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1.
Thermostable phytase production by Thermoascus aurantiacus in submerged fermentation 总被引:2,自引:0,他引:2
Nampoothiri KM Tomes GJ Roopesh K Szakacs G Nagy V Soccol CR Pandey A 《Applied biochemistry and biotechnology》2004,118(1-3):205-214
Phytases act on phytic acid, an antinutrient factor present in animal feeds, and release inorganic phosphate. We optimized
the production parameters for phytase production using Thermoascus aurantiacus (TUB F 43), a thermophilic fungal culture, by submerged fermentation. A semisynthetic medium containing glucose, starch,
peptone, and minerals supplemented with 3.75% (w/v) wheat bran particles was found to be the best production medium among
the various combinations tried. Further supplementation of this medium with surfactants such as Tween-20 and Tween-80 considerably
enhanced the enzyme yield. A maximum phytase activity (468.22 U/mL) was obtained using this production medium containing 2%
(v/v) Tween-20 after 72 h of fermentation at 45°C in shake-flask cultures with a rotation of 150 rpm. Herein we present details
of a few of the process parameter optimizations. The phytase enzyme was found to be thermostable, and the optimal temperature
for phytase activity was found to be 55°C. However, 80% of the activity still remained when the temperature was shifted to
70°C. 相似文献
2.
Alejandro Colina Betzabé Sulbarán-de-Ferrer Cateryna Aiello Alexis Ferrer 《Applied biochemistry and biotechnology》2003,108(1-3):715-724
Xylanase production of Trichoderma reesei Rut C-30 was examined at different initial pH values (4.8, 5.9, and 7.0) on rice straw in shake flasks, and in a fermentor,
for the best pH condition. Enzyme performance was tested on ammonia-treated dwarf elephant grass. The maximum xylanase activities,
92 and 122 IU/mL, were obtained at pH 4.8 in the shake flasks and fermentor, respectively, in which good growth of the fungus
was observed during the first 24 h and consumption of proteins dissolved from the rice straw caused the pH to rise later to
values between 6.4 and 6.7 (optimal for xylanase production). The xylanases from T. reesei were as effective as Multifect XL, a commercial enzyme preparation, in hydrolyzing ammonia-treated elephant grass. 相似文献
3.
Shin Chul Seung Lee Joon Pyo Lee Jin Suk Park Soon Chul 《Applied biochemistry and biotechnology》2000,84(1-9):237-245
Economical production of cellulase enzyme is key for feasible bioethanol production from ligh ocellulosics using an enzyme-based
process. On-site cellulase production can be more feasible with the process of separate hydrolysis and fermentation (SHF)
than with simultaneous saccharification and fermentation, since the cost of enzyme is more important and a variety of substrates
are available for the SHF process. Cellulase production using various biomass substrates available for SHF, including paper
sludge, pretreated wood (steam exploded), and their hydrolysis residues, was investigated in shake flasks and a fermenter
for their productivities and titers. Among the newspaper sludge, office paper sludge, and steam-exploded woods treated in
various ways, the steam-exploded wood showed the best properties for substrate in cellulase production. The besttiter of 4.29
IU/mL was obtained using exploded wood of 2% (w/v) slurry in the shake flask, and the titer with the same substrate was duplicated
to about 4.30 IU/mL in a 3.7-L fermenter. Also, the yield of enzyme reached 215 1U/g of substrate or 363 IU/g of cellulose.
Despite various pretreatment attempts, newspaper and office paper substrate was inferior to the exploded-wood substrate for
cellulase production. However, hydrolysis residues of papers showed quite promising results. The hydrolysis residue of office
paper produced 2.48 IU/mL of cellulase in 7 d. Hence, the utilization of hydrolysis residues for cellulase production will
be further investigated in the future. 相似文献
4.
