Showing papers in "Applied Biochemistry and Biotechnology in 2004"

Lipases and their industrial applications: an overview.

Abstract: Lipases (triacylglycerol acylhydrolase, EC 3.1.1.3) are part of the family of hydrolases that act on carboxylic ester bonds. The physiologic role of lipases is to hydrolyze triglycerides into diglycerides, monoglycerides, fatty acids, and glycerol. These enzymes are widely found throughout the animal and plant kingdoms, as well as in molds and bacteria. Of all known enzymes, lipases have attracted the most scientific attention. In addition to their natural function of hydrolyzing carboxylic ester bonds, lipases can catalyze esterification, interesterification, and transesterification reactions in nonaqueous media. This versatility makes lipases the enzymes of choice for potential applications in the food, detergent, pharmaceutical, leather, textile, cosmetic, and paper industries. The most significant industrial applications of lipases have been mainly found in the food, detergent, and pharmaceutical sectors. Limitations of the industrial use of these enzymes have mainly been owing to their high production costs, which may be overcome by molecular technologies, enabling the production of these enzymes at high levels and in a virtually purified form.

Effects of Sugar Inhibition on Cellulases and β-Glucosidase During Enzymatic Hydrolysis of Softwood Substrates

Abstract: A quantitative approach was taken to determine the inhibition effects of glucose and other sugar monomers during cellulase and β-Glucosidase hydrolysis of two types of cellulosic material: Avicel and acetic acid-pretreated softwood. The increased glucose content in the hydrolysate resulted in a dramatic increase in the degrees of inhibition on both β-Glucosidase and cellulase activities. Supplementation of mannose, xylose, and galactose during cellobiose hydrolysis did not show any inhibitory effects on β-Glucosidase activity. However, these sugars were shown to have significant inhibitory effects on cellulase activity during cellulose hydrolysis. Our study suggests that high-substrate consistency hydrolysis with supplementation of hemicellulose is likely to be a practical solution to minimizing end-product inhibition effects while producing hydrolysate with high glucose concentration.

Two-step preparation for catalyst-free biodiesel fuel production

Abstract: Biodiesel fuel was prepared by a two-step reaction: hydrolysis and methyl esterification. Hydrolysis was carried out at a subcritical state of water to obtain fatty acids from triglycerides of rapeseed oil, while the methyl esterification of the hydrolyzed products of triglycerides was treated near the supercritical methanol condition to achieve fatty acid methyl esters. Consequently, the two-step preparation was found to convert rapeseed oil to fatty acid methyl esters in considerably shorter reaction time and milder reaction condition than the direct supercritical methanol treatment. The optimum reaction condition in this two-step preparation was 270°C and 20 min for hydrolysis and methyl esterification, respectively. Variables affecting the yields in hydrolysis and methyl esterification are discussed.

Production of ethanol from cellulosic biomass hydrolysates using genetically engineered Saccharomyces yeast capable of cofermenting glucose and xylose.

Abstract: Recent studies have proven ethanol to be the idael liquid fuel for transportation, and renewable ligno cellulosic materials to be the attractive feed stocks for ethanol fuel production by fermentation. The major fermentable sugars from hydrolysis of most cellulosic biomass are D-glucose and D-xylose. The naturally occurring Saccharomyces yeasts that are used by industry to produce ethanol from starches and cane sugar cannot metabolize xylose. Our group at Purdue University succeded in developing genetically engineered Saccharomyces yeasts capable of effectively cofermenting glucose and xylose to ethanol, which was accomplished by cloning three xylose-metabolizing genes into the yeast. In this study, we demonstrated that our stable recombinant Sacharomyces yeast, 424A (LNH-ST), which contains the cloned xylose-metabolizing genes stably integrated into the yeast chromosome in high copy numbers, can efficiently ferment glucose and xylose present in hydrolysates from different cellulosic biomass to ethanol.

