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Canan Tari - One of the best experts on this subject based on the ideXlab platform.
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Microparticle-enhanced polygalacturonase production by wild type Aspergillus sojae
3 Biotech, 2017Co-Authors: Ercan Karahalil, Canan Tari, Fadime Demirel, Ezgi Evcan, Mustafa Germeç, Irfan TurhanAbstract:Polygalacturonases (PGs), an important industrial enzyme group classified under depolymerases, catalyze the hydrolytic cleavage of the polygalacturonic acid chain through the introduction of water across the oxygen bridge. In order to produce and increase the concentration of this enzyme group in fermentation processes, a new approach called microparticle cultivation, a promising and remarkable method, has been used. The aim of this study was to increase the PG activity of Aspergillus sojae using aluminum oxide (Al_2O_3) as microparticles in shake flask fermentation medium. Results indicated that the highest PG activity of 34.55 ± 0.5 U/ml was achieved with the addition of 20 g/L of Al_2O_3 while the lowest activity of 15.20 ± 0.2 U/mL was obtained in the presence of 0.1 g/L of Al_2O_3. In fermentation without microparticles as control, the activity was 15.64 ± 3.3 U/mL. Results showed that the maximum PG activity was 2.2-fold higher than control. Additionally, smaller pellets formed with the addition of Al_2O_3 where the lowest pellet diameter was 955.1 µm when 10 g/L of the microparticle was used. Also, it was noticed that biomass concentration gradually increased with increasing microparticle concentration in the fermentation media. Consequently, the PG activity was significantly increased in microparticle-enhanced shake flask fermentation. In fact, these promising preliminary data can be of significance to improve the enzyme activity in large-scale bioreactors.
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partial purification of a polygalacturonase from a new Aspergillus sojae mutant and its application in grape mash maceration
International Journal of Food Science and Technology, 2017Co-Authors: Canan Tari, Marco A Matagomez, Semanur Yildiz, Marco RitopalomaresAbstract:Summary The use of polygalacturonase (PG) preparations in winemaking promotes the release of phenolic compounds. A PG from a new source, Aspergillus sojae mutant, was semi-purified and tested for grape mash maceration. Crude extract (CE), a commercial pectinase, and two high PG activity semi-purified preparations, FI and FII, were applied for maceration at PG activity of 3.5 U g−1 of grape for 46 h. Enzyme-assisted maceration significantly (P < 0.05) increased the total phenolic content from 255.8 to 916.3 ± 5.2, 5732.9 ± 9.9, 563.4 ± 6.7 and 620.6 ± 18.4 mg L−1 for CE, commercial pectinase, FI and FII, respectively. The content of individual phenolics such as gallic, protocatechuic, chlorogenic and p-coumaric acids was improved. Principal component and hierarchical clustering analyses suggested that CE has a better performance upon the release of phenols. Semi-purified preparations acted similar to commercial pectinase. These findings open an opportunity for the potential use of PG from the mutant strain as an alternative macerating enzyme.
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control of agitation rate and aeration for enhanced polygalacturonase production in submerged fermentation by Aspergillus sojae using agro industrial wastes
Journal of Chemical Technology & Biotechnology, 2017Co-Authors: Dante Fratebianchi, Juan Manuel Crespo, Canan Tari, Sebastian Fernando CavalittoAbstract:BACKGROUND The koji mold Aspergillus sojae, an industrially important microorganism, can produce high levels of pectinases utilizing agro-industrial wastes. This study introduces apricot and peach pomace, two agro-industrial wastes barely considered as raw material for the generation of value-added products, and focuses on its utilization together with orange peel for polygalacturonase production in submerged cultures using A. sojae. RESULTS A Doehlert response surface methodology design conducted in shake flasks and applied individually with these three by-products led to 60–80 U mL−1 polygalacturonase activity. In bioreactor studies performed with a mixture of apricot pomace and orange peel, by fixing stirrer speed to 600 rpm and cascading airflow to the dissolved oxygen tension up to 1.7 vvm, oxygen limitation problems were overcome and polygalacturonase activity values of 380 U mL−1 were achieved. CONCLUSION A simple and efficient strategy to minimize oxygen limitation with the lowest possible shear stress is provided for stirred-tank bioreactors working with highly viscous broths, so as to ultimately enhance microbial enzyme production. The polygalacturonase activity yields obtained in our study are among the highest reported in the literature. © 2016 Society of Chemical Industry