Koivula A Ruohonen L Wohlfahrt G Reinikainen T Teeri TT Piens K Claeyssens M Weber M Vasella A Becker D Sinnott ML Zou JY Kleywegt GJ Szardenings M Ståhlberg J Jones TA 《Journal of the American Chemical Society》2002,124(34):10015-10024
Trichoderma reesei cellobiohydrolase Cel6A is an inverting glycosidase. Structural studies have established that the tunnel-shaped active site of Cel6A contains two aspartic acids, D221 and D175, that are close to the glycosidic oxygen of the scissile bond and at hydrogen-bonding distance from each other. Here, site-directed mutagenesis, X-ray crystallography, and enzyme kinetic studies have been used to confirm the role of residue D221 as the catalytic acid. D175 is shown to affect protonation of D221 and to contribute to the electrostatic stabilization of the partial positive charge in the transition state. Structural and modeling studies suggest that the single-displacement mechanism of Cel6A may not directly involve a catalytic base. The value of (D2O)(V) of 1.16 +/- 0.14 for hydrolysis of cellotriose suggests that the large direct effect expected for proton transfer from the nucleophilic water through a water chain (Grotthus mechanism) is offset by an inverse effect arising from reversibly breaking the short, tight hydrogen bond between D221 and D175 before catalysis. 相似文献
5.
Effect of acetic acid and furfural on cellulase production of Trichoderma reesei RUT C30 总被引:4,自引:0,他引:4
Because of the high temperature applied in the steam pretreatment of lignocellulosic materials, different types of inhibiting
degradation products of saccharides and lignin, such as acetic acid and furfural, are formed. The main objective of the present
study was to examine the effect of acetic acid and furfural on the cellulase production of a filamentous fungus Trichoderma reesei RUT C30, which is known to be one of the best cellulase-producing strains. Mandels’s mineral medium, supplemented with steam-pretreated
willow as the carbon source at a concentration corresponding to 10 g/L of carbohydrate, was used. Four different concentration
levels of acetic acid (0–3.0 g/L) and furfural (0–1.2 g/L) were applied alone as well as in certain combinations. Two enzyme
activities, cellulase and β-glucosidase, were measured. The highest cellulase activity obtained after a 7-d incubation was
1.55 FPU/mL with 1.0 g/L of acetic acid and 0.8 g/L of furfural added to the medium. This was 17% higher than that obtained
without acetic acid and furfural. Furthermore, the results showed that acetic acid alone did not influence the cellulase activity
even at the highest concentration. However, β-glucosidase activity was increased with increasing acetic acid concentration.
Furfural proved to be an inhibiting agent causing a significant decrease in both cellulase and β-glucosidase production. 相似文献
6.
It is commonly observed that the rate of enzymatic hydrolysis of solid cellulose substrates declines markedly with time. In
this work the mechanism behind the rate reduction was investigated using two dominant cellulases of Trichoderma reesei: exoglucanase Cel7A (formerly known as CBHI) and endoglucanase Cel7B (formerly EGI). Hydrolysis of steam-pretreated spruce
(SPS) was performed with Cel7A and Cel7B alone, and in reconstituted mixtures. Throughout the 48-h hydrolysis, soluble products,
hydrolysis rates, and enzyme adsorption to the substrate were measured. The hydrolysis rate for both enzymes decreases rapidly
with hydrolysis time. Both enzymes adsorbed rapidly to the substrate during hydrolysis. Cel7A and Cel7B cooperate synergistically,
and synergism was approximately constant during the SPS hydrolysis. Thermal instability of the enzymes and product inhibition
was not the main cause of reduced hydrolysis rates. Adding fresh substrate to substrate previously hydrolyzed for 24 h with
Cel7A slightly increased the hydrolysis of SPS; however, the rate increased even more by adding fresh Cel7A. This suggests
that enzymes become inactivated while adsorbed to the substrate and that unproductive binding is the main cause of hydrolysis
rate reduction. The strongest increase in hydrolysis rate was achieved by adding Cel7B. An improved model is proposed that
extends the standard endo-exo synergy model and explains the rapid decrease in hydrolysis rate. It appears that the processive
action of Cel7A becomes hindered by obstacles in the lignocellulose substrate. Obstacles created by disordered cellulose chains
can be removed by the endo activity of Cel7B, which explains some of the observed synergism between Cel7A and Cel7B. The improved
model is supported by adsorption studies during hydrolysis. 相似文献
7.
