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Category Archives: Biotechnology

Effects of alkaline or liquid-ammonia treatment on crystalline cellulose: Changes in crystalline structure and effects on enzymatic digestibility

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Background:
In converting biomass to bioethanol, pretreatment is a key step intended to render cellulose more amenable and accessible to cellulase enzymes and thus, increase glucose yields. In this study, four cellulose samples with different degrees of polymerization and crystallinity indexes were subjected to aqueous NaOH and anhydrous liquid ammonia treatments. The effects of the treatments on cellulose crystalline structure were studied, in addition to the effects on the digestibility of the celluloses by a cellulase complex.
Results:
From X-ray diffractograms and NMR spectra, it was revealed that treatment with liquid-ammonia produced the cellulose III allomorph; however, crystallinity depended on treatment conditions. Treatment at low temperature (25degreesC) resulted in a less crystalline product, whereas treatment at elevated temperatures (130degreesC or 140degreesC) gave a more crystalline product. Treatment of cellulose I with aqueous NaOH (16.5 wt%) resulted in formation of cellulose II, but also produced a much less crystalline cellulose. The relative digestibilities of the different cellulose allomorphs were tested by exposing the treated and untreated cellulose samples to a commercial enzyme mixture (Genencor-Danisco; GC 220). The digestibility results showed that the starting cellulose I samples were the least digestible (except for corn stover cellulose, which had a high amorphous content). Treatment with NaOH produced the most digestible cellulose, followed by treatment with liquid-ammonia at low temperature. Factor analysis indicated that initial rates of digestion (up to 24 h) were most strongly correlated with amorphous content. Correlation of allomorph type with digestibility was weak, but was strongest with cellulose conversion at later times. The cellulose III samples produced at higher temperature had comparable crystallinities to the initial cellulose I samples, but achieved higher levels of cellulose conversion, at longer digestion times.
Conclusions:
Earlier studies have focused on determining which cellulose allomorph is the most digestible. In this study we have found that the chemical treatments to produce different allomorphs also changed the crystallinity of the cellulose, and this had a significant effect on the digestibility of the substrate. When determining the relative digestibilities of different cellulose allomorphs it is essential to also consider the relative crystallinities of the celluloses being tested.Source:
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Transcriptome analysis of Aspergillus niger grown on sugarcane bagasse

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Background:
Considering that the costs of cellulases and hemicellulases contribute substantially to the price of bioethanol, new studies to understand and improve cellulase efficiency and productivity are of paramount importance. Aspergillus niger has been shown to produce a wide spectrum of polysaccharide-hydrolytic enzymes. In order to understand how to improve enzymatic cocktails that can hydrolyze pre-treated sugarcane bagasse, we used a genomics approach to investigate which genes and pathways are transcriptionally modulated during growth of A. niger on steam-exploded sugarcane bagasse (SEB).
Results:
Here, we report the main cellulase- end hemicellulase-encoding genes with increased expression during growth on SEB. We also examined if the mRNA accumulation of several SEB-induced genes encoding putative transporters was induced by xylose and dependent on glucose. We identified 18 (58 % of A. niger predicted cellulases) and 21 (58 % of A. niger predicted hemicellulases) cellulases- and hemicellulases-encoding genes, respectively, that were highly expressed during growth on SEB.
Conclusions:
Degradation of sugarcane bagasse requires production of many different enzymes, which are regulated by the type and complexity of the available substrate. Our work opens new possibilities for understanding sugarcane biomass saccharification by A. niger hydrolases and for the construction of more efficient enzymatic cocktails for second generation bioethanol.Source:
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Application of a Burkholderia cepacia lipase-immobilized silica monolith to the batch and continuous biodiesel production with a stoichiometric mixture of methanol and crude Jatropha oil

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Background:
The enzymatic production of biodiesel through alcoholysis of triglycerides has become more attractive because it shows potential in overcoming the drawbacks of chemical processes. In this study, we investigate the production of biodiesel from crude, non-edible Jatropha oil and methanol to characterize Burkholderia cepacia lipase immobilized in an n-butyl-substituted hydrophobic silica monolith. We also evaluate the performance of a lipase-immobilized silica monolith bioreactor in the continuous production of biodiesel.
Results:
The Jatropha oil used contained 18% free fatty acid, which is problematic in a base-catalyzed process. In the lipase-catalyzed reaction, the presence of free fatty acid made the reaction mixture homogeneous and allowed bioconversion to proceed to 90% biodiesel yield after a 12 h reaction time. The optimal molar ratio of methanol to oil was 3.3-3.5:1 with water content of 0.6% (w/w). Further experiments revealed that B. cepacia lipase immobilized in hydrophobic silicates was sufficiently tolerant to methanol, and glycerol adsorbed on the support disturbed the reaction to some extent in the present reaction system. The continuous production of biodiesel was performed at steady state using a lipase-immobilized silica monolith bioreactor loaded with 1.67 g of lipase. The yield of 95% was reached at a flow rate of 0.6 mL h-1, although the performance of the continuous bioreactor was somewhat below that predicted from the batch reactor. The bioreactor was operated successfully for almost 50 days with 80% retention of the initial yield.
Conclusions:
The presence of free fatty acids originally contained in Jatropha oil improved the reaction efficiency of the biodiesel production. A combination of B. cepacia lipase and its immobilization support, n-butyl-substituted silica monolith, was effective in the production of biodiesel. This procedure is easily applicable to the design of a continuous flow-through bioreactor system.Source:
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A cellular automaton model of crystalline cellulose hydrolysis by cellulases

