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.

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.

Xylooligosaccharides Production from Alkali-Pretreated Sugarcane Bagasse Using Xylanases from Thermoascus aurantiacus

Sugarcane bagasse hemicellulose was isolated in a one-step chemical extraction using hydrogen peroxide in alkaline media. The polysaccharide containing 80.9% xylose and small amounts of L-arabinose, 4-O-methyl-D-glucuronic acid and glucose, was hydrolyzed by crude enzymatic extracts from Thermoascus aurantiacus at 50 degrees C. Conditions of enzymatic hydrolysis leading to the best yields of xylose and xylooligosaccharides (DP 2-5) were investigated using substrate concentration in the range 0.5-3.5% (w/v), enzyme load 40-80 U/g of the substrate, and reaction time from 3 to 96 h, applying a 22 factorial design. The maximum conversion to xylooligosaccharides (37.1%) was obtained with 2.6% of substrate and xylanase load of 60 U/g. The predicted maximum yield of xylobiose by a polynomial model was 41.6%. Crude enzymatic extract of T. aurantiacus generate from sugarcane bagasse hemicellulose 39% of xylose, 59% of xylobiose, and 2% of other xylooligosaccharides.

The roles of xylan and lignin in oxalic acid pretreated corncob during separate enzymatic hydrolysis and ethanol fermentation

High yields of hemicellulosic and cellulosic sugars are critical in obtaining economical conversion of agricultural residues to ethanol. To optimize pretreatment conditions, we evaluated oxalic acid loading rates, treatment temperatures and times in a 2(3) full factorial design. Response-surface analysis revealed an optimal oxalic acid pretreatment condition to release sugar from the cob of Zea mays L ssp. and for Pichia stipitis CBS 6054. To ferment the residual cellulosic sugars to ethanol following enzymatic hydrolysis, highest saccharification and fermentation yields were obtained following pretreatment at 180 degrees C for 50 min with 0.024 g oxalic acid/g substrate. Under these conditions, only 7.5% hemicellulose remained in the pretreated substrate. The rate of cellulose degradation was significantly less than that of hemicellulose and its hydrolysis was not as extensive. Subsequent enzymatic saccharification of the residual cellulose was strongly affected by the pretreatment condition with cellulose hydrolysis ranging between 26.0% and 76.2%. The residual xylan/lignin ratio ranged from 0.31 to 1.85 depending on the pretreatment condition. Fermentable sugar and ethanol were maximal at the lowest ratio of xylan/lignin and at high glucan contents. The model predicts optimal condition of oxalic acid pretreatment at 168 degrees C, 74 min and 0.027 g/g of oxalic acid. From these findings, we surmised that low residual xylan was critical in obtaining maximal glucose yields from saccharification. Published by Elsevier Ltd.

Production, characterization and application of activated carbon from brewer`s spent grain lignin

Different types of activated carbon were prepared by chemical activation of brewer`s spent grain (BSG) lignin using H(3)PO(4) at various acid/lignin ratios (1, 2, or 3 g/g) and carbonization temperatures (300, 450, or 600 degrees C), according to a 2(2) full-factorial design. The resulting materials were characterized with regard to their surface area, pore volume, and pore size distribution, and used for detoxification of BSG hemicellulosic hydrolysate (a mixture of sugars, phenolic compounds, metallic ions, among other compounds). BSG carbons presented BET surface areas between 33 and 692 m(2)/g, and micro- and mesopores with volumes between 0.058 and 0.453 cm(3)/g. The carbons showed high capacity for adsorption of metallic ions, mainly nickel, iron, chromium, and silicon. The concentration of phenolic compounds and color were also reduced by these sorbents. These results suggest that activated carbons with characteristics similar to those commercially found and high adsorption capacity can be produced from BSG lignin. (C) 2009 Elsevier Ltd. All rights reserved.