In an earlier binding study conducted in our laboratory using Thermobifida fusca cellulases Cel6B, Cel9A, and Cel5A (formally Thermomonospora fusca E3, E4, and E5), it was observed that binding capacities for these three cellulases were 18–30 times higher on BMCC than on Avicel. These
results stimulated an interest in how the difference in accessibility between the two cellulosic substrates would affect synergism
observed with cellulase mixtures. To explore the impact of substrate, accessibility on the extent of conversion and synergism,
three binary T. fusca cellulase mixtures were tested over a range of cellulase ratios and total molar cellulase concentrations on Avicel and BMCC.
Higher extents of conversion were observed for BMCC due to the higher enzyme to substrate ratio resulting from the higher
binding The processive endoglucanase, Cel9A, had four times the extent of conversion of the end endocellulase Cel5A, while
the exocellulase Cel6B had three times the extent of conversion of Cel5A. Approximately 500 nmol/g of the cel9A+Cel6B mixture
was needed to obtain 80% conversion, while the Cel6B+Cel5A and Cel9A+Cel5A mixtures required 1500 and 1250 nmol/g, respectively,
to obtain 80% conversion. Thus, it appears that the more accessible structure of BMCC, as reflected by its binding capacity,
results in relative higher processive activity. 相似文献
8.
Henriksson H Muñoz IG Isaksson R Pettersson G Johansson G 《Journal of chromatography. A》2000,898(1):63-74
Cellobiohydrolase 58 (EC 3.2.1.91, P.c. Cel 7D) from Phanerochaete chrysosporium was immobilized on silica and the resulting material, CBH 58-silica, was then used as a chiral stationary phase (CSP) in liquid chromatographic separations of enantiomers. The enantioselectivities obtained on CBH 58-silica were compared with those on CBH I-silica (a phase based on a corresponding cellulase from Trichoderma reesei). CBH 58-silica displayed higher selectivity than CBH I-silica for the more hydrophilic compounds, such as atenolol and metoprolol, although great similarities in chiral separation of beta-adrenergic antagonists were found between the two phases. None of the acidic compounds tested could be resolved on the CBH 58 phase. Moreover, the solutes were retained more on the CBH 58 phase in general, indicating an improved application potential in bioanalysis. Addition of cellobiose or lactose, both of which are inhibitors of cellulases, to the mobile phase impaired the enantioselectivity, indicating an overlap of the enantioselective and catalytic sites. The chiral analytes also functioned as competitive inhibitors and their inhibition constants were determined. 相似文献
9.
A cellulase production process was developed by growing the fungi Trichoderma reesei and Aspergillus phoenicis on dairy manure. T. reesei produced a high total cellulase titer (1.7 filter paper units [FPU]/mL, filter paper activity) in medium containing 10 g/L
of manure (dry basis [w/w]), 2 g/L KH2PO4, 2 mL/L of Tween-80, and 2mg/L of CoCl2. However, β-glucosidase activity in the T. reesei-enzyme system was very low. T. reesei was then cocultured with A. phoenicis to enhance the β-glucosidase level. The mixed culture resulted in a relatively high level of total cellulase (1.54 FPU/mL)
and β-glucosidase (0.64 IU/mL). The ratio of β-glucosidase activity to filter paper activity was 0.41, suitable for hydrolyzing
manure cellulose. The crude enzyme broth from the mixed culture was used for hydrolyzing the manure cellulose, and the produced
glucose was significantly (p<0.01) higher than levels obtained by using the commercial enzyme or the enzyme broth of the pure culture T. reesei. 相似文献
10.
