Fermentation kinetics for xylitol production by a Pichia stipitis D-Xylulokinase mutant previously grown in spent sulfite liquor

Spent sulfite pulping liquor (SSL) contains lignin, which is present as lignosulfonate, and hemicelluloses that are present as hydrolyzed carbohydrates. To reduce the biological oxygen demand of SSL associated with dissolved sugars, we studied the capacity of Pichia stipitis FPL-YS30 (xyl3 Delta) to convert these sugars into useful products. FPL-YS30 produces a negligible amount of ethanol while converting xylose into xylitol. This work describes the xylose fermentation kinetics of yeast strain P.stipitis FPL-YS30. Yeast was grown in rich medium supplemented with different carbon sources: glucose, xylose, or ammonia-base SSL. The SSL and glucose-acclimatized cells showed similar maximum specific growth rates (0.146 h(-1)). The highest xylose consumption at the beginning of the fermentation process occurred using cells precultivated in xylose, which showed relatively high specific activity of glucose-6-phosphate dehydrogenase (EC 1.1.1.49). However, the maximum specific rates of xylose consumption (0.19 g(xylose)/g(cel) h) and xylitol production (0.059 g(xylitol)/g(cel) h) were obtained with cells acclimatized in glucose, in which the ratio between xylose reductase (EC 1.1.1.21) and xylitol dehydrogenase (EC 1.1.1.9) was kept at higher level (0.82). In this case, xylitol production (31.6 g/l) was 19 and 8% higher than in SSL and xylose-acclimatized cells, respectively. Maximum glycerol (6.26 g/l) and arabitol (0.206 g/l) production were obtained using SSL and xylose-acclimatized cells, respectively. The medium composition used for the yeast precultivation directly reflected their xylose fermentation performance. The SSL could be used as a carbon source for cell production. However, the inoculum condition to obtain a high cell concentration in SSL needs to be optimized.

Alkaline-sulfite chemithermomechanical pulping of Eucalyptus grandis biotreated by Ceriporiopsis subvermispora under varied culture conditions

The effect of different culture conditions have been evaluated concerning the extracellular enzyme activities of the white-rot fungus Ceriporiopsis subvermispora growing on Eucalyptus grandis wood. The consequence of the varied fungal pretreatment on a subsequent chemithermomechanical pulping (CTMP) was addressed. In all cultures, manganese peroxidase (MnP) and xylanase were the predominant extracellular enzymes. The biopulping efficiency was evaluated based on the amount of fiber bundles obtained after the first fiberizing step and the fibrillation levels of refined pulps. It was found that the MnP levels in the cultures correlated positively with the biopulping benefits. On the other hand, xylanase and total oxalate levels did not vary significantly. Accordingly, it was not possible to determine whether MnP accomplishes the effect alone or depends on synergic action of other extracellular agents. Pulp strength and fiber size distribution were also evaluated. The average fiber length of CTMP pulps prepared from untreated wood chips was 623 mu m. Analogous values were observed for most of the biopulps; however, significant amounts of shorter fibers were found in the biopulp prepared from wood chips biotreated in cultures supplemented with glucose plus corn-steep liquor. Despite evidence of reduced average fiber length, biopulps prepared from these wood chips presented the highest improvement in tensile indexes (+28% at 23 degrees Schopper-Riegler).

Estimation of hexenuronic acids and kappa number in kraft pulps of Eucalyptus globulus by Fourier transform near infrared spectroscopy and multivariate analysis

Fourier transform near infrared (FT-NIR) spectroscopy was evaluated as an analytical too[ for monitoring residual Lignin, kappa number and hexenuronic acids (HexA) content in kraft pulps of Eucalyptus globulus. Sets of pulp samples were prepared under different cooking conditions to obtain a wide range of compound concentrations that were characterised by conventional wet chemistry analytical methods. The sample group was also analysed using FT-NIR spectroscopy in order to establish prediction models for the pulp characteristics. Several models were applied to correlate chemical composition in samples with the NIR spectral data by means of PCR or PLS algorithms. Calibration curves were built by using all the spectral data or selected regions. Best calibration models for the quantification of lignin, kappa and HexA were proposed presenting R-2 values of 0.99. Calibration models were used to predict pulp titers of 20 external samples in a validation set. The lignin concentration and kappa number in the range of 1.4-18% and 8-62, respectively, were predicted fairly accurately (standard error of prediction, SEP 1.1% for lignin and 2.9 for kappa). The HexA concentration (range of 5-71 mmol kg(-1) pulp) was more difficult to predict and the SEP was 7.0 mmol kg(-1) pulp in a model of HexA quantified by an ultraviolet (UV) technique and 6.1 mmol kg(-1) pulp in a model of HexA quantified by anion-exchange chromatography (AEC). Even in wet chemical procedures used for HexA determination, there is no good agreement between methods as demonstrated by the UV and AEC methods described in the present work. NIR spectroscopy did provide a rapid estimate of HexA content in kraft pulps prepared in routine cooking experiments.