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effect of physicochemical parameters on the polygalacturonase of an Aspergillus sojae mutant using wheat bran an agro industrial waste via solid state fermentation
Journal of the Science of Food and Agriculture, 2016Co-Authors: Hande Demir, Canan TariAbstract:BACKGROUND Polygalacturonases (PGs) are valuable enzymes of the food industry; therefore it is of great importance to discover new and GRAS PG-producing microbial strains. In this study, PG enzyme produced from a high PG activity producer mutant Aspergillus sojae using wheat bran at the flask scale under pre-optimized conditions of solid-state fermentation (SSF) was biochemically characterized. RESULTS The crude PG enzyme showed optimum activity in the pH range 4.0-5.0 and was stable in the pH range 3.0-7.0. The optimum temperature for the PG was 40 °C and it retained 99% of its activity at 50 °C. The mutant A. sojae PG could preserve more than 50% of its stability between 25 and 50 °C, both for 30 and 60 min, and was found to be stable in the presence of most of the tested compounds and metal ions. The inactivation energy (Ed ) was determined as 125.3 kJ mol(-1) . The enthalpy (ΔH*), free energy (ΔG*) and entropy (ΔS*) of inactivation were found to be stable with increasing temperature. CONCLUSION The mutant A. sojae PG could be suitable for the clarification (depectinization) of orange and grape juices and wine. © 2015 Society of Chemical Industry.
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utilization of orange peel a food industrial waste in the production of exo polygalacturonase by pellet forming Aspergillus sojae
Bioprocess and Biosystems Engineering, 2015Co-Authors: Ali Oguz Buyukkileci, Marcelo Fernandez Lahore, Canan TariAbstract:The production of exo-polygalacturonase (exo-PG) from orange peel (OP), a food industrial waste, using Aspergillus sojae was studied in submerged culture. A simple, low-cost, industrially significant medium formulation, composed of only OP and (NH4)2SO4 (AS) was developed. At an inoculum size of 2.8 × 103 spores/mL, growth was in the form of pellets, which provided better mixing of the culture broth and higher exo-PG activity. These pellets were successfully used as an inoculum for bioreactors and 173.0 U/mL exo-PG was produced. Fed-batch cultivation further enhanced the exo-PG activity to 244.0 U/mL in 127.5 h. The final morphology in the form of pellets is significant to industrial fermentation easing the subsequent downstream processing. Furthermore, the low pH trend obtained during this fermentation serves an advantage to fungal fermentations prone to contamination problems. As a result, an economical exo-PG production process was defined utilizing a food industrial by-product and producing high amount of enzyme.
Sebastian Fernando Cavalitto - One of the best experts on this subject based on the ideXlab platform.
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harnessing soybean hulls for improved polygalacturonase production by Aspergillus sojae through fine tuning of ambient ph
Journal of Chemical Technology & Biotechnology, 2018Co-Authors: Dante Fratebianchi, Marina Alejandra Acosta, Sebastian Fernando CavalittoAbstract:BACKGROUND Soybean hulls result from the processing of the bean for producing oil and protein products. This by-product generated massively in America has virtually no commercial value, so substantial effort is being paid into its exploitation for generating value-added goods. This work evaluates soybean hulls as inducer of the production of pectinolytic enzymes, through optimization studies regarding polygalacturonase production by Aspergillus sojae in submerged cultures. RESULTS A 2-fold improvement in polygalacturonase yield was found by varying the initial pH of the culture in a very narrow acid pH range (2.40-2.80). The optimized fermentation process was successfully transferred to stirred-tank bioreactors in terms of volumetric productivity, and final polygalacturonase yields were 42 U/ml and 1.39 U/g soybean hulls, which are among the highest reported with this by-product. Morphological characterization of A. sojae during cultivation showed that the fungus mainly developed in dispersed mycelia at initial pH of 2.40-2.80 whilst, conversely, fungal pellets predominated in cultures performed at initial pH of 5.40. CONCLUSION High enzyme titers are possibly connected to the formation of dispersed mycelia, as well as to acid-induced expression of the respective gene/s. We foresee this data will be helpful regarding the production of fungal pectinases or other acid-induced enzymes.