LiFePO4/C composites are prepared by using two types of carbon source: one using polymer (PAALi) and the other using sucrose. The physical characteristics of LiFePO4/C composites are investigated by X-ray diffraction), scanning electron microscopy, BET, laser particle analyzer, and Raman spectroscopy. Their electrochemical properties are characterized by cyclic voltammograms, constant current charge–discharge, and electrochemical impedance spectra. These analyses indicate that the carbon source and carbon content have a great effect on the physical and electrochemical performances of LiFePO4/C composites. An ideal carbon source and appropriate carbon content can effectively increase the lithium-ion diffusion coefficient and exchange current density, decrease the charge transfer resistance (R ct), and enhance the electrochemical performances of LiFePO4/C composite. The results show that PAALi is a better carbon source for the synthesis of LiFePO4/C composites. When the carbon content is 4.11 wt.% (the molar ratio of PAALi/Li2C2O4 was 2:1), as-prepared LiFePO4/C composite shows the best combination between electrochemical performances and tap density. 相似文献
11.
Santos Alexandre S. Sampaio Ana Paula W. Vasquez Gina S. Santa Anna Lídia M. Pereira Nei Freire Denise M. G. 《Applied biochemistry and biotechnology》2002,98(1-9):1025-1035
Culture conditions involving variations in carbon and nitrogen sources and different C:N ratios were examined with the aim
of increasing productivity in the process of rhamnolipid synthesis by Pseudomonas aeruginosa. In addition to the differences in productivity, the use of different carbon sources resulted in several proportions related
to the types of rhamnolipids synthesized (monorhamnolipids and dirhamnolipids). Furthermore, the variation in nutrients, mainly
the nitrogen source, resulted in different amounts of virulence factors, as phenazines and extracellular proteins. The data
point out a new concern in the choice of substrate to be used for rhamnolipid production by P. aeruginosa: toxic byproducts. 相似文献
12.
Polypropylene powders as the adsorbent for organic solution containing n-hexadecane and olive oil were employed as the carbon source for producing an alkaline lipase from Acinetobacter radioresistens. The best volumetric ratio of n-hexadecane to olive oil around 5 for lipase production was determined from shake-flask and fermentation cultivations. The
existence of a maximum time course lipase activity of the aqueous phase was attributed to the compensation effects of olive
oil on cell growth and lipase production, repression of lipase synthesis by oleic acid, and lipase adsorption on the supports.
A linear relationship between the average cell growth rate in the exponential phase and the ratio of surface areas of the
supports was found. The benefits of using the present fermentation process include less foaming and emulsion of the broth,
less organic phase used, higher lipase production, and easy recovery of the lipase in the centrifugation step. 相似文献
13.
Genetic engineering has been used for fusion of the peptide tag, Trp-Pro-Trp-Pro, on a protein to study the effect on partitioning in aqueous two-phase systems. As target protein for the fusions the cellulase, endoglucanase I (endo-1,4-beta-Dglucan-4-glucanohydrolase, EC 3.2.1.4, EGI, Cel7B) of Trichoderma reesei was used. For the first time a glycosylated two-domain enzyme has been utilized for addition of peptide tags to change partitioning in aqueous two-phase systems. The aim was to find an optimal fusion localization for EGI. The peptide was (1) attached to the C-terminus end of the cellulose binding domain (CBD), (2) inserted in the glycosylated linker region, (3) added after a truncated form of EGI lacking the CBD and a small part of the linker. The different constructs were expressed in the filamentous fungus T. reesei under the gpdA promoter from Aspergillus nidulans. The expression levels were between 60 and 100 mg/l. The partitioning behavior of the fusion proteins was studied in an aqueous two-phase model system composed of the thermoseparating ethylene oxide (EO)-propylene oxide (PO) random copolymer EO-PO (50:50) (EO50PO50) and dextran. The Trp-Pro-Trp-Pro tag was found to direct the fusion protein to the top EO50PO50 phase. The partition coefficient of a fusion protein can be predicted with an empirical correlation based on independent contributions from partitioning of unmodified protein and peptide tag in this model system. The fusion position at the end of the CBD, with the spacer Pro-Gly, was shown to be optimal with respect to partitioning and tag efficiency factor (TEF) was 0.87, where a fully exposed tag would have a TEF of 1.0. Hence, this position can further be utilized for fusion with longer tags. For the other constructs the TEF was only 0.43 and 0.10, for the tag fused to the truncated EGI and in the linker region of the full length EGI, respectively. 相似文献
14.
