula glutinis was found to be the best source of this enzyme production. The effect of medium composition and incubation conditions for fermentation process were established. The maximum β-glucosidase production (270units/ml) was occurred after 24h of incubation at 30°C on optimized medium containing cellobiose (2%) as a sole source of carbon and 0.5% (NH 4) 2 SO 4 as a nitrogen source. The enzyme was found to be produced intracellular by the strain under study, therefore, physical and chemical treatments were applied to release the enzyme.The maximum enzyme release was observed when grinding with sterile sea sand (250 units/ml). The enzyme was partially purified by ammonium sulfate precipitation and the recovery was 75% of the total enzyme activity which affords 3.02 fold purification. The hydrolytic activity of...

Background: Biomass produced as a byproduct from the β-mannanase production process by Aspergillus tamarii NRC 3was evaluated as a biosorbent for the removal and recovery of some heavy metal ions. Results: Under optimal conditions, the isolated strain recorded the highest β-mannanase activity (31.88 Uml −1). Thus, the biomass produced from mannanase production process as a byproduct was evaluated as a biosorbent for the removal and recovery of some heavy metal ions from aqueous solutions and an industrial wastewater. The fungal biomass was found to be efficient for the removal of Cu +2 and some heavy metal ions. The biosorption process of copper(II) by Aspergillus tamarii NRC 3 biomass was affected by changing of time, temperature, pH, metal ions concentration, the presence of some heavy metals, and biomass concentration. The rate of Cu +2 uptake from Cu +2 solution proceeded rapidly, and it appeared to be virtually complete during the initial 5 min (92%); the maximum uptake of Cu +2 appeared at 30°C, pH 5, and biomass concentration 5 g w/w. On the other hand, the fungal biomass was to remove considerable proportion of Pb 2+ , Co +2 , Ni 2+ , Fe +3 , and Cr 3+ in addition to Cu 2+. The uptake of Cu +2 by pretreated biomass was studied. Recovery of the sorbed metal ions by desorbing agents and the potential reuse of the regenerated biomass in metal ions uptake (reloading) were evaluated. Conclusions: Aspergillus tamarii NRC 3 biomass seems to be quite feasible in the removal of heavy metal ions especially Cu +2 from aqueous solutions.

Among 20 fungal strains grown on the solid glucose salts medium, 10 Aspergillus spp. have a strong red purple color in plates and the appearance of red color was taken as the production of kojic acid. The fungi in the plates showing the highest color intensities were selected by visual observation followed by a second selection using a250 ml Erlenmeyer flasks fermentation. Of the ten Aspergillus strains grown statically or shaken on a fermentation medium (pH, 4.0) for 18 days at 28°C, A. oryzae var.effusus NRC14, A. flavus NRC13, A. tamarii NRC18 and A. parasiticus were found to be the most highly active organisms for kojic acid production, whereas in two cases,( static and shaken cultures), kojic acid produced by A. oryzae var.effusus NRC14 was higher than that produced by A. flavus NRC13, A. tamarii NRC18 and A. parasiticus. On the other hand, a static culture of A. oryzae var.effusus NRC14 produced kojic acid higher than a shaken culture did (42.0, 25.5g kojic acid/L) at 150 rpm....

Microbial protease represents the most important industrial enzymes, which have an active role in biotechnological processes. The objective of this study was to isolate new strain of Streptomyces that produce proteolytic enzymes with novel properties and the development of the low-cost medium. An alkaline protease producer strain NRC-15 was isolated from Egyptian soil sample. The cultural, morphological, physiological characters and chemotaxonomic evidence strongly indicated that the NRC-15 strain represents a novel species of the genus Streptomyces, hence the name Strptomyces pseudogrisiolus NRC-15. The culture conditions for higher protease production by NRC-15 were optimized with respect to carbon and nitrogen sources, metal ions, pH and temperature. Maximum protease production was obtained in the medium supplemented with 1% glucose, 1% yeast extract, 6% NaCl and 100 µmol/L of Tween 20, initial pH 9.0 at 50 ºC for 96 h. The current results confirm that for this strain, a great ability to produce alkaline proteases, which supports the use of applications in industry.