Continuous Production of Butanol by Clostridium acetobutylicum Immobilized in a Fibrous Bed Bioreactor

Abstract: We explored the influence of dilution rate and pH in continuous cultures of Clostridium acetobutylicum. A 200-mL fibrous bed bioreactor was used to produce high cell density and butyrate concentrations at pH 5.4 and 35°C. By feeding glucose and butyrate as a cosubstrate, the fermentation was maintained in the solventogenesis phase, and the optimal butanol productivity of 4.6 g/(L h) and a yield of 0.42 g/g were obtained at a dilution rate of 0.9 h-1 and pH 4.3. Compared to the conventional acetone-butanol-ethanol fermentation, the new fermentation process greatly improved butanol yield, making butanol production from corn an attractive alternative to ethanol fermentation.

Yields from glucose, xylose, and paper sludge hydrolysate during hydrogen production by the extreme thermophile Caldicellulosiruptor saccharolyticus

Abstract: This study addressed the utilization of an industrial waste stream, paper sludge, as a renewable cheap feedstock for the fermentative production of hydrogen by the extreme thermophile Caldicellulosiruptor saccharolyticus. Hydrogen, acetate, and lactate were produced in medium in which paper sludge hydrolysate was added as the sole carbon and energy source and in control medium with the same concentration of analytical grade glucose and xylose. The hydrogen yield was dependent on lactate formation and varied between 50 and 94% of the theoretical maximum. The carbon balance in the medium with glucose and xylose was virtually 100%. The carbon balance was not complete in the paper sludge medium because the measurement of biomass was impaired owing to interfering components in the paper sludge hydrolysate. Nevertheless, >85% of the carbon could be accounted for in the products acetate and lactate. The maximal volumetric hydrogen production rate was 5 to 6 mmol/ (L•h), which was lower than the production rate in media with glucose, xylose, or a combination of these sugars (9–11 mmol/ [L•h]). The reduced hydrogen production rate suggests the presence of inhibiting components in paper sludge hydrolysate.

Evaluation of wet oxidation pretreatment for enzymatic hydrolysis of softwood.

Abstract: The wet oxidation pretreatment (water, oxygen, elevated temperature, and pressure) of softwood (Picea abies) was investigated for enhancing enzymatic hydrolysis. The pretreatment was preliminarily optimized. Six different combinations of reaction time, temperature, and pH were applied, and the compositions of solid and liquid fractions were analyzed. The solid fraction after wet oxidation contained 58–64% cellulose, 2–16% hemicellulose, and 24–30% lignin. The pretreatment series gave information about the roles of lignin and hemicellulose in the enzymatic hydrolysis. The temperature of the pretreatment, the residual hemicellulose content of the substrate, and the type of the commercial cellulase preparation used were the most important factors affecting the enzymatic hydrolysis. The highest sugar yield in a 72-h hydrolysis, 79% of theoretical, was obtained using a pretreatment of 200°C for 10 min at neutral pH.

Kinetics of Glucose Decomposition During Dilute-Acid Hydrolysis of Lignocellulosic Biomass

Abstract: Recent research work in-house both at Auburn University and National Renewable Energy Laboratory has demonstrated that extremely low concentrations of acid (e.g., 0.05–0.2 wt% sulfuric acid) and high temperatures (e.g., 200–230°C) are reaction conditions that can be effectively applied for hydrolysis of the cellulosic component of biomass. These conditions are far from those of the conventional dilute-acid hydrolysis processes, and the kinetic data for glucose decomposition are not currently available. We investigated the kinetics of glucose decomposition covering pH values of 1.5–2.2 and temperatures of 180–230°C using glass ampoule reactors. The primary factors controlling glucose decomposition are the reaction medium, acid concentration, and temperature. Based on the experimental data, a kinetic model was developed and the best-fit kinetic parameters were determined. However, a consistent discrepancy in the rate of glucose disappearance was found between that of the model based on pure glucose data and that observed during the actual process of lignocellulosic biomass hydrolysis. This was taken as an indication that glucose recombines with acid-soluble lignin during the hydrolysis process, and this conclusion was incorporated accordingly into the overall model of glucose decomposition.

Optimization of steam pretreatment of corn stover to enhance enzymatic digestibility.