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purification and biochemical and kinetic properties of an endo polygalacturonase from the industrial fungus Aspergillus sojae
Journal of Molecular Microbiology and Biotechnology, 2017Co-Authors: Dante Fratebianchi, Ivana Alejandra Cavello, Sebastian Fernando CavalittoAbstract:An endo-polygalacturonase secreted by Aspergillus sojae was characterized after being purified to homogeneity from submerged cultures with orange peel as the sole carbon source by gel filtration and ion-exchange chromatographies. According to SDS-PAGE and analytical isoelectric focusing analyses, the enzyme presents a molecular weight of 47 kDa and pI value of 4.2. This enzyme exhibits considerable stability under highly acidic to neutral conditions (pH 1.5-6.5) and presents a half-life of 2 h at 50°C. Besides its activity towards pectin and polygalacturonic acid, the enzyme displays pectin-releasing activity, acting best in a pH range of 3.3-5.0. Thin-layer chromatographic analysis revealed that tri-galacturonate is the main enzymatic end product of polygalacturonic acid hydrolysis, indicating that it is an endo-polygalacturonase. The enzyme exhibits Michaelis-Menten kinetics, with KM and VMAX values of 0.134 mg/mL and 9.6 µmol/mg/min, respectively, and remained stable and active in the presence of SO2, ethanol, and various cations assayed except Hg2+.
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control of agitation rate and aeration for enhanced polygalacturonase production in submerged fermentation by Aspergillus sojae using agro industrial wastes
Journal of Chemical Technology & Biotechnology, 2017Co-Authors: Dante Fratebianchi, Juan Manuel Crespo, Canan Tari, Sebastian Fernando CavalittoAbstract:BACKGROUND The koji mold Aspergillus sojae, an industrially important microorganism, can produce high levels of pectinases utilizing agro-industrial wastes. This study introduces apricot and peach pomace, two agro-industrial wastes barely considered as raw material for the generation of value-added products, and focuses on its utilization together with orange peel for polygalacturonase production in submerged cultures using A. sojae. RESULTS A Doehlert response surface methodology design conducted in shake flasks and applied individually with these three by-products led to 60–80 U mL−1 polygalacturonase activity. In bioreactor studies performed with a mixture of apricot pomace and orange peel, by fixing stirrer speed to 600 rpm and cascading airflow to the dissolved oxygen tension up to 1.7 vvm, oxygen limitation problems were overcome and polygalacturonase activity values of 380 U mL−1 were achieved. CONCLUSION A simple and efficient strategy to minimize oxygen limitation with the lowest possible shear stress is provided for stirred-tank bioreactors working with highly viscous broths, so as to ultimately enhance microbial enzyme production. The polygalacturonase activity yields obtained in our study are among the highest reported in the literature. © 2016 Society of Chemical Industry
Yasuji Koyama - One of the best experts on this subject based on the ideXlab platform.
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identification of the glutaminase genes of Aspergillus sojae involved in glutamate production during soy sauce fermentation
Bioscience Biotechnology and Biochemistry, 2013Co-Authors: Kotaro Ito, Yasuji Koyama, Yoshiki HanyaAbstract:Glutaminase, an enzyme that catalyzes the conversion of L-glutamine to L-glutamate, enhances the umami taste in soy sauce. The Aspergillus sojae genome contains 10 glutaminase genes. In this study, we estimated that approximately 60% of the glutamate in soy sauce is produced through the glutaminase reaction. To determine which glutaminase is involved in soy sauce glutamate production, we prepared soy sauces using single and multiple glutaminase gene disruptants of A. sojae. The glutamate concentration in soy sauce prepared using the ΔgahA-ΔgahB-ΔggtA-Δgls disruptant was approximately 60% lower than that in the control strain, whereas it was decreased by approximately 20–30% in the ΔgahA-ΔgahB disruptant. However, the glutamate concentration was unchanged in the soy sauces prepared using the ΔgahA-ΔggtA-Δgls and ΔgahB-ΔggtA-Δgls disruptants. These results indicate that four glutaminases are involved in glutamate production in soy sauce, and that the peptidoglutaminase activities of GahA and GahB increase t...