A study on the pretreatment of a sugarcane bagasse sample with dilute sulfuric acid

Experiments based on a 2(3) central composite full factorial design were carried out in 200-ml stainless-steel containers to study the pretreatment, with dilute sulfuric acid, of a sugarcane bagasse sample obtained from a local sugar-alcohol mill. The independent variables selected for study were temperature, varied from 112.5A degrees C to 157.5A degrees C, residence time, varied from 5.0 to 35.0 min, and sulfuric acid concentration, varied from 0.0% to 3.0% (w/v). Bagasse loading of 15% (w/w) was used in all experiments. Statistical analysis of the experimental results showed that all three independent variables significantly influenced the response variables, namely the bagasse solubilization, efficiency of xylose recovery in the hemicellulosic hydrolysate, efficiency of cellulose enzymatic saccharification, and percentages of cellulose, hemicellulose, and lignin in the pretreated solids. Temperature was the factor that influenced the response variables the most, followed by acid concentration and residence time, in that order. Although harsher pretreatment conditions promoted almost complete removal of the hemicellulosic fraction, the amount of xylose recovered in the hemicellulosic hydrolysate did not exceed 61.8% of the maximum theoretical value. Cellulose enzymatic saccharification was favored by more efficient removal of hemicellulose during the pretreatment. However, detoxification of the hemicellulosic hydrolysate was necessary for better bioconversion of the sugars to ethanol.

Simultaneous saccharification and ethanol fermentation of oxalic acid pretreated corncob assessed with response surface methodology

Response surface methodology was used to evaluate optimal time, temperature and oxalic acid concentration for simultaneous saccharification and fermentation (SSF) of corncob particles by Pichia stipitis CBS 6054. Fifteen different conditions for pretreatment were examined in a 2(3) full factorial design with six axial points. Temperatures ranged from 132 to 180 degrees C, time from 10 to 90 min and oxalic acid loadings from 0.01 to 0.038 g/g solids. Separate maxima were found for enzymatic saccharification and hemicellulose fermentation, respectively, with the condition for maximum saccharification being significantly more severe. Ethanol production was affected by reaction temperature more than by oxalic acid and reaction time over the ranges examined. The effect of reaction temperature was significant at a 95% confidence level in its effect on ethanol production. Oxalic acid and reaction time were statistically significant at the 90% level. The highest ethanol concentration (20 g/l) was obtained after 48 h with an ethanol volumetric production rate of 0.42 g ethanol l(-1) h(-1). The ethanol yield after SSF with P. stipitis was significantly higher than predicted by sequential saccharification and fermentation of substrate pretreated under the same condition. This was attributed to the secretion of beta-glucosidase by P. stipitis. During SSF, free extracellular beta-glucosidase activity was 1.30 pNPG U/g with P. stipitis, while saccharification without the yeast was 0.66 pNPG U/g. Published by Elsevier Ltd.

Search for optimum conditions of sugarcane bagasse hemicellulose extraction

A process has been elaborated for one-step low lignin content sugarcane bagasse hemicellulose extraction using alkaline solution of hydrogen peroxide. To maximize the hemicellulose yields several extraction conditions were examined applying the 2(4) factorial design: H(2)O(2) concentration from 2 to 6% (w/v), reaction time from 4 to 16 h, temperature from 20 to 60 degrees C, and magnesium sulfate absence or presence (0.5%, w/v). This approach allowed selection of conditions for the extraction of low and high lignin content hemicellulose. At midpoint the yield of hemicellulose was 94.5% with more than 88% of lignin removed. Lignin removal is suppressed at low extraction temperatures and in the absence of magnesium sulfate. Hemicellulose in 86% yield with low lignin content (5.9%) was obtained with 6% H(2)O(2) treatment for 4 h and 20 degrees C. This hemicellulose is much lighter in color than samples obtained at the midpoint condition and was found suitable for subsequent enzymatic hydrolysis. (C) 2009 Elsevier B.V. All rights reserved.

PVA-Hydrogel Entrapped Candida Guilliermondii for Xylitol Production from Sugarcane Hemicellulose Hydrolysate

Viable cells of Candida guilliermondii were immobilized by inclusion into polyvinyl alcohol (PVA) hydrogel using the freezing-thawing method. Entrapment experiments were planned according to a 2(3) full factorial design, using the PVA concentration (80, 100, and 120 g L(-1)), the freezing temperature (-10, -15, and -20 degrees C), and the number of freezing-thawing cycles (one, three, and five) as the independent variables, integrated with three additional tests to estimate the errors. The effectiveness of the immobilization procedure was checked in Erlenmeyer flasks as the pellet capability to catalyze the xylose-to-xylitol bioconversion of a medium based on sugarcane bagasse hemicellulosic hydrolysate. To this purpose, the yield of xylitol on consumed xylose, xylitol volumetric productivity, and cell retention yield were selected as the response variables. Cell pellets were then used to perform the same bioconversion in a stirred tank reactor operated at 400 rpm, 30 degrees C, and 1.04 vvm air flowrate. At the end of fermentation, a maximum xylitol concentration of 28.7 g L(-1), a xylitol yield on consumed xylose of 0.49 g g(-1) and a xylitol volumetric productivity of 0.24 g L(-1) h(-1) were obtained.

