Lignocellulose degradation in mushroom composts

The edible mushroomAgaricus bisporus is grown commercially on composted manure/straw mixtures. However, this proven composting procedure is wasteful of raw materials. A nonmanure compost was developed (Smith, 1980) with two main aims:

1. To conserve raw materials, while still producing a compost favoringAgaricus bisporus colonization and giving an economic yield of mushrooms.
2. To speed up composting, hence making more efficient use of labor, farm equipment, and buildings.

A “conservation compost” (wheat straw, bran, whey, urea, peat, and gypsum) is ready for inoculation with mushroom mycelium (spawning) after 7 d preparation, i.e., 2 d pre-wetting of straw, then 4–5 d composting under controlled conditions. Whereas a traditional manure/wheat straw compost is produced by composting in windrows (8–11 d) followed by a controlled pasteurization phase (5–7 d). In the preparation of a traditional mushroom compost, as much as 60% of the initial dry matter is lost by microbial degradation prior to spawning. By shortening the composting process to 7 d conservation of cellulose and hemicellulose is achieved with only some 30% loss in dry matter. Straw hemicelluloses are degraded much quicker than cellulose during composting. Hence, the measurable extracellular laminarinase and xylanase activities of the compost microflora appear much greater than their cellulase activities at this period in both composts. A peak in laminarinase and xylanase activity after 48 h in manure compost corresponds with the increase in microbial populations. A pronounced increase in thermophilic bacterial and actinomycete populations occurs in “conservation composts” as readily available soluble carbohydrates are assimilated. Initially, this results in higher uniform compost temperatures (60?C+) and leads to a reduced thermophilic fungal population (10³ viable propagules g^-1 dry wt compost), which may explain the lowered enzyme activities found in the “conservation composts” and thus the reduced degradation of lignocellulose. The compost microflora showed no laccase activity during composting, and little if any lignin was degraded. However,Agaricus bisporus does possess a moderately active lignolytic system and a strongly active cellulolytic system. Subsequent experiments have shown that increased mushroom yields may be obtained from these composts when urea is replaced by chicken manure as the nitrogen supplement (Smith, 1983); this has not affected compost “selectivity” for mushroom growth, dry matter loss, or the duration of the process. Although yield of mushrooms, based on compost weights at spawning tend to be lower than what would be expected from traditional composts, yield calculated on the basis of weight of starting materials is usually much higher.     

Alkaline peroxide mechanical pulping of wheat straw with enzyme treatment

Alkaline peroxide mechanical pulping (APMP) of wheat straw with enzyme treatment was studied. Instead of direct enzyme pretreatment on wheat straw, an alternative treatment method was used, in which coarse pulps from refiner defibrated wheat straw rather than wheat straw were pretreated with a crude enzyme containing mainly xylanase, then impregnated with alkaline H₂O₂ solution and further refined. The optimum conditions of enzyme treatment were xylanase dosage of 10–15 IU/g of oven-dried wheat straw, 90 min, 50–60°C, pulp consistency of 5–10%, and initial pH of 5.0, and those for chemical impregnation were 6% NaOH, 70–80°C, 60–90 min, and 4 to 5% H₂O₂. Enzyme treatment improved pulpability of wheat straw by the APMP process, and final pulp quality such as brightness, breaking length, and burst index of pulp. Pulp from the APMP process with enzyme treatment could be bleached to a brightness of 70.5% ISO by two-stage H₂O₂ bleaching sequence with only 4% H₂O₂, and breaking length of the bleach pulp reached 4470 m.     