Abstract: Among the available agricultural byproducts, corn stover, with its yearly production of 10 million t (dry basis), is the most abundant promising raw material for fuel ethanol production in Hungary In the United States, more than 216 million t of corn stover is produced annually, of which a portion also could possibly be collected for conversion to ethanol However, a network of lignin and hemicellulose protects cellulose, which is the major source of fermentable sugars in corn stover (approx 40% of the dry matter [DM]) Steam pretreatment removes the major part of the hemicellulose from the solid material and makes the cellulose more susceptible to enzymatic digestion We studied 12 different combinations of reaction temperature, time, and pH during steam pretreatment The best conditions (200 degrees C, 5 min, 2% H2SO4) increased the enzymatic conversion (from cellulose to glucose) of corn stover more then four times, compared to untreated material However, steam pretreatment at 190 degrees C for 5 min with 2% sulfuric acid resulted in the highest overall yield of sugars, 561 g from 100 g of untreated material (DM), corresponding to 73% of the theoretical The liquor following steam explosion was fermented using Saccharomyces cerevisiae to investigate the inhibitory effect of the pretreatment The achieved ethanol yield was slightly higher than that obtained with a reference sugar solution This demonstrates that baker's yeast could adapt to the pretreated liquor and ferment the glucose to ethanol efficiently

Two-step preparation for catalyst-free biodiesel fuel production: hydrolysis and methyl esterification.

Abstract: Biodiesel fuel was prepared by a two-step reaction: hydrolysis and methyl esterification. Hydrolysis was carried out at a subcritical state of water to obtain fatty acids from triglycerides of rapeseed oil, while the methyl esterification of the hydrolyzed products of triglycerides was treated near the supercritical methanol condition to achieve fatty acid methyl esters. Consequently, the two-step preparation was found to convert rapeseed oil to fatty acid methyl esters in considerably shorter reaction time and milder reaction condition than the direct supercritical methanol treatment. The optimum reaction condition in this two-step preparation was 270°C and 20 min for hydrolysis and methyl esterification, respectively. Variables affecting the yields in hydrolysis and methyl esterification are discussed.

Ammonia fiber explosion treatment of corn stover.

Abstract: Optimizing process conditions and parameters such as ammonia loading, moisture content of biomass, temperature, and residence time is necessary for maximum effectiveness of the ammonia fiber explosion process. Approximate optimal pretreatment conditions for corn stover were found to be temperature of 90°C, ammonia:dry corn stover mass ratio of 1:1, moisture content of corn stover of 60% (dry weight basis), and residence time (holding at target temperature), of 5 min. Approximately 98% of the theoretical glucose yield was obtained during enzymatic hydrolysis of the optimal treated corn stover using 60 filter paper units (FPU) of cellulase enzyme / g of glucan (equal to 22 FPU/g of dry corn stover). The ethanol yield from this sample was increased up to 2.2 times over that of untreated sample. Lowering enzyme loading to 15 and 7.5 FPU/g of glucan did not significantly affect the glucose yield compared with 60 FPU, and any differences between effects at different enzyme levels decreased as the treatment temperature increased.

Opportunities in the industrial biobased products industry.

Abstract: Approximately 89 million metric t of organic chemicals and lubricants, the majority of which are fossil based, are produced annually in the United States. The development of new industrial bioproducts, for production in standalone facilities or biorefineries, has the potential to reduce our dependence on imported oil and improve energy security. Advances in biotechnology are enabling the optimization of feedstock composition and agronomic characteristics and the development of new and improved fermentation organisms for conversion of biomass to new end products or intermediates. This article reviews recent biotechnology efforts to develop new industrial bioproducts and improve renewable feedstocks and key market opportunities.

Biosurfactant production by Bacillus subtilis using cassava-processing effluent.

Abstract: A cassava flour-processing effluent (manipueira) was evaluated as a substrate for surfactant production by two Bacillus subtilis strains. B. subtilis ATCC 21332 reduced the surface tension of the medium to 25.9 mN/m, producing a crude biosurfactant concentration of 2.2 g/L. The wild-type strain, B. subtilis LB5a, reduced the surface tension of the medium to 26.6 mN/m, giving a crude biosurfactant concentration of 3.0 g/L. A decrease in surfactant concentration observed for B. subtilis ATCC 21332 seemed to be related to an increase in protease activity. The biosurfactant produced on cassava effluent medium by B. subtilis LB5a was similar to surfactin.