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Background:
Cellulose from plant biomass is an abundant, renewable material which could be a major feedstock for low emissions transport fuels such as cellulosic ethanol. Cellulase enzymes that break down cellulose into fermentable sugars are composed of different types - cellobiohydrolases I and II (CBHI and CBHII), endoglucanase (EG) and beta-glucosidase (BG) - with separate functions. They form a complex interacting network between themselves, soluble hydrolysis product molecules, solution and solid phase substrates and inhibitors. There have been many models proposed for enzymatic saccharification, however, none have yet employed a cellular automaton approach which allows important phenomena such as enzyme crowding on surface of solid substrates, denaturation and substrate inhibition to be considered in the model.
Results:
The Cellulase 4D model was developed de novo taking into account the size and composition of the substrate and surface-acting enzymes were ascribed behaviors based on their movements, catalytic activities and rates, affinity for, and potential for crowding of, the cellulose surface, substrates and inhibitors, and denaturation rates. A basic case modeled on literature-derived parameters obtained from Trichoderma reesei cellulases resulted in cellulose hydrolysis curves that closely matched curves obtained from published experimental data. Scenarios were tested in the model which included variation of enzyme loadings, adsorption strengths of surface acting enzymes and reaction periods, and the effect on saccharide production over time was assessed. The model simulations indicated an optimal enzyme loading of between 0.5 and 2 of the base case concentrations where a balance was obtained between enzyme crowding on the cellulose crystal, and that the affinities of enzymes for the cellulose surface had a large effect on cellulose hydrolysis. In addition, improvements to the CBHI activity period substantially improved overall glucose production.
Conclusions:
Cellulase 4D simulates the enzymatic hydrolysis of cellulose to glucose by surface and solution phase-acting enzymes and accounts for complex phenomena that have previously not been included in cellulose hydrolysis models. The model is intended as a tool for industry, researchers and educators alike to explore options for enzyme engineering and process development and to test hypotheses regarding cellulase mechanisms.Source:
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Biotechnology – Video

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The biotechnology, we have to know it, is not only attached to be aplicated to medical/health things. agriculture clearly fits the broad definition of "using a biotechnological system to make products" such that the cultivation of plants may be viewed as the earliest biotechnological enterprise. Since the human`s begins, the biotechnology is present, no that advance as our technology but with some relation, for example the Indians, select the best herbs to cure themselves The processes and methods of agriculture have been refined by other mechanical and biological sciences since its inception

Here is the original post:
Biotechnology - Video

Improving L-arabinose utilization of pentose fermenting Saccharomyces cerevisiae cells by heterologous expression of L-arabinose transporting sugar transporters

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Background:
Hydrolysates of plant biomass used for the production of lignocellulosic biofuels typically contain sugar mixtures consisting mainly of D-glucose and D-xylose, and minor amounts of L-arabinose. The yeast Saccharomyces cerevisiae is the preferred microorganism for the fermentative production of ethanol but is not able to ferment pentose sugars. Although D-xylose and L-arabinose fermenting S. cerevisiae strains have been constructed recently, pentose uptake is still a limiting step in mixed sugar fermentations.
Results:
Here we described the cloning and characterization of two sugar transporters, AraT from the yeast Scheffersomyces stipitis and Stp2 from the plant Arabidopsis thaliana, which mediate the uptake of L-arabinose but not of D-glucose into S. cerevisiae cells. A yeast strain lacking all of its endogenous hexose transporter genes and expressing a bacterial L-arabinose utilization pathway could no longer take up and grow with L-arabinose as the only carbon source. Expression of the heterologous transporters supported uptake and utilization of L-arabinose especially at low L-arabinose concentrations but did not, or only very weakly, support D-glucose uptake and utilization. In contrast, the S. cerevisiae D-galactose transporter, Gal2, mediated uptake of both L-arabinose and D-glucose, especially at high concentrations.
Conclusions:
Using a newly developed screening system we have identified two heterologous sugar transporters from a yeast and a plant which can support uptake and utilization of L-arabinose in L-arabinose fermenting S. cerevisiae cells, especially at low L-arabinose concentrations.Source:
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The promoting effect of by-products from Irpex lacteus on subsequent enzymatic hydrolysis of bio-pretreated cornstalks

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Background:
Irpex lacteus, a versatile lignin-degrading fungus with various extracellular enzymes, has been widely used for biological pretreatment. However, most studies have focused on the change of substrate structure after biological pretreatment, and the effect of these changes on the enzymatic hydrolysis, but the effect of byproducts from biological pretreatment process on subsequent enzymatic hydrolysis is not well understood.
Methods:
We developed a biological pretreatment process with I. lacteus that can produce stimulatory byproducts that enhance the enzymatic hydrolysis of cornstalks.
Results:
The maximum hydrolysis yield of glucan (82%) was obtained after pretreatment for 28 days. The maximum reducing sugar yield decreased from 313.5 to 200.1 mg/g raw cornstalks after water-soluble byproducts of biological pretreatment were removed from pretreated cornstalks. The effect of byproducts on enzymatic hydrolysis was also investigated. We found that the hydrolysis efficiency of commercial cellulase preparation on cornstalks could be improved by water extracts from bio-pretreated cornstalks with hydrolytic enzyme activity and iron-reducing activity.
Conclusion:
The key finding suggested that byproducts from biological pretreatment play important roles in enhancing downstream hydrolysis, which might be attributable to hydrolytic enzymes and iron-reducing compounds produced by I. lacteus.Source:
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  1. Biotechnology