Background Although microbial fuel cells (MFCs) represent a promising technology for capturing renewable energy from wastewater, their scaling-up is significantly limited by a slow-rate cathodic oxygen reduction reaction (ORR) and the development of a resilient anodic microbial community. In this study, mixed transition metal oxides of nickel and copper (Ni and Cu), supported on a graphene (G) (NiO–CuO/G) electrocatalyst, were synthesized and tested as a cost-effective cathode for ORR in MFCs. Electrochemical measurements of electrocatalyst were conducted using a rotating disk electrode (RDE) and linear sweep voltammetry (LSV) in a neutral electrolyte, and compared with a benchmark Pt/C catalyst. Furthermore, the long-term performance of the as-synthesized electrocatalyst was evaluated in a single-chamber MFC by measuring organic matter removal and polarization behavior. The successful enrichment of electroactive biofilm was also monitored using transmission electron microscopy and ...

Immobilization technique using different matrix (agar, polyacrylamide and gelatin) to enhancement protease inhibitor activity and specific activity from Streptomyces lavendulae (Egyptian isolate from marine) which identified by 16SrRNA with accession number AB795940. The best specific activity of protease inhibitor using numerous factors plus UV radiation was 1055 u/mg however the specific activity without any factor 234 u/mg. Comparing free cells data with immobilization data great different observed, specific activity of protease inhibitor when Streptomyces lavendulae immobilized on gelatin after three days less cell leakage 0.7 and specific activity was 660 u/mg. when gelatin matrix exposure different doses of gamma radiation then immobilized bacterial cells specific activity reached to 1900 u/mg at 8 KGy. The gelatin immobilized cells of Streptomyces lavendulae can be proposed as an effective biocatalyst for repeated usage for maximum production of protease inhibitor.

Research focused on isolation and characterization on new strain of Streptomyces pseudogrisiolus from Egyptian soil have proteinases which possesses activity (MCE) identified by 16S rRNA with universal primer and submit in genbank with accession number AB739622. In this work, study optimizes the extraction of milk-clotting enzyme (MCE) according to its adequacy to be employed as substitute of rennin. The organism grew well in medium 3 after 4days of incubation at temperatures of 35° ±2, pH 6.0. Cultural and environmental conditions for milk clotting enzyme production by Streptomyces were studied. Glucose (3%, w/v) and ammonium sulphate (0.2%w/v), were found to be the best C and N sources for milk clotting enzyme production respectively. Partially purified and characterized. MCE was obtained by fractional precipitation with ammonium sulphate, followed by the chromatography of the most active fraction on Sephadex G-100 with 20.39 purification fold and recovery about 80%. The maximum enzyme activity was at pH (6.5) and 45ºC. The clotting activity of the purified enzyme was stimulated with increasing CaCl 2 concentration up to 10mM. However, a gradual reduction of the activity was observed by increasing NaCl concentration between 4-8%. Whereas, Cu +2 and Zn +2 significant inhibition it. Interestingly MnCl 2 and MgSO 4 were found to have stimulatory effect, which has been rarely reported. Also study includes some kinetics calculation about MCE. Gene control MCE expression was detected by specific primer design using DNA star program, sequence illustrate gene found clt a and clt b, and submitted in genbank as new sequence with accession number AB739621. The milk clotting enzyme (MCE) obtained from S. pseudogrisiolus has potential as calf rennet substitutes in dairy products and it can be used in lypholyzed form as tablets in pharmaceutical products for patients with digestive system.

Allelopathy is the chemical interaction between plants (including microorganisms), and reported as an ecological process structuring plant communities. Bioavailability and persistence of allelochemicals are much affecting by soil microbes. In the same time we find (according to the different studies) growth inhibition in filter paper bioassay enriched with an allelochemical or an extract, such results have not been verifiable in more natural settings with plants growing in soil. We examined in vitro the allelopathic potential of rice straw extract incubated 40 and 70 days with different isolates of Fusarium moniliforme (F1) and F. oxysporium (F2, and F3), besides a combination treatment including them all against seedling root and shoot growth of Triticum aestivum, Raphanus sativus, Lactuca sativa and Eruca sativa as bioassay. Rice straw was also incubated under natural conditions of soil (for 40 & 70 days) and the leachates were used in bioassay-plants reaction experiments. A quali...