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purification and characterization of a glutaminase enzyme accounting for the majority of glutaminase activity in Aspergillus sojae under solid state culture
Applied Microbiology and Biotechnology, 2013Co-Authors: Kotaro Ito, Yoshiki Hanya, Yasuji KoyamaAbstract:Glutaminase, an enzyme that hydrolyzes l-glutamine to l-glutamate, plays an important role in the production of fermented foods by enhancing the umami taste. In this study, we found ten glutaminase genes in the Aspergillus sojae genome by conducting a BLAST search of the characterized glutaminase sequence. We subsequently constructed glutaminase gene disruptants. The glutaminase activity of the gahB disruptant was decreased by approximately 90 % in A. sojae and Aspergillus oryzae, indicating that this enzyme (GahB) accounted for the majority of the glutaminase activity in Aspergillus species. Subsequently, GahB protein was purified from the AsgahB-overexpressing transformant and characterized. The molecular mass was estimated to be approximately 110 and 259 kDa by SDS-PAGE and gel filtration chromatography, respectively, indicating that the native form of AsGahB was a dimer. The optimal pH was 9.0, and the optimal temperature was 50 °C. Analysis of substrate specificity revealed that AsGahB had peptidoglutaminase-asparaginase activity, similar to AsGahA, but preferred free l-glutamine to free l-asparagine, C-terminal glutaminyl, and asparaginyl residues in peptides.
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gene cloning purification and characterization of a novel peptidoglutaminase asparaginase from Aspergillus sojae
Applied and Environmental Microbiology, 2012Co-Authors: Kotaro Ito, Kenichiro Matsushima, Yasuji KoyamaAbstract:Glutaminase is an enzyme that catalyzes the hydrolysis of l-glutamine to l-glutamate, and it plays an important role in the production of fermented foods by enhancing the umami taste. By using the genome sequence and expressed sequence tag data available for Aspergillus oryzae RIB40, we cloned a novel glutaminase gene (AsgahA) from Aspergillus sojae, which was similar to a previously described gene encoding a salt-tolerant, thermostable glutaminase of Cryptococcus nodaensis (CnGahA). The structural gene was 1,929 bp in length without introns and encoded a glutaminase, AsGahA, which shared 36% identity with CnGahA. The introduction of multiple copies of AsgahA into A. oryzae RIB40 resulted in the overexpression of glutaminase activity. AsGahA was subsequently purified from the overexpressing transformant and characterized. While AsGahA was located at the cell surface in submerged culture, it was secreted extracellularly in solid-state culture. The molecular mass of AsGahA was estimated to be 67 kDa and 135 kDa by SDS-PAGE and gel filtration chromatography, respectively, indicating that the native form of AsGahA was a dimer. The optimal pH of the enzyme was 9.5, and its optimal temperature was 50°C in sodium phosphate buffer (pH 7.0). Analysis of substrate specificity revealed that AsGahA deamidated not only free l-glutamine and l-asparagine but also C-terminal glutaminyl or asparaginyl residues in peptides. Collectively, our results indicate that AsGahA is a novel peptidoglutaminase-asparaginase. Moreover, this is the first report to describe the gene cloning and purification of a peptidoglutaminase-asparaginase.
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draft genome sequencing and comparative analysis of Aspergillus sojae nbrc4239
DNA Research, 2011Co-Authors: Atsushi Sato, Yasuji Koyama, Tadashi Takahashi, Kenshiro Oshima, Hideki Noguchi, Masahiro Ogawa, Tetsuya Oguma, Takehiko Itoh, Masahira Hattori, Yoshiki HanyaAbstract:We conducted genome sequencing of the filamentous fungus Aspergillus sojae NBRC4239 isolated from the koji used to prepare Japanese soy sauce. We used the 454 pyrosequencing technology and investigated the genome with respect to enzymes and secondary metabolites in comparison with other Aspergilli sequenced. Assembly of 454 reads generated a non-redundant sequence of 39.5-Mb possessing 13 033 putative genes and 65 scaffolds composed of 557 contigs. Of the 2847 open reading frames with Pfam domain scores of >150 found in A. sojae NBRC4239, 81.7% had a high degree of similarity with the genes of A. oryzae. Comparative analysis identified serine carboxypeptidase and aspartic protease genes unique to A. sojae NBRC4239. While A. oryzae possessed three copies of a-amyalse gene, A. sojae NBRC4239 possessed only a single copy. Comparison of 56 gene clusters for secondary metabolites between A. sojae NBRC4239 and A. oryzae revealed that 24 clusters were conserved, whereas 32 clusters differed between them that included a deletion of 18 508 bp containing mfs1, mao1, dmaT, and pks-nrps for the cyclopiazonic acid (CPA) biosynthesis, explaining the no productivity of CPA in A. sojae. The A. sojae NBRC4239 genome data will be useful to characterize functional features of the koji moulds used in Japanese industries.