Preparation and characterization of Mo/W bimetallic carbides by using different synthesis methods

Molybdenum and tungsten bimetallic oxides were synthetized according to the following methods: Pechini, coprecipitation and solid state reaction (SSR). After the characterization, those solids were carbureted at programmed temperature. The carburation process was monitored by checking the consumption of carburant hydrocarbon and CO produced. The monitoring process permits to avoid or to diminish the formation of pirolytic carbon.

Optimal fermentation conditions for maximizing the ethanol production by Kluyveromyces fragilis from cheese whey powder

Cheese whey powder (CWP) is an attractive raw material for ethanol production since it is a dried and concentrated form of CW and contains lactose in addition to nitrogen, phosphate and other essential nutrients. In the present work, deproteinized CWP was utilized as fermentation medium for ethanol production by Kluyveromyces fragilis. The individual and combined effects of initial lactose concentration (50-150 kg m(-3)), temperature (25-35 degrees C) and inoculum concentration (1-3 kg m(-3)) were investigated through a 2(3) full-factorial central composite design, and the optimal conditions for maximizing the ethanol production were determined. According to the statistical analysis, in the studied range of values, only the initial lactose concentration had a significant effect on ethanol production, resulting in higher product formation as the initial substrate concentration was increased. Assays with initial lactose concentration varying from 150 to 250 kg m(-3) were thus performed and revealed that the use of 200 kg m(-3) initial lactose concentration, inoculum concentration of 1 kg m(-3) and temperature of 35 degrees C were the best conditions for maximizing the ethanol production from CWP solution. Under these conditions, 80.95 kg m(-3) of ethanol was obtained after 44 h of fermentation. (C) 2011 Elsevier Ltd. All rights reserved.

Xylitol production from DEO hydrolysate of corn stover by Pichia stipitis YS-30

Corn stover that had been treated with vapor-phase diethyl oxalate released a mixture of mono- and oligosaccharides consisting mainly of xylose and glucose. Following overliming and neutralization, a d-xylulokinase mutant of Pichia stipitis, FPL-YS30 (xyl3-a dagger 1), converted the stover hydrolysate into xylitol. This research examined the effects of phosphoric or gluconic acids used for neutralization and urea or ammonium sulfate used as nitrogen sources. Phosphoric acid improved color and removal of phenolic compounds. d-Gluconic acid enhanced cell growth. Ammonium sulfate increased cell yield and maximum specific cell growth rate independently of the acid used for neutralization. The highest xylitol yield (0.61 g(xylitol)/g(xylose)) and volumetric productivity (0.18 g(xylitol)/g(xylose) l) were obtained in hydrolysate neutralized with phosphoric acid. However, when urea was the nitrogen source the cell yield was less than half of that obtained with ammonium sulfate.

Lignocellulosic polysaccharides and lignin degradation by wood decay fungi: the relevance of nonenzymatic Fenton-based reactions

The brown rot fungus Wolfiporia cocos and the selective white rot fungus Perenniporia medulla-panis produce peptides and phenolate-derivative compounds as low molecular weight Fe(3+)-reductants. Phenolates were the major compounds with Fe(3+)-reducing activity in both fungi and displayed Fe(3+)-reducing activity at pH 2.0 and 4.5 in the absence and presence of oxalic acid. The chemical structures of these compounds were identified. Together with Fe(3+) and H(2)O(2) (mediated Fenton reaction) they produced oxygen radicals that oxidized lignocellulosic polysaccharides and lignin extensively in vitro under conditions similar to those found in vivo. These results indicate that, in addition to the extensively studied Gloeophyllum trabeum-a model brown rot fungus-other brown rot fungi as well as selective white rot fungi, possess the means to promote Fenton chemistry to degrade cellulose and hemicellulose, and to modify lignin. Moreover, new information is provided, particularly regarding how lignin is attacked, and either repolymerized or solubilized depending on the type of fungal attack, and suggests a new pathway for selective white rot degradation of wood. The importance of Fenton reactions mediated by phenolates operating separately or synergistically with carbohydrate-degrading enzymes in brown rot fungi, and lignin-modifying enzymes in white rot fungi is discussed. This research improves our understanding of natural processes in carbon cycling in the environment, which may enable the exploration of novel methods for bioconversion of lignocellulose in the production of biofuels or polymers, in addition to the development of new and better ways to protect wood from degradation by microorganisms.