Screening Genus Penicillium for Producers of Cellulolytic and Xylanolytic Enzymes

Abstract: For enzymatic hydrolysis of lignocellulosic material, cellulolytic enzymes from Trichoderma reesei are most commenly used, but, there is a need for more efficient enzyme cocktails. In this study, the production of cellulolytic and xylanolytic enzymes was investigated in 12 filamentous fungi from genus Penicillium and compared with that of T. reesei. Either Solka-Floc cellulose or oat spelt xylan was used as carbon source in shake flask cultivations. All the fungi investigated showed coinduction of cellulolytic and xylanolytic enzymes during growth on cellulose as well as on xylan. The highest filter paper activity was measured after cultivation of Penicillium brasilianum IBT 20888 on cellulose.

Optimization of enzymatic production of biodiesel from castor oil in organic solvent medium.

Abstract: We studied the production of fatty acid ethyl esters from castor oil using n-hexane as solvent and two commercial lipases, Novozym 435 and Lipozyme IM, as catalysts. For this purpose, a Taguchi experimental design was adopted considering the following variables: temperature (35–65°C), water (0–10 wt/wt%), and enzyme (5–20 wt/wt%) concentrations and oil-to-ethanol molar ratio (1∶3 to 1∶10). An empirical model was then built so as to assess the main and cross-variable effects on the reaction conversion and also to maximize biodiesel production for each enzyme. For the system containing Novozym 435 as tatalyst the maximum conversion obtained was 81.4% at 65°C, enzyme concentration of 20 wt/wt%, water concentration of 0 wt/wt%, and oil-to-ethanol molar ratio of 1∶10. When the catalyst was Lipozyme IM, a conversion as high as 98% was obtained at 65°C, enzyme concentration of 20 wt/wt%, water concentration of 0 wt/wt%, and oil-to-ethanol molar ratio of 1∶3.

Lactic acid bacteria used in inoculants for silage as probiotics for ruminants

Abstract: Many studies have shown the beneficial effects on ruminant performance of feeding them with silages inoculated with lactic acid bacteria (LAB). These benefits might derive from probiotic effects. The purpose of the current study was to determine whether LAB included in inoculants for silage can survive in rumen fluid (RF), as the first step in studying their probiotic effects. Experiments were conducted in the United States and Israel with clarified (CRF) and strained RF (SRF) that were inoculated at 10(6)-10(8) microorganisms/mL with and without glucose at 5 g/L. RF with no inoculants served as control. Ten commercial inoculants were used. The RF was incubated at 39 degrees C and sampled in duplicates at 6, 12, 24, 48, 72, and 96 h for pH and LAB counts. The results indicate that with glucose the pH of the RF decreased during the incubation period. In the SRF, the pH of the inoculated samples was higher than that of the controls in most cases. This might be a clue to the mechanism by which LAB elicit the enhancement in animal performance. LAB counts revealed that the inoculants survived in the RF during the incubation period. The addition of glucose resulted in higher LAB counts.

Discrimination among eight modified michaelis-menten kinetics models of cellulose hydrolysis with a large range of substrate/enzyme ratios: inhibition by cellobiose.

Abstract: The kinetics of exoglucanase (Cel7A) from Trichoderma reesei was investigated in the presence of cellobiose and 24 different enzyme/Avicel ratios for 47 h, in order to establish which of the eight available kinetic models best explained the factors involved. The heterogeneous catalysis was studied and the kinetic parameters were estimated employing integrated forms of Michaelis-Menten equations through the use of nonlinear least squares. It was found that cellulose hydrolysis follows a model that takes into account competitive inhibition by cellobiose (final product) with the following parameters: K m=3.8 mM, K ic=0.041 mM, k cat=2 h−1 (5.6×10−4 s−1). Other models, such as mixed type inhibition and those incorporating improvements concerning inhibition by substrate and parabolic inhibition, increased the modulation performance very slightly. The results support the hypothesis that nonproductive enzyme substrate complexes, parabolic inhibition, and enzyme inactivation (Selwyn test) are not the principal constraints in enzymatic cellulose hydrolysis. Under our conditions, the increment in hydrolysis was not significant for substrate/enzyme ratios <6.5.