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Understanding nonaflatoxigenicity of Aspergillus sojae: a windfall of aflatoxin biosynthesis research
Applied Microbiology and Biotechnology, 2007Co-Authors: Perng-kuang Chang, Yasuji Koyama, Tadashi Takahashi, Kenichiro Matsushima, Keietsu Abe, Deepak Bhatnagar, Gwo-fang Yuan, Thomas E. ClevelandAbstract:Aspergillus section Flavi includes aflatoxin-producing and nonproducing fungi. Aspergillus sojae is unable to produce aflatoxins and is generally recognized as safe for food fermentation. However, because of its taxonomical relatedness to aflatoxin-producing Aspergillus parasiticus and A. flavus , it is necessary to decipher the underlying mechanisms for its inability to produce aflatoxins. This review addresses the relationship between A. sojae and A. parasiticus and the advances that have been made in aflatoxin biosynthesis research, especially with regard to gene structure, genome organization, and gene regulation in A. parasiticus and A. flavus and how this has been used to assure the safety of A. sojae as an organism for food fermentation. The lack of aflatoxin-producing ability of A. sojae results primarily from an early termination point mutation in the pathway-specific aflR regulatory gene, which causes the truncation of the transcriptional activation domain of AflR and the abolishment of interaction between AflR and the AflJ co-activator. Both are required for gene expression. In addition, a defect in the polyketide synthase gene also contributes to its nonaflatoxigenicity.
Nihan Gogus - One of the best experts on this subject based on the ideXlab platform.
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evaluation of orange peel an industrial waste for the production of Aspergillus sojae polygalacturonase considering both morphology and rheology effects
Turkish Journal of Biology, 2014Co-Authors: Nihan Gogus, Canan Tari, Sevcan Unluturk, Hande Demir, Bengi Hakguder Taze, Marcelo Fernandez LahoreAbstract:Orange peel is an agroindustrial waste rich in pectin and known to be an inducer for pectinase production. The use of this low-cost substrate for the production of an industrially important enzyme, polygalacturonase (PG), can be an alternative way to turn this waste into a value-added product, contributing to the reduction of environmental waste disposal problems. Enzyme productions by fungal microorganisms are affected by environmental and nutritional factors, demanding the determination of optimum conditions for maximum enzyme production with the desired fungal morphology and broth rheology. Therefore, complex and additional carbon sources were optimized with respect to PG production by Aspergillus sojae using statistical approaches. Effect of pH, another significant parameter affecting the rheology and morphology of the strain, was investigated in the serial bioreactor system using the optimized medium composition. Highest PG enzyme yield and productivity together with the maximum PG enzyme production (93.48 U/mL) were obtained under uncontrolled pH conditions. Under these conditions, morphologically, pellet sizes exhibited a normal distribution ranging between 0.5-1.0 mm and 1.0-1.5 mm, and rheological measurements revealed that fermentation broths showed non-Newtonian flow. The low pH trend observed during the course of the fermentation was another important positive outcome for industrial fermentations, prone to contamination problems.
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optimization of the process parameters for the utilization of orange peel to produce polygalacturonase by solid state fermentation from an Aspergillus sojae mutant strain
Turkish Journal of Biology, 2012Co-Authors: Hande Demir, Canan Tari, Doreen Heerd, Nihan Gogus, Marcelo Fernandez LahoreAbstract:Th e eff ect of orange peel concentration, HCl concentration, incubation time and temperature, and inoculum size on the spore count and activity of polygalacturonase (PG) enzyme produced from Aspergillus sojae M3 by solid- state fermentation was screened using 2 k factorial design. Orange peel and HCl concentrations and incubation time were signifi cant factors aff ecting the responses. Optimum conditions favoring both PG and spore production from Aspergillus sojae M3 were determined as 2% orange peel and 50 mM HCl concentrations at 22 °C and 4.3 days of incubation. An overlay plot was constructed for use as a practical chart for production of high enzyme activity (>35.0 U/g substrate) and spore count (9.0 × 10 8 to 2.0 × 10 9 spore/mL) by superimposing the contours of PG activity and spore count responses. Th e accuracy and reliability of the constructed models on the responses was validated with the maximum calculated error rate between the predicted and actual activities at 14.1% and 22.4%, respectively.