ETHANOL PRODUCTION FROM XYLOSE BY Pichia stipitis NRRL Y-7124 IN A STIRRED TANK BIOREACTOR

The ethanol production by Pichia stipitis was evaluated in a stirred tank bioreactor using semi-defined medium containing xylose (90.0 g/l) as the main carbon source. Experimental assays were performed according to a 2(2) full factorial design to evaluate the influence of aeration (0.25 to 0.75 vvm) and agitation (150 to 250 rpm) conditions on ethanol production. In the studied range of values, the agitation increase and aeration decrease favored ethanol production, which was maximum (26.7 g/l) using 250 rpm and 0.25 vvm, conditions that gave a volumetric oxygen transfer coefficient (k(L)a value) of 4.9 h(-1). Under these conditions, the ethanol yield factor, ethanol productivity, and the process efficiency were 0.32 g/g, 0.32 g/l.h, and 63%, respectively. These results are promising and contribute to the development of a suitable process for ethanol production from xylose by Pichia stipitis.

Linoleic acid peroxidation initiated by Fe(3+)-reducing compounds recovered from Eucalyptus grandis biotreated with Ceriporiopsis subvermispora

This work work evaluates linoleic acid peroxidation reactions initiated by Fe(3+)-reducing compounds recovered from Eucalyptus grandis, biotreated with the biopulping fungus Ceriporiopsis subvermispora. The aqueous extracts from biotreated wood had the ability to reduce Fe(3+) ions from freshly prepared solutions. The compounds responsible for the Fe(3+)-reducing activity corresponded to UV-absorbing substances with apparent molar masses from 3 kDa to 5 kDa. Linoleic acid peroxidation reactions conducted in the presence of Fe(3+) ions and the Fe(3+)-reducing compounds showed that the rate of O(2) consumption during peroxidation was proportional to the Fe(3+)-reducing activity present in each extract obtained from biotreated wood. This peroxidation reaction was coupled with in-vitro treatment of ball-milled E. grandis wood. Ultraviolet data showed that the reaction system released lignin fragments from the milled wood. Size exclusion chromatography data indicated that the solubilized material contained a minor fraction representing high-molar-mass molecules excluded by the column and a main low-molar-mass peak. Overall evaluation of the data suggested that the Fe(3+)-reducing compounds formed during wood biodegradation by C subvermispora can mediate lignin degradation through linoleic acid peroxidation. (C) 2010 Elsevier Ltd. All rights reserved.

Immobilization and stabilization of microbial lipases by multipoint covalent attachment on aldehyde-resin affinity: Application of the biocatalysts in biodiesel synthesis

Microbial lipase preparations from Thermomyces lanuginosus (TLL) and Pseudomonas fluorescens (PFL) were immobilized by multipoint covalent attachment on Toyopearl AF-amino-650M resin and the most active and thermal stable derivatives used to catalyze the transesterificanon reaction of babassu and palm oils with ethanol in solvent-free media For this different activating agents mainly glutaraldehyde glycidol and epichlorohydrin were used and immobilization parameters were estimated based on the hydrolysis of olive oil emulsion and butyl butyrate synthesis ILL immobilized on glyoxyl-resin allowed obtaining derivatives with the highest hydrolytic activity (HA(der)) and thermal stability between 27 and 31 times more stable than the soluble lipase Although PFL derivatives were found to be less active and thermally stables similar formation of butyl butyrate concentrations were found for both ILL and PFL derivatives The highest conversion into biodiesel was found in the transesterification of palm oil catalyzed by both ILL and PFL glyoxyl-derivatives (c) 2010 Elsevier B V All rights reserved

Inhibitory action of toxic compounds present in lignocellulosic hydrolysates on xylose to xylitol bioconversion by Candida guilliermondii

The inhibitory action of acetic acid, ferulic acid, and syringaldehyde on metabolism of Candida guilliermondii yeast during xylose to xylitol bioconversion was evaluated. Assays were performed in buffered and nonbuffered semidefined medium containing xylose as main sugar (80.0 g/l), supplemented or not with acetic acid (0.8-2.6 g/l), ferulic acid (0.2-0.6 g/l), and/or syringaldehyde (0.3-0.8 g/l), according to a 2(3) full factorial design. Since only individual effects of the variables were observed, assays were performed in a next step in semidefined medium containing different concentrations of each toxic compound individually, for better understanding of their maximum concentration that can be present in the fermentation medium without affecting yeast metabolism. It was concluded that acetic acid, ferulic acid, and syringaldehyde are compounds that may affect Candida guilliermondii metabolism (mainly cell growth) during bioconversion of xylose to xylitol. Such results are of interest and reveal that complete removal of toxic compounds from the fermentation medium is not necessary to obtain efficient conversion of xylose to xylitol by Candida guilliermondii. Fermentation in buffered medium was also considered as an alternative to overcome the inhibition caused by these toxic compounds, mainly by acetic acid.