Biodiesel Fuel from Vegetable Oil by Various Supercritical Alcohols

Abstract: Biodiesel was prepared in various supercritical alcohol treatments with methanol, ethanol, 1-propanol, 1-butanol, or 1-octanol to study transesterification of rapeseed oil and alkyl esterification of fatty acid at temperatures of 300 and 350°C. The results showed that in transesterification, the reactivity was greatly correlated to the alcohol: the longer the alkyl chain of alcohol, the longer the reaction treatment. In alkyl esterification of fatty acids, the conversion did not depend on the alcohol type because they had a similar reactivity. Therefore, the selection of alcohol in biodiesel production should be based on consideration of its performance of properties and economics.

Thermostable phytase production by Thermoascus aurantiacus in submerged fermentation.

Abstract: 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.

Enzyme production and profile by Aspergillus niger during solid substrate fermentation using palm kernel cake as substrate.

Abstract: The oil palm sector is one of the major plantation industries in Malaysia. Palm kernel cake is a byproduct of extracted palm kernel oil. Mostly palm kernel cake is wasted or is mixed with other nutrients and used as animal feed, especially for ruminant animals. Recently, palm kernel cake has been identified as an important ingredient for the formulation of animal feed, and it is also exported especially to Europe, South Korea, and Japan. It can barely be consumed by nonruminant (monogastric) animals owing to the high percentages of hemicellulose and cellulose contents. Palm kernel cake must undergo suitable pretreatment in order to decrease the percentage of hemicellulose and cellulose. One of the methods employed in this study is fermentation with microorganisms, particularly fungi, to partially degrade the hemicellulose and cellulose content. This work focused on the production of enzymes by Aspergillus niger and profiling using palm kernel cake as carbon source.

Effect of corn stover concentration on rheological characteristics.

Abstract: Corn stover, a well-known example of lignocellulosic biomass, is a potential renewable feed for bioethanol production. Dilute sulfuric acid pretreatment removes hemicellulose and makes the cellulose more susceptible to bacterial digestion. The rheologic properties of corn stover pretreated in such a manner were studied. The Power Law parameters were sensitive to corn stover suspension concentration becoming more non-Newtonian with slope n, ranging from 0.92 to 0.05 between 5 and 30% solids. The Casson and the Power Law models described the experimental data with correlation coefficients ranging from 0.90 to 0.99 and 0.85 to 0.99, respectively. The yield stress predicted by direct data extrapolation and by the Herschel-Bulkley model was similar for each concentration of corn stover tested.

Quantitative Analysis of Cellulose-Reducing Ends

Abstract: Methods for the quantification of total and accessible reducing ends on traditional cellulose substrates have been evaluated because of their relevance to enzyme-catalyzed cellulose saccharification. For example, quantification of accessible reducing ends is likely to be the most direct measure of substrate concentration for the exo-acting, reducing end-preferring cellobiohydrolases. Two colorimetric assays (dinitrosalicylic acid [DNS] and bicinchoninic acid [BCA] assay) and a radioisotope approach (NaB3H4 labeling) were evaluated for this application. Cellulose substrates included microcrystalline celluloses, bacterial celluloses, and filter paper. Estimates of the number of reducing ends per unit mass cellulose were found to be dependent on the assay system (i.e. the DNS and BCA assays gave strikingly different results). DNS-based values were several-fold higher than those obtained using the BCA assay, with fold-differences being substrate specific. Sodium borohydride reduction of celluloses, using cold or radiolabeled reagent under relatively mild conditions, was used to assess the number of surface (solvent-accessible) reducing ends. The results indicate that 30–40% of the reducing ends on traditional cellulose substrates are not solvent accessible; that is, they are buried in the interior of cellulose structures and thus not available to exo-acting enzymes.

Effect of pH on cellulase production of Trichoderma reesei RUT C30.