Plant Growth Promotion by an Extracellular HAP-Phytase of a Thermophilic Mold <Emphasis Type="Italic">Sporotrichum thermophile</Emphasis>

Phytase of the thermophilic mold Sporotrichum thermophile Apinis hydrolyzed and liberated inorganic phosphate from Ca⁺², Mg⁺², and Co⁺² phytates more efficiently than those of Al³⁺, Fe²⁺, Fe³⁺, and Zn²⁺. The hydrolysis rate was higher at 60 °C as compared to 26 °C. Among all the organic acids tested, citrate was more effective in enhancing solubilization of insoluble phytate salts by phytase than others. The dry weight and inorganic phosphate contents of the wheat plants were high when supplemented with phytase or fungal spores. The plants provided with 5 mg phytate per plant exhibited enhanced growth and inorganic phosphate. With increase in the dosage of phytase, there was increase in growth and inorganic phosphate of plants, the highest being at 20 U per plant. The compost made employing the combined native microflora of the wheat straw and S. thermophile promoted growth of the plants. The plant-growth-promoting effect was also higher with the compost made using S. thermophile than that from only the native microflora.     

Ethanol production from steam-explosion pretreated wheat straw

Bioconversion of cereal straw to bioethanol is becoming an attractive alternative to conventional fuel ethanol production from grains. In this work, the best operational conditions for steam-explosion pretreatment of wheat straw for ethanol production by a simultaneous saccharification and fermentation process were studied, using diluted acid [H₂SO₄ 0.9% (w/w)] and water as preimpregnation agents. Acid-or water-impregnated biomass was steam-exploded at different temperatures (160–200°C) and residence times (5, 10, and 20 min). Composition of solid and filtrate obtained after pretreatment, enzymatic digestibility and ethanol production of pretreated wheat straw at different experimental conditions was analyzed. The best pretreatment conditions to obtain high conversion yield to ethanol (approx 80% of theoretical) of cellulose-rich residue after steam-explosion were 190°C and 10 min or 200°C and 5 min, in acid-impregnated straw. However, 180°C for 10 min in acid-impregnated biomass provided the highest ethanol yield referred to raw material (140 L/t wheat straw), and sugars recovery yield in the filtrate (300 g/kg wheat straw).     

High solids simultaneous saccharification and fermentation of pretreated wheat straw to ethanol

Wheat straw was pretreated with dilute (0.5%) sulfuric acid at 140°C for 1 h. Pretreated straw solids were washed with deionized water to neutrality and then stored frozen at –20°C. The approximate composition of the pretreated straw solids was 64% cellulose, 33% lignin, and 2% xylan. The cellulose in the pretreated wheat straw solids was converted to ethanol in batch simultaneous saccharification and fermentation experiments at 37°C using cellulase enzyme fromTrichoderma reesei (Genencor 150 L) with or without supplementation with β–glucosidase fromAspergillus niger (Novozyme 188) to produce glucose sugar and the yeastSaccharomyces cerevisiae to ferment the glucose into ethanol. The initial cellulose concentrations were adjusted to 7.5, 10, 12.5, 15, 17.5, and 20% (w/w). Since wheat straw particles do not form slurries at these concentrations and cannot be mixed with conventional impeller mixers used in laboratory fermenters, a simple rotary fermenter was designed and fabricated for these experiments. The results of the simultaneous saccharification and fermentation (SSF) experiments indicate that the cellulose in pretreated wheat straw can be efficiently fermented into ethanol for up to a 15% cellulose concentration (24.4% straw concentration).     

The Effect of Drying Temperature on the Composition of Biomass

The compositional quality of different lignocellulosic feedstocks influences their performance and potential demand at a biorefinery. Many analytical protocols for determining the composition or performance characteristics of biomass involve a drying step, where the drying temperature can vary depending on the specific protocol. To get reliable data, it is important to determine the correct drying temperature to vaporize the water without negatively impacting the compositional quality of the biomass. A comparison of drying temperatures between 45 °C and 100 °C was performed using wheat straw and corn stover. Near-infrared (NIR) spectra were taken of the dried samples and compared using principal component analysis (PCA). Carbohydrates were analyzed using quantitative saccharification to determine sugar degradation. Analysis of variance was used to determine if there was a significant difference between drying at different temperatures. PCA showed an obvious separation in samples dried at different temperatures due to sample water content. However, quantitative saccharification data shows, within a 95% confidence interval, that there is no significant difference in sugar content for drying temperatures up to 100 °C for wheat straw and corn stover.     