Optimal recovery process conditions for manganese-peroxidase obtained by solid-state fermentation of eucalyptus residue using Lentinula edodes

Enzyme production is a growing field in biotechnology and increasing attention has been devoted to the solid-state fermentation (SSF) of lignocellulosic biomass for production of industrially relevant lignocellulose deconstruction enzymes, especially manganese-peroxidase (MnP), which plays a crucial role in lignin degradation. However, there is a scarcity of studies regarding extraction of the secreted metabolities that are commonly bound to the fermented solids, preventing their accurate detection and limiting recovery efficiency. In the present work, we assessed the effectiveness of extraction process variables (pH, stirring rate, temperature, and extraction time) on recovery efficiency of manganese-peroxidase (MnP) obtained by SSF of eucalyptus residues using Lentinula edodes using statistical design of experiments. The results from this study indicated that of the variables studied, pH was the most significant (p < 0.05%) parameter affecting MnP recovery yield, while temperature, extraction time, and stirring rate presented no statistically significant effects in the studied range. The optimum pH for extraction of MnP was at 4.0-5.0, which yielded 1500-1700 IU kg (1) of enzyme activity at extraction time 4-5 h, under static condition at room temperature. (C) 2011 Elsevier Ltd. All rights reserved.

Thermodynamic and kinetic investigations of phosphate adsorption onto hydrous niobium oxide prepared by homogeneous solution method

The adsorption kinetics of phosphate onto Nb(2)O(5)center dot nH(2)O was investigated at initial phosphate concentrations 10 and 50 mg L(-1). The kinetic process was described by a pseudo second-order rate model very well. The adsorption thermodynamics was carried out at 298, 308, 318, 328 and 338 K. The positive values of both Delta H and Delta S suggest an endothermic reaction and increase in randomness at the solid-liquid interface during the adsorption. Delta G values obtained were negative indicating a spontaneous adsorption process. The Langmuir model described the data better than the Freundlich isotherm model. The peak appearing at 1050 cm(-1) in IR spectra after adsorption was attributed to the bending vibration of adsorbed phosphate. The effective desorption could be achieved using water at pH 12. (C) 2010 Elsevier B.V. All rights reserved.

High Solids Content, Soap-Free, Film-Forming Latexes Stabilized by Laponite Clay Platelets

High solids content film-forming poly[styrene-co-(n-butyl acrylate)] [poly(Sty-co-BuA)] latexes armored with Laponite clay platelets have been synthesized by soap-free emulsion copolymerization of styrene and n-butyl acrylate. The polymerizations were performed in batch in the presence of Laponite and a methyl ether acrylateterminated poly(ethylene glycol) macromonomer in order to promote polymer/clay association. The overall polymerization kinetics showed a pronounced effect of clay on nucleation and stabilization of the latex particles. Cryo-transmission electron microscopy observation confirmed the armored morphology and indicated that the majority of Laponite platelets were located at the particle surface. The resulting nanostructured films displayed enhanced mechanical properties.

The Influence of Initial Xylose Concentration, Agitation, and Aeration on Ethanol Production by Pichia stipitis from Rice Straw Hemicellulosic Hydrolysate

Rice straw hemicellulosic hydrolysate was used as fermentation medium for ethanol production by Pichia stipitis NRRL Y-7124. Shaking bath experiments were initially performed aiming to establish the best initial xylose concentration to be used in this bioconversion process. In the sequence, assays were carried out under different agitation (100 to 200 rpm) and aeration ((V) under bar (flask)/V(medium) ratio varying from 2.5 to 5.0) conditions, and the influence of these variables on the fermentative parameters values (ethanol yield factor, Y(P/S); cell yield factor, Y(X/S); and ethanol volumetric productivity, Q(P)) was investigated through a 2(2) full-factorial design. Initial xylose concentration of about 50 g/l was the most suitable for the development of this process, since the yeast was able to convert substrate in product with high efficiency. The factorial design assays showed a strong influence of both process variables in all the evaluated responses. The agitation and aeration increase caused a deviation in the yeast metabolism from ethanol to biomass production. The best results (Y(P/S) = 0.37 g/g and Q(P) = 0.39 g/l. h) were found when the lowest aeration (2.5 V(flask)/V(medium) ratio) and highest agitation (200 rpm) levels were employed. Under this condition, a process efficiency of 72.5% was achieved. These results demonstrated that the establishment of adequate conditions of aeration is of great relevance to improve the ethanol production from xylose by Pichia stipitis, using rice straw hemicellulosic hydrolysate as fermentation medium.