Abstract: Currently, the high market price of cellulases prohibits commercialization of the lignocellulosics-to-fuel ethanol process, which utilizes enzymes for saccharification of cellulose. For this reason research aimed at understanding and improving cellulase production is still a hot topic in cellulase research. Trichoderma reesei RUT C30 is known to be one of the best hyper producing cellulolytic fungi, which makes it an ideal test organism for research. New findings could be adopted for industrial strains in the hope of improving enzyme yields, which in turn may result in lower market price of cellulases, thus making fuel ethanol more cost competitive with fossil fuels. Being one of the factors affecting the growth and cellulase production of T. reesei, the pH of cultivation is of major interest. In the present work, numerous pH-controlling strategies were compared both in shake-flask cultures and in a fermentor. Application of various buffer systems in shake-flask experiments was also tested. Although application of buffers resulted in slightly lower cellulase activity than that obtained in non-buffered medium, β-glucosidase production was increased greatly.

Methodology for estimating removable quantities of agricultural residues for bioenergy and bioproduct use.

Abstract: A methodology was developed to estimate quantities of crop residues that can be removed while maintaining rain or wind erosion at less than or equal to the tolerable soil-loss level. Six corn and wheat rotations in the 10 largest corn-producing states were analyzed. Residue removal rates for each rotation were evaluated for conventional, mulch/reduced, and no-till field operations. The analyses indicated that potential removable maximum quantities range from nearly 5.5 million dry metric t/yr for a continuous corn rotation using conventional till in Kansas to more than 97 million dry metric t/yr for a corn-wheat rotation using no-till in Illinois.

Effect of temperature, moisture, and carbon supplementation on lipase production by solid-state fermentation of soy cake by Penicillium simplicissimum.

Abstract: The production of lipases by Penicillium simplicissimum using solid-state fermentation and soy cake as substrate was investigated. The effects of temperature, cake moisture, and carbon supplementation on lipase production were studied using a two-level experimental plan. Moisture, pH, and lipase activity were followed during fermentation. Statistical analysis of the results was performed to evaluate the effect of the studied variables on the maximum lipase activity. Incubation temperature was the variable that most affected enzyme activity, showing a negative effect. Moisture and carbon supplementation presented a positive effect on activity. It was possible to obtain lipase activity as high as 21 U/g of dry cake in the studied range of process variables.

Biosynthesis of Poly(3-hydroxybutyrate- co -3-hydroxyalkanoates) by Metabolically Engineered Escherichia coli Strains

Abstract: Biosynthesis of polyhydroxyalkanoates (PHAs) consisting of 3-hydroxyalkanoates (3HAs) of 4 to 10 carbon atoms was examined in metabolically engineered Escherichia coli strains. When the fadA and / or fadB mutant E. coli strains harboring the plasmid containing the Pseudomonas sp. 61–3 phaC2 gene and the Ralstonia eutropha phaAB genes were cultured in Luria-Bertani (LB) medium supplemented with 2 g/L of sodium decanoate, all the recombinant E. coli strains synthesized PHAs consisting of C4, C6, C8, and C10 monomer units. The monomer composition of PHA was dependent on the E. col i strain used. When the fadA mutant E. coli was employed, PHA containing up to 63 mol% of 3-hydroyhexanoate was produced. In fadB and fad B mutant E. coli strains, 3-hydroxybutyrate (3HB) was efficiently incorporated into PHA up to 86 mol%. Cultivation of recombinant fadA and / or fadB mutant E. coli strains in LB medium containing 10 g/L of sodium gluconate and 2 g/L of sodium decanoate resulted in the production of PHA copolymer containing a very high fraction of 3HB up to 95 mol%. Since the material properties of PHA copolymer consisting of a large fraction of 3HB and a small fraction of medium-chain-length 3HA are similar to those of low-density polyethylene, recombinant E. coi strains constructed in this study should be useful for the production of PHAs suitable for various commercial applications.

Siderophore production by a marine Pseudomonas aeruginosa and its antagonistic action against phytopathogenic fungi.

Abstract: A marine isolate of fluorescent Pseudomonas sp. having the ability to produce the pyoverdine type of siderophores under low iron stress (up to 10 microM iron in the succinate medium) was identified as Pseudomonas aeruginosa by using BIOLOG Breathprint and siderotyping. Pyoverdine production was optimum at 0.2% (w/v) succinate, pH 6.0, in an iron-deficient medium. Studies carried out in vitro revealed that purified siderophores and Pseudomonas culture have good antifungal activity against the plant deleterious fungi, namely, Aspergillus niger, Aspergillus flavus, Aspergillus oryzae, Fusarium oxysporum, and Sclerotium rolfsii. Siderophore-based maximum inhibition was observed against A. niger. These in vitro antagonistic actions of marine Pseudomonas against phytopathogens suggest the potential of the organism to serve as a biocontrol agent.