Present Study target on alkaline Protease production which have multi- medical application in pharmaceuticals products. Streptomyces flavogriseus isolated and identified by 16SrRNA, it have accessions number AB723783.1. Alkaline Protease produced by free and immobilized cells. Entrapment method of immobilization used with gelatin, polyacrylamide, and agar matrices. The novelty in this work, data showed that specific activity of alkaline protease with gelatin entrapped immobilization produce 1042 U/mg ; followed by agar 998.3 U/mg, and polyacrylamide, 626.7.3. The study suggests the gelatin, a better matrix than others. It is possible used matrix immobilized with Streptomyces flavogriseus cells several times with highly activity the decreasing in activity in each time very low. With different gamma radiation doses exposure to gelatin matrix the maximum value of protease specific activity 22152 U/mg protein at dose 10 kGy. On the otherwise submerged fermentation using free cells only ...

Collections of 13 yeast strains isolated from different sources of marine water (Ras Al Bar, Alexandria and Marsa Matruh) were screened for β-glucosidase production. Rhodotorula glutinis was found to be the best source of this enzyme production. The effect of medium composition and incubation conditions for fermentation process were established. The maximum β-glucosidase production (270units/ml) was occurred after 24h of incubation at 30°C on optimized medium containing cellobiose (2%) as a sole source of carbon and 0.5% (NH 4) 2 SO 4 as a nitrogen source. The enzyme was found to be produced intracellular by the strain under study, therefore, physical and chemical treatments were applied to release the enzyme.The maximum enzyme release was observed when grinding with sterile sea sand (250 units/ml). The enzyme was partially purified by ammonium sulfate precipitation and the recovery was 75% of the total enzyme activity which affords 3.02 fold purification. The hydrolytic activity of...

The objective of this study is the production of an extra cellular alkaline protease by a new local isolate which was characterized and identified by 16S rRNA as Streptomyces griseus sub sp griseus and it was submitted in gene bank with accession number AB723782. The culture conditions for higher protease production by Streptomyces griseus were optimized with respect to carbon source, nitrogen source and pH. Maximum protease production was obtained in medium no. 5 supplemented with 3% glucose as a carbon source, 0.4 % NaNO 3 as a nitrogen source, the optimum pH was 8 at 30°C and the highest production was obtained after 6 days of incubation. Wheat bran proved as the best food waste for protease production (9208.5 U/mg protein) compared to other wastes, and finally the optimum gamma radiation was (1.6 kGy) led to increase the specific activity to 14450.5 U/mg protein. The protease profile of the isolate shows its potential as a good source for industrial application.

Protease inhibitorstrypsin (STI1, Streptomyces trypsin inhibitor 1) has been identified, purified by ammonium sulfate precipitation and Sephadex G-100 gel filtration. SDS-PAGE of protease inhibitor showed molecular weight of approximately 10 KDa. PCR product (~1615 bp) ofsti1gene was cloned in expression vectorpACYC177/ET3dand transformed inEscherichia coliJM109.Protease inhibitorstrypsin was purified and used as antivirus against Coxsackievirus B3 (CVB3). CVB3 is one of the major causative agents of chronic, subacute, acute, and fulminant myocarditis as well as pancreatitis and aseptic meningitis. It has been reported that more than 50% of human myocarditis is associated with CVB3 infection.

The aim of the present study is to purify L-methioninase fromCandida tropicalis34.19-fold with 27.98% recovery after ion exchange chromatography followed by gel filtration. The purified enzyme revealed a single band on SDS-PAGE gel with a molecular weight of 46 KDa. Its optimum temperature was 45 to 55 and thermal stability was 55°C for 15 min. The enzyme had optimum pH at 6.5 and stability at a pH range of 5.5 to 7.0 for 24 hr. The maximum activity was observed with substrate concentration of 30 µM and Km was 0.5 mM. The enzyme was strongly inhibited by Cd+2and Cu+2while it was enhanced by Na+, Ni+2, and Mg+2at 10 mM while Ca+2had slight activation at 20 mM. In addition, the potential application of the L-methioninase as an anticancer agent against various types of tumor cell lines is discussed.