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effect of substrate concentration on the polygalacturonase production by Aspergillus sojae mutant strain in serial bioreactor system
2nd International ISEKI_FOOD Conference, 2011Co-Authors: Nihan Gogus, Canan Tari, Hande Demir, Bengi Hakguder, Marcelo Fernandez LahoreAbstract:Pectinases include a number of related enzymes involved in breaking down pectic substances. This degradative process plays an important role in food technology, due to reduction in time of filtration and to the volume increase, and juice clarification. This process leads to a more stable and concentrated product. The main sources of the pectinolytic complex enzymes are yeast, bacteria and a large variety of filamentous fungi, for which the most relevant ones are Aspergillus . Polygalacturonase (PG) attracts the most attention among the family of pectinolytic enzymes. As the pectinases are induced enzymes it is necessary to supplement the culture medium with pectin or with raw materials rich in pectin, like orange peel. In this study dry orange peels which are rich in pectin, cellulose and hemicellulose as a fermentation substrate were used. Submerged fermentation (SmF) is generally used for the production of industrially important enzymes. It is known that different from the shake flask studies, bioreactors are a better controlled environment where the effect of common factors such as pH, agitation and dissolved oxygen tension can be observed and controlled. In the proposed study it was aimed to investigate the effect of substrate concentration on the PG activity and biomass using Aspergillus sojae M5/6 mutant strain. With this perspective, using Sartorius BIOSTAT Qplus-6 serial bioreactor fermentations with six different substrate concentrations (60, 40, 20, 15, 10, 5 g/l) at 750 ml scale, were performed. In conclusion maximum PG activity was achieved at 40 g/l orange peel concentration (108,866 U/ml) at the end of the fermentation. Besides as the substrate concentration was increased PG activity and biomass was increased, too. Maximum product yield (Y P/S ) was calculated for 20 and 15 g/l substrate concentrations as 14,47 and 14,35 Uactivity/mgsubstrate, respectively. Additionally maximum productivity was obtained at 40 g/l concentration as 0,75 U/ml.h.
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biochemical and thermal characterization of crude exo polygalacturonase produced by Aspergillus sojae
Food Chemistry, 2008Co-Authors: Canan Tari, Nergiz Dogan, Nihan GogusAbstract:Abstract Crude exo-polygalacturonase enzyme (produced by Aspergillus sojae ), significant for industrial processes, was characterized with respect to its biochemical and thermal properties. The optimum pH and temperature for maximum crude exo-polygalacturonase activity were pH 5 and 55 °C, respectively. It retained 60–70% of its activity over a broad pH range and 80% of its initial activity at 65 °C for 1 h. The thermal stability study indicated an inactivation energy of E d = 152 kJ mol −1 . The half lives at 75 and 85 °C were estimated as 3.6 and 1.02 h, respectively. Thermodynamic parameters, Δ H ∗ , Δ S ∗ and Δ G ∗ , were determined as a function of temperature. The kinetic constants K m and V max , using polygalacturonic acid as substrate, were determined as 0.424 g l −1 and 80 μmol min −1 , respectively. SDS-PAGE profiling revealed three major bands with molecular weights of 36, 53 and 68 kDa. This enzyme can be considered as a potential candidate in various applications of waste treatment, in food, paper and textile industries.
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optimization of biomass pellet size and polygalacturonase production by Aspergillus sojae atcc 20235 using response surface methodology
Enzyme and Microbial Technology, 2007Co-Authors: Canan Tari, Nihan Gogus, Figen TokatliAbstract:A two-step optimization procedure using central composite design with four factors (concentrations of maltrin and corn steep liquor (CSL), agitation speed and inoculation ratio) was used in order to investigate the effect of these parameters on the polygalacturonase (PG) enzyme activity, mycelia growth (biomass) and morphology (pellet size) of Aspergillus sojae ATCC 20235. According to the results of response surface methodology (RSM), initial concentrations of maltrin and CSL and agitation speed were significant (p < 0.05) on both PG enzyme production and biomass formation. As a result of this optimization, maximum PG activity (13.5 U/ml) was achievable at high maltrin (120 g/l), at low CSL (0 g/l), high agitation speed (350 rpm) and high inoculation ratio (2 × 107 total spore). Similarly, maximum biomass (26 g/l) could be obtained under the same conditions with only the difference for higher level of CSL requirement. The diameter of pellets in all optimization experiments ranged between 0.05 and 0.76 cm. The second optimization step improved the PG activity by 74% and the biomass by 40%.