Effect of alkaline pretreatment on delignification of wheat straw

This study was conducted to analyse structural changes through scanning electron microscopy (SEM) and Fourier transform infrared (FTIR) after alkaline pretreatment of wheat straw for optimum steaming period. During the study, 2 mm size of substrate was soaked in 2.5% NaOH for 1 h at room temperature and then autoclaved at 121°C for various steaming time (30, 60, 90 and 120 min). Results revealed that residence time of 90 min at 121°C has strong effect on substrate, achieving a maximum cellulose content of 83%, delignification of 81% and hemicellulose content of 10.5%. Further SEM and FTIR spectroscopy confirmed structural modification caused by alkaline pretreatment in substrate. Maximum saccharification yield of 52.93% was achieved with 0.5% enzyme concentration using 2.5% substrate concentration for 8 h of incubation at 50°C. This result indicates that the above-mentioned pretreatment conditions create accessible areas for enzymatic hydrolysis.     

Controlled composting of straw

Production of composts on the farm from surplus straw might provide a low-cost biotechnological approach for increasing the value of this lignocellulosic waste. Successful composting depends on the conversion of the polysaccharides (cellulose and hemicelluloses) of straw by inoculated microorganisms to products that can promote plant growth when applied to the land. None of the potentially useful products we have identified are produced by cellulolytic organisms. We have therefore studied mixed populations in which noncellulolytic bacteria depend for growth on the products of fungal cellulolysis. The nature and yield of bacterial products depends not only on conditions within the compost, but also on the microbial inoculants used. Under defined laboratory conditions, using pure cellulose, N₂ is fixed by the anaerobic bacteriumClostridium butyricum in association with a cellulolytic fungus such asTrichoderma sp. A similar association has been achieved on straw withPenicillium corylophilum as the cellulolytic fungus. Cellulolytic fungi can also provide available substrates for the production of bacterial polysaccharides that can improve the structure of unstable soils. The yield of polysaccharide and its efficacy in soil aggregate stabilization again varies with the inoculants used. Such composts can thus contribute to plant nutrition and to soil structure. The adoption ofTrichoderma spp. as the cellulolytic inoculants would further extend the potential value of the compost to include the biocontrol of plant pathogens.     

Screening and Production Study of Microbial Xylanase Producers from Brazilian Cerrado

Hemicelluloses are polysaccharides of low molecular weight containing 100 to 200 glycosidic residues. In plants, the xylans or the hemicelluloses are situated between the lignin and the collection of cellulose fibers underneath. The xylan is the most common hemicellulosic polysaccharide in cell walls of land plants, comprising a backbone of xylose residues linked by β-1,4-glycosidic bonds. So, xylanolytic enzymes from microorganism have attracted a great deal of attention in the last decade, particularly because of their biotechnological characteristics in various industrial processes, related to food, feed, ethanol, pulp, and paper industries. A microbial screening of xylanase producer was carried out in Brazilian Cerrado area in Selviria city, Mato Grosso do Sul State, Brazil. About 50 bacterial strains and 15 fungal strains were isolated from soil sample at 35 °C. Between these isolated microorganisms, a bacterium Lysinibacillus sp. and a fungus Neosartorya spinosa as good xylanase producers were identified. Based on identification processes, Lysinibacillus sp. is a new species and the xylanase production by this bacterial genus was not reported yet. Similarly, it has not reported about xylanase production from N. spinosa. The bacterial strain P5B1 identified as Lysinibacillus sp. was cultivated on submerged fermentation using as substrate xylan, wheat bran, corn straw, corncob, and sugar cane bagasse. Corn straw and wheat bran show a good xylanase activity after 72 h of fermentation. A fungus identified as N. spinosa (strain P2D16) was cultivated on solid-state fermentation using as substrate source wheat bran, wheat bran plus sawdust, corn straw, corncob, cassava bran, and sugar cane bagasse. Wheat bran and corncobs show the better xylanase production after 72 h of fermentation. Both crude xylanases were characterized and a bacterial xylanase shows optimum pH for enzyme activity at 6.0, whereas a fungal xylanase has optimum pH at 5.0–5.5. They were stable in the pH range 5.0–10.0 and 5.5–8.5 for bacterial and fungal xylanase, respectively. The optimum temperatures were 55C and 60 °C for bacterial and fungal xylanase, respectively, and they were thermally stable up to 50 °C.     