Background: Biomass produced as a byproduct from the β-mannanase production process by Aspergillus tamarii NRC 3was evaluated as a biosorbent for the removal and recovery of some heavy metal ions. Results: Under optimal conditions, the isolated strain recorded the highest β-mannanase activity (31.88 Uml −1). Thus, the biomass produced from mannanase production process as a byproduct was evaluated as a biosorbent for the removal and recovery of some heavy metal ions from aqueous solutions and an industrial wastewater. The fungal biomass was found to be efficient for the removal of Cu +2 and some heavy metal ions. The biosorption process of copper(II) by Aspergillus tamarii NRC 3 biomass was affected by changing of time, temperature, pH, metal ions concentration, the presence of some heavy metals, and biomass concentration. The rate of Cu +2 uptake from Cu +2 solution proceeded rapidly, and it appeared to be virtually complete during the initial 5 min (92%); the maximum uptake of Cu +2 appeared at 30°C, pH 5, and biomass concentration 5 g w/w. On the other hand, the fungal biomass was to remove considerable proportion of Pb 2+ , Co +2 , Ni 2+ , Fe +3 , and Cr 3+ in addition to Cu 2+. The uptake of Cu +2 by pretreated biomass was studied. Recovery of the sorbed metal ions by desorbing agents and the potential reuse of the regenerated biomass in metal ions uptake (reloading) were evaluated. Conclusions: Aspergillus tamarii NRC 3 biomass seems to be quite feasible in the removal of heavy metal ions especially Cu +2 from aqueous solutions.

Background: Several species of streptomycetes, saprophytic bacteria found widely distributed in soil, water and plants, produce bioactive compounds such as intra and extracellular hydrolases including lytic enzymes which reflecting on their importance in the biological control of insects and parasites. This study assessed the in vitro effects of Streptomyces tyrosinase, produced from Streptomyces spp. isolated from Egyptian soil, on animal-parasitic nematode Toxocara vitulorum, in terms of egg development and adult worm’s cuticular structure, and as an alternative strategy to alleviate this infection. Methods: This study was conducted at the National Research Centre, Egypt in 2018. Five different concentrations of tyrosinase, ranged from 1%-30% were tested against the development of T. vitulorum eggs. The concentration induced the highest inhibitory activity was tested against adult T. vitulorum cuticle, which is essential for the protective and nutritive functions. The results were...

The present study target is alkaline protease production by Streptomyces flavogriseus which isolated and identified by 16S rRNA and was deposited in the genebank sequence database under accessions number AB723783.1. Alkaline protease was produced by free and immobilized cells. Entrapment method of immobilization was used with agar, gelatin and polyacrylamide matrices. Data showed that gelatin entrapped cells gave the maximum specific activity (1042.0 U/mg protein); followed by agar (930.8 U/mg protein) and polyacrylamide (690.7 U/ mg protein) after 48 h of incubation. The reusability test proved that the gelatin entrapped cells can be reused for 4 cycles with retention of about 75% of enzyme specific activity followed by polyacrylamide and agar; the reusability was for 2 and 4 cycles, respectively. After gamma radiation exposure of gelatin entrapped cells, the maximum value of protease specific activity reached 19152 U/mg protein at dose 12 kGy. On the other hand, studies on submerg...

Background Grey mould caused by Botrytis cinerea of cucumber immature fruits is the most epidemic fungal diseases causing significantly losses of fruits during development on cucumber plant in winter season. Material and methods Crude and nanoemulsions of clove, black seed, lemon and orange essential oils at (5000 ppm) and also different formulations of clove, black seed and lemon oils nanoemulsions as single or in combination were tested against mycelial linear growth of B. cinerea (MF996363) as well sclerotial formation and pathological activity on the development of germinated seeds of cucumber. Results Essential oils nanoemulsion formulation of clove + black seed (2:1) at 5000 ppm was the best formulation significantly reduced mycelia linear growth of B. cinerea by 61.0% followed by 28% formulation (1:1) and (1:2). In addition, nanoemulsions formulations of clove, black seed essential oils (2:1) were highly reduced the count of sclerotial formation of B. cinerea and completely (...

An alkaline protease was purified from a halophilic and thermotolerant potent alkaline protease-producing strain Streptomyces pseudogrisiolus NRC-15 using ammonium sulphate precipitation and Sephadex G-100 column chromatography. The enzyme was purified to 77.24-folds with a yield of 91.8% and the specific activity was 112 U/mg of protein. The protease showed a single band on SDS-PAGE with its molecular mass at 20 kDa and exhibited a maximum relative activity of 100% using casein as a substrate and. The enzyme had an optimum pH of 9.5 and displayed optimum activity at 50 degrees C. The enzyme activity was completely inhibited by the serine protease inhibitor PMSF, suggesting the presence of serine residue in the active site. The enzyme activity was increased by the metal ions Ca2+, Co2+, K+ and Mg2+. The enzyme significantly enhanced the removal of stains when used with wheel detergent, indicating the potential of the enzyme for using as a laundry detergent additive to improve the pe...