Dante Fratebianchi - One of the best experts on this subject based on the ideXlab platform.
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harnessing soybean hulls for improved polygalacturonase production by Aspergillus sojae through fine tuning of ambient ph
Journal of Chemical Technology & Biotechnology, 2018Co-Authors: Dante Fratebianchi, Marina Alejandra Acosta, Sebastian Fernando CavalittoAbstract:BACKGROUND Soybean hulls result from the processing of the bean for producing oil and protein products. This by-product generated massively in America has virtually no commercial value, so substantial effort is being paid into its exploitation for generating value-added goods. This work evaluates soybean hulls as inducer of the production of pectinolytic enzymes, through optimization studies regarding polygalacturonase production by Aspergillus sojae in submerged cultures. RESULTS A 2-fold improvement in polygalacturonase yield was found by varying the initial pH of the culture in a very narrow acid pH range (2.40-2.80). The optimized fermentation process was successfully transferred to stirred-tank bioreactors in terms of volumetric productivity, and final polygalacturonase yields were 42 U/ml and 1.39 U/g soybean hulls, which are among the highest reported with this by-product. Morphological characterization of A. sojae during cultivation showed that the fungus mainly developed in dispersed mycelia at initial pH of 2.40-2.80 whilst, conversely, fungal pellets predominated in cultures performed at initial pH of 5.40. CONCLUSION High enzyme titers are possibly connected to the formation of dispersed mycelia, as well as to acid-induced expression of the respective gene/s. We foresee this data will be helpful regarding the production of fungal pectinases or other acid-induced enzymes.
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purification and biochemical and kinetic properties of an endo polygalacturonase from the industrial fungus Aspergillus sojae
Journal of Molecular Microbiology and Biotechnology, 2017Co-Authors: Dante Fratebianchi, Ivana Alejandra Cavello, Sebastian Fernando CavalittoAbstract:An endo-polygalacturonase secreted by Aspergillus sojae was characterized after being purified to homogeneity from submerged cultures with orange peel as the sole carbon source by gel filtration and ion-exchange chromatographies. According to SDS-PAGE and analytical isoelectric focusing analyses, the enzyme presents a molecular weight of 47 kDa and pI value of 4.2. This enzyme exhibits considerable stability under highly acidic to neutral conditions (pH 1.5-6.5) and presents a half-life of 2 h at 50°C. Besides its activity towards pectin and polygalacturonic acid, the enzyme displays pectin-releasing activity, acting best in a pH range of 3.3-5.0. Thin-layer chromatographic analysis revealed that tri-galacturonate is the main enzymatic end product of polygalacturonic acid hydrolysis, indicating that it is an endo-polygalacturonase. The enzyme exhibits Michaelis-Menten kinetics, with KM and VMAX values of 0.134 mg/mL and 9.6 µmol/mg/min, respectively, and remained stable and active in the presence of SO2, ethanol, and various cations assayed except Hg2+.
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control of agitation rate and aeration for enhanced polygalacturonase production in submerged fermentation by Aspergillus sojae using agro industrial wastes
Journal of Chemical Technology & Biotechnology, 2017Co-Authors: Dante Fratebianchi, Juan Manuel Crespo, Canan Tari, Sebastian Fernando CavalittoAbstract:BACKGROUND The koji mold Aspergillus sojae, an industrially important microorganism, can produce high levels of pectinases utilizing agro-industrial wastes. This study introduces apricot and peach pomace, two agro-industrial wastes barely considered as raw material for the generation of value-added products, and focuses on its utilization together with orange peel for polygalacturonase production in submerged cultures using A. sojae. RESULTS A Doehlert response surface methodology design conducted in shake flasks and applied individually with these three by-products led to 60–80 U mL−1 polygalacturonase activity. In bioreactor studies performed with a mixture of apricot pomace and orange peel, by fixing stirrer speed to 600 rpm and cascading airflow to the dissolved oxygen tension up to 1.7 vvm, oxygen limitation problems were overcome and polygalacturonase activity values of 380 U mL−1 were achieved. CONCLUSION A simple and efficient strategy to minimize oxygen limitation with the lowest possible shear stress is provided for stirred-tank bioreactors working with highly viscous broths, so as to ultimately enhance microbial enzyme production. The polygalacturonase activity yields obtained in our study are among the highest reported in the literature. © 2016 Society of Chemical Industry