利用几种常见的液态烃作为碳源对MoO3/Al2O3进行了程序升温碳化(TPC),通过在线质谱(MS)的跟踪分析以及对所得催化剂的XRD表征,研究了这些烃在TPC过程中对MoO3/Al2O3物相转变的影响以及这些烃在不断转化催化剂表面的化学行为.结果显示在不同温度范围内这些烃具有不同的碳化活性,对MoO3/Al2O3的完全碳化来说,正己烷表现出了最好的活性.TPC-MS的分析结果也见证了MoO3/Al2O3碳化时的中间物相及其对链烃的脱氢芳构化作用,这对于不同用途的钼基碳化物催化剂制备时合适碳源的选择极具参考意义. 相似文献
15.
The production of pigment-free pullulan byAureobasidium pullulans, using olive oil and sucrose as carbon (C) sources, in shake flasks, was investigated. Optimum medium composition for pullulan
elaboration was 80 g/L sucrose, 25 mL/L olive oil, 5 mL/L Tween-80, 10 g/L glutamic acid, and an initial pH of 5.5. Maximum
pullulan concentration (51.5 g/L), productivity (8.6 g/L·d), and yield (80.3%) were achieved under these conditions after
120 h of fermentation. The principal advantage of using olive oil and sucrose simultaneously as C sources was the elimination
of the inhibitory effect of high sucrose concentrations (> 60 g/L) on pullulan production by the microorganism. Structural
characterization by13C-NMR, monosaccharide, and methylation analyses, and pullulanase digestion, combined with size-exclusion chromatography, confirmed
the identity of pullulan and the homogeneity of the released polysaccharide in the fermentation broths. There were no significant
differences in structure between pullulan samples isolated from either olive oil-supplemented media or olive oil-free media.
The molecular size of pullulan from the combined olive oil-sucrose fermentation was slightly lower (1.1 X 106) than that of conventional fermentation with sucrose as a single C source (1.4 X 106). Lowering the initial pH of the medium resulted in increased molecular size for the released polymer, but a lower pullulan
yield. 相似文献
16.
Sybil O’Reilly Sonia Erazo Victoriano Campos Erika Salas Jaime Baeza AndrÉ Ferraz Jaime Rodriguez Nelson DurÁn 《Applied biochemistry and biotechnology》1991,27(3):267-276
The single cell protein and extracellular enzyme production from a lignocellulolytic fungus,Chrysonilia sitophila, using different carbon sources were evaluated. The mycelial dry mass composition showed a high protein (39.2%) and low nucleic
acid content (3.3%), as well as carbohydrate, fatty acid, fiber, and ash levels comparable with single cell proteins currently
studied. Mycelial protein showed amino acid content similar to or higher than FAO standard requirements. The amino acid, fatty
acid, and carotenoid composition, as well as mycelial mass yield and enzyme production, were dependent on the carbon source
used. Glucose, saccharose, cellobiose, cellulose, microcrystal-line cellulose, lactose, and rice hull as carbon sources were
studied. 相似文献
17.
Xu G Hou AJ Wang RR Liang GY Zheng YT Liu ZY Li XL Zhao Y Huang SX Peng LY Zhao QS 《Organic letters》2006,8(20):4453-4456
Przewalskin A (1), a novel C23 terpenoid with a 6/6/7 carbon ring skeleton, was isolated from Salvia przewalskii. Its structure was determined by comprehensive 1D NMR, 2D NMR, and MS spectroscopic analysis and subsequently confirmed by a single-crystal X-ray diffraction study of its PDC oxidation derivative (2). Compounds 1 and 2 showed modest anti-HIV-1 activity with EC50 = 41 and 89 microg/mL, respectively. 相似文献
18.