Enhanced Enzymatic Hydrolysis of Steam-Exploded Douglas Fir Wood by Alkali-Oxygen Post-treatment

Abstract: Good enzymatic hydrolysis of steam-exploded Douglas fir wood (SEDW) cannot be achieved owing to the very high lignin content (>40%) that remains associated with this substrate. Thus, in this study, we investigated the use of alkali-oxygen treatment as a posttreatment to delignify SEDW and also considered the enzymatic hydrolyzability of the delignified SEDW. The results showed that under optimized conditions of 15% NaOH, 5% consistency, 110°C, and 3 h, approx 84% of the lignin in SEDW could be removed. The resulting delignified SEDW had good hydrolyzability, and cellulose-to-glucose conversion yields of over 90 and 100% could be achieved within 48 h with 20 and 40 filter paper units / g of cellulose enzyme loadings, respectively. It was also indicated that severe conditions, such as high NaOH concentration and high temperature, should not be utilized in oxygen delignification of SEDW in order to avoid extensive condensation of lignin and significant degradation of cellulose.

High-cell-density cultivation of Pseudomonas putida IPT 046 and medium-chain-length polyhydroxyalkanoate production from sugarcane carbohydrates.

Abstract: We studied high-density cultures of Pseudomonas putida IPT 046 for the production of medium-chain-length polyhydroxyalkanoates (PHAMCL) using an equimolar mixture of glucose and fructose as carbon sources. Kinetics studies of P. putida growth resulted in a maximum specific growth rate of 0.65h−1. Limitation and inhibition owing to NH4 + ions were observed, respectively, at 400 and 3500 mg of NH4 +/L. The minimum concentration of dissolved oxygen in the broth must be 15% of saturation. Fed-batch strategies for high-cell-density cultivation were proposed. Pulse feed followed by constant feed produced a cell concentration of 32 g/L in 18 h of fermentation and low PHAMCL content. Constant feed produced a cell concentration of 35 g/L, obtained in 27 h of fermentation, with up to 15% PHAMCL. Exponential feed produced a cell concentration of 30 g/L in 20 h of fermentation and low PHAMCL content. Using the last strategy, 21% PHAMCL was produced during a period of 34 h of fed-batch operation, with a final cell concentration of 40 g/L and NH4 + limitation. Using phosphate limitation, 50 g/L cell concentration, 63% PHAMCL and a productivity of 0.8 g/(L·h) were obtained in 42 h of fed-batch operation. The PHAMCL yield factors from consumed carbohydrate for N-limited and P-limited experiments were, respectively, 0.15 and 0.19 g/g.

Impact of fluid velocity on hot water only pretreatment of corn stover in a flowthrough reactor.

Abstract: Flowthrough pretreatment with hot water only offers many promising features for advanced pretreatment of biomass, and a better understanding of the mechanisms responsible for flowthrough behavior could allow researchers to capitalize on key attributes while overcoming limitations In this study, the effect of fluid velocity on the fate of total mass, hemicellulose, and lignin was evaluated for hot water only pretreatment of corn stover in tubular flowthrough reactors Increasing fluid velocity significantly accelerated solubilization of total mass, hemicellulose, and lignin at early times For example, when fluid velocity was increased from 28 to 107 cm/min, xylan removal increased from 60 to 82% for hot water only pretreatment of corn stover at 200°C after 8 min At the same time, lignin removal increased from 30 to 46% Dissolved hemicellulose was almost all in oligomeric form, and solubilization of hemicellulose was always accompanied by lignin release The increase in removal of xylan and lignin with velocity, especially in the early reaction stage, suggests that chemical reaction is not the only factor controlling hemicellulose hydrolysis and that mass transfer and other physical effects may also play an important role in hemicellulose and lignin degradation and removal