Zn uptake, physiological response and stress attenuation in mycorrhizal jack bean growing in soil with increasing Zn concentrations

The influence of arbuscular mycorrhizal fungi (AMF) inoculation on Canavalia ensiformis growth. nutrient and Zn uptake, and on some physiological parameters in response to increasing soil Zn concentrations was studied. Treatments were applied in seven replicates in a 2 x 4 factorial design, consisting of the inoculation or not with the AMF Glomus etunicatum, and the addition of Zn to soil at the concentrations of 0, 100, 300 and 900 mg kg(-1). AMF inoculation enhanced the accumulation of Zn in tissues and promoted biomass yields and root nodulation. Mycorrhizal plants exhibited relative tolerance to Zn up to 300 mg kg(-1) without exhibiting visual symptoms of toxicity, in contrast to non-mycorrhizal plants which exhibited a significant growth reduction at the same soil Zn concentration. The highest concentration of Zn added to soil was highly toxic to the plants. Leaves of plants grown in high Zn concentration exhibited a Zn-induced proline accumulation and also an increase in soluble amino acid contents; however proline contents were lower in mycorrhizal jack beans. Plants in association or not with the AMF exhibited marked differences in the foliar soluble amino acid profile and composition in response to Zn addition to soil. In general, Zn induced oxidative stress which could be verified by increased lipid peroxidation rates and changes in catalase, ascorbate peroxidase, glutathione reductase and superoxide dismutase activities. In summary, G. etunicatum was able to maintain an efficient symbiosis with jack bean plants in moderately contaminated Zn-soils, improving plant performance under those conditions, which is likely to be due to a combination of physiological and nutritional changes caused by the intimate relation between fungus and plant. The enhanced Zn uptake by AMF inoculated jack bean plants might be of interest for phytoremediation purposes. (C) 2009 Elsevier Ltd. All rights reserved.

Nitrogen and potassium fertilization affect apple fruit quality in southern Brazil

Nitrogen (N) and potassium (K) are usually found in higher concentrations than other macronutrients in apple (Malus x domestica Borkh) fruits and are most frequently associated with changes in fruit quality. The aim of this article was to evaluate the effects of N and K fertilization on some fruit quality attributes of Fuji apple. The experiment was conducted at Sao Joaquim, State of Santa Catarina, Brazil, during 2004 and 2005. A factorial design was used with N and K annual fertilizer rates (0, 50, 100, and 200 kg ha(-1) of N and K2O) replicated in three orchards. Fifteen days prior to harvest, three fruit samples were collected from each treatment and site. One sample was used for total soluble solid content (TSS), titratable acidity, pulp firmness, and fruit color parameter analyses, and the other samples were refrigerated in a conventional atmosphere for 3 and 6 months for subsequent determination of fruit quality. Nitrogen fertilization negatively affected fruit color, flesh firmness, and TSS content. These same variables were positively affected by K fertilization, except for flesh firmness.

The development and production of leaves and tillers by Marandu palisadegrass fertilised with nitrogen and sulphur

The effects of combined nitrogen and sulphur fertilisation on the dynamics of leaf and tiller appearance in Marandu palisadegrass (Brachiaria brizantha cv. Marandu) and its impact on dry matter production were evaluated in a greenhouse study. Grass seedlings were grown in pots filled with a soil classified as an Entisol and were harvested after 43 days, a further 35 days and finally after 48 more days. Five rates of N (0, 100, 200, 300 and 400 mg/dm(3)) and 5 rates of S (0, 10, 20, 30 and 40 mg/dm(3)) were tested in an incomplete factorial design with 4 replications. Leaf and tiller development were monitored every 3 days by counting the appearance of recently expanded leaves and new basal tillers. The phyllochron and thermal time between appearance of tillers decreased as N and S fertiliser levels increased to about 300 and 25 mg/dm(3), respectively, then tended to increase. In contrast, leaf and tiller appearance rates increased with the supply of these nutrients to similar levels, then tended to decline. Leaf and tiller production and dry matter yields were affected by both N and S levels, with the role of S increasing as the growth phases increased.