Elisa M. P. Ciapina Walber C. Melo Lidia M. M. Santa Anna Alexandre S. Santos Denise M. G. Freire Nei Pereira Jr. 《Applied biochemistry and biotechnology》1996,131(1-3):880-886
The production of biosurfactant by Rhodococcus erythropolis during the growth on glycerol was investigated. The process was carried out at 28°C in a 1.5-L bioreactor using glycerol
as carbon source. The bioprocess was monitored through measurements of biosurfactant concentration and glycerol consumption.
After 51 h of cultivation, 1.7 g/L of biosurfactant, surface, and interfacial tensions values (with n-hexadecane) of 43 and 15 mN/m, respectively, 67% of Emulsifying Index (E
24), and 94% of oil removal were obtained. The use of glycerol rather than what happens with hydrophobic carbon source allowed
the release of the biosurfactant, originally associated to the cell wall. 相似文献
19.
Ingrid Persson Folke Tjerneld Bärbel Hähn-Hagerdal 《Applied biochemistry and biotechnology》1991,27(1):9-25
Cellulolytic enzyme production in aqueous two-phase systems withTrichoderma reesei Rutgers C30 has been investigated. The influ ence of different phase systems, as well as addition of media compo nents and substrate on enzyme production have been studied. Extractive enzyme production in fed-batch cultivations was per formed in a phase system composed of PEG 8000 5%-Dextran T500 7% with 1% Solka-Floc BW 200 as substrate. The cellulolytic enzyme system was intermittently withdrawn with the top phase. Addition of media components every 24 h and cellulose every 72 h gave an aver age enzyme activity in the withdrawn top phase of 2.2 FPU/mL dur ing 170 h cultivation. The corresponding productivity was 18 FPU/lh. The productivity was increased to 24 FPU/l.h when media compo nents and cellulose were added every 72 h. The average enzyme con centration was then 1.6 FPU/mL. The results are discussed in relation to methods for cellulolytic enzyme production involving immobiliza tion and cell recycling. 相似文献
20.
《Electrochemistry communications》2007,9(3):480-484
In this work we report on the analytical performance of glassy carbon electrodes modified with a dispersion of multi-wall carbon nanotubes in polyethylenimine (GCE/(PEI/CNT)). The resulting electrodes show an excellent electrocatalytic activity toward different bioanalytes like ascorbic acid, dopamine, 3,4-dihydroxyphenylacetic acid (dopac) and hydrogen peroxide. An important decrease in the overvoltages for the oxidation of ascorbic acid (505 mV) and hydrogen peroxide (350 mV) and for the reduction of hydrogen peroxide (450 mV), as well as a dramatic improvement in the reversibility of the electrochemical behavior of dopamine and dopac is obtained. The currents are higher than those obtained with other dispersant agents like Nafion, concentrated acids or chitosan, evidencing the high efficiency of the dispersion in PEI. The GCE/(PEI/CNT) demonstrated to be highly reproducible, with 3.0% RSD for the sensitivity of hydrogen peroxide for 10 electrodes prepared with five different dispersions. Differences in sensitivity of 10.0% were obtained for hydrogen peroxide with electrodes prepared using the same dispersion even after 14 days preparation. The CNT/PEI layer immobilized on glassy carbon electrodes has been also used as a platform for building supramolecular architectures based on the self-assembling of polyelectrolytes without any pretreatment of the electrode surface, oxidation or derivatization of the carbon nanotubes, just taking advantages of the polycationic nature of the polymer used for dispersing the nanotubes. The self-assembling of glucose oxidase has allowed us to obtain a supramolecular multistructure for glucose biosensing, with detection limits of 11 μM (0.02 g/L). Such an excellent performance of GCE/(PEI/CNT) toward hydrogen peroxide and the effectiveness of the use of CNT/PEI as a platform for obtaining supramolecular multistructures, represents a very good alternative for developing other enzymatic biosensors. 相似文献