Influence of light and leaf epicuticular wax layer on Phakopsora pachyrhizi infection in soybean

Influence of light and leaf epicuticular wax layer on Phakopsora pachyrhizi infection in soybean Asian rust, caused by the fungus Phakopsora pachyrhizi, is one of the most serious phytosanitary problems of soybean in Brazil, especially because no cultivars with satisfactory resistance levels as yet exist. The objective of this study was to evaluate the influence of luminosity and of leaf epicuticular wax on the infection of soybean by P. pachyrhizi. The adaxial and abaxial leaflet surfaces of the first trifoliate leaf from cultivar BRS 154, phenological stage V2, were inoculated with a suspension of 105 uredospores/mL. The plants were kept for 24 hours in a humid chamber at temperature of 23 degrees C, in light or dark conditions, using a factorial design. Subsequently, the plants were maintained for 14 days under a 12-hour photoperiod. The disease severity and density were evaluated. For in vitro experiments, in light or dark conditions, the evaluation was done in terms of uredospore germination and appressorium formation. The wax content of adaxial and abaxial leaflets was analyzed quantitatively using chloroform extraction and ultrastructurally using scanning electron microscope. Higher density and severity were observed when the adaxial surface was inoculated, with later incubation of the plants in the dark, with no significant interaction between these factors. Spore germination in the dark (40.7%) was statistically different from spore germination in the light (28.5%). The same effect was observed with appressorium formation, in the dark (24.7%) and in the light (12.8%). The quantity and the ultrastructural aspects of epicuticular wax content did not show differences between the adaxial and abaxial surfaces; nor did they show any effect on infection by Phakopsora pachyrhizi in the soybean cultivar studied.

Changes in bone mass, biomechanical properties, and microarchitecture of calcium- and iron-deficient rats fed diets supplemented with inulin-type fructans

Feeding mineral-deficient diets enhances absorptive efficiency as an attempt of the body to compensate for the lack of an essential nutrient. Under certain circumstances, it does not succeed, and nutritional deficiency is produced Our hypothesis was that mulin-type fructans (ITF), which arc known to affect mineral absorption, could increase Ca and Fe bioavailability in Ca- and Fe-deficient rats. Male Wistar rats (n = 48, 4 weeks old) were assigned to I of 8 groups derived from 2 x 2 x 2 factorial design with 2 levels of added Fe (0 and 35 mg/kg), Ca (0 and 5 g/kg), and ITF (0 and 100 g/kg) for 33 days. The Fe status (hemoglobin, serum Fe, total Fe-binding capacity, transferrin saturation, liver minerals) was evaluated. Tibia minerals (Ca, Mg, and Zn), bone strength, and histomorphometry were determined In nondeficient rats, ITF supplementation did not affect Fe status or organ minerals, with the exception of tibia Mg Moreover, ITF improved bone resilience and led to a reduction in eroded surface per body surface and number of osteoclasts per area In Ca-deficient rats, ITF increased liver (Fe and Zn) and tibia (Zn) mineral levels but impaired tibia Mg, yield load, and resilience. In conclusion, ITF worsened the tibia Mg levels and elastic properties when supplemented in Ca-deficient diets In contrast, although bone Ca was not affected in nondeficient rats under the present experimental conditions, bone quality improved, as demonstrated by a moderate reduction in femur osteoclast resorption and significant increases in tibia Mg content and elasticity. (C) 2009 Elsevier Inc. All rights reserved.

Effect of Different Cooking Conditions on Phenolic Compounds and Antioxidant Capacity of Some Selected Brazilian Bean (Phaseolus vulgaris L.) Cultivars

The effects of different cooking conditions such as soaking, atmospheric (100 degrees C) or pressure boiling (121 degrees C), and draining of cooking water following thermal treatment on phenolic compounds and the DPPH radical scavenging capacity from two selected Brazilian bean cultivars (black and yellow-brown seed coat color) were investigated using a factorial design (2(3)). Factors that significantly reduced the total phenolic contents and antioxidant capacity in both cultivars were the soaking and draining stage. Independent of cooking temperature, total phenolics and antioxidant capacities were enhanced in treatments without soaking and where cooking water was not discarded, and this was likely linked to an increase of specific phenolic compounds detected by high performance liquid chromatography such as flavonols and free phenolic acids in both cultivars. Cooking of beans either at 100 or 121 degrees C, without a soaking stage and keeping the cooking water, would be recommendable for retaining antioxidant phenolic compounds.