Changes to the physicochemical characteristics of wheat straw by mechanical ultrafine grinding

To investigate changes on the physicochemical characteristics of wheat straw by mechanical ultrafine grinding, wheat straw powders of four different particle sizes and distributions were produced using a sieve-based Retsch ZM100 grind mill and CJM-SY-B ultrafine vibration grind mill. Changes on the microstructure and physicochemical characteristics of the different powders were assessed by scanning electron microscopy, X-ray diffractometry, X-ray photoelectron spectroscopy, Fourier transform infrared spectroscopy, thermogravimetric analysis and relevant standard laboratory analysis methods. Ultrafine grinding reduced the crystallite size and crystallinity of the wheat straw. New surfaces were exposed on the ultrafine powder with high levels of cellulose/hemicelluloses components but there was no apparent change in chemical structure. Wheat straw powders were smaller in size but had a higher bulk density (from 0.19 to 0.54 g/mL) and angle of repose (from 46.02° to 55.61°) and slide (from 37.26° to 41.00°). The hydration properties (water-holding capacity and swelling capacity) decreased with reduction in particle size of the wheat straw. Both the sieve-based and ultrafine powder exhibited a good ability to remove Pb²⁺ and Cd²⁺ and there was marginal improvement when using the ultrafine powder. The thermal stability of the ultrafine powder measured by thermogravimetric analysis decreased significantly because of the low cellulose crystallinity.     

Pyrolytic behaviour of different biomasses (angiosperms) (maize plants,straws, and wood) in low temperature pyrolysis

The thermal degradation of agricultural products and by-products (two kinds of maize plants, wheat, and barley straw) has been investigated by means of thermogravimetric/mass spectrometric analysis at heating rates from 1 to 10 °C/min. Large differences were found in the pyrolytic behaviour of the untreated samples, mainly caused by the high content of inorganics (ash content of about 4–6 wt%). These differences could be reduced by washing the samples with cold water. A kinetic model based on the formal kinetic parameters for the pyrolysis of the main components (hemicelluloses, lignin, and cellulose) and their degradable amounts was applied. To reduce the complexity of the model, only largely ash reduced samples were used. The formal kinetic parameters for the main components of barley straw and Gavott were individually determined. Although, different monomeric lignin degradation products were found for the angiosperms of grassy biomass in comparison to woody biomass, the formal kinetic parameters for lignin degradation are similar. The transferability of the formal kinetic parameters was successfully tested by applying them to a different straw type (wheat) and to a different maize cultivar (Doge) using the results of the biochemical analysis for the main components (hemicelluloses, lignin, and cellulose).     

Production, Purification, and Characterization of Exoglucanase by Aspergillus fumigatus

Fungi are considered good producers of industrially valuable enzymes with higher enzymatic activities. Among these cellulases are group of extracellular enzymes commonly employed in many industries for the hydrolysis of cellulolytic material. Aspergillus fumigatus produced exoglucanase having high enzymatic activity (83 U/gds) during the solid-state fermentation of wheat straw under optimum physical and nutritional conditions. Maximum production was obtained after 72 h of fermentation, at 55 °C temperature, pH 5.5, 80 % moisture level, and 2 mL fungal inoculum. Production was further increased by the addition of fructose (0.3 %) as additional carbon source, peptone (0.4 %) as nitrogen source, Tween-80 (0.3 %) as surfactant, and ammonium sulfate (0.2 %) in media. Exoglucanase was 2.30-folds purified by adding 40 % ammonium sulfate with volumetric activity 95.4 U/gds and specific activity 14.74 U/mg. Further, it was 5.18-folds purified by gel filtration chromatography with volumetric activity 115.2 U/gds and specific activity 33.10 U/mg. Purified exoglucanase has maximum activity at 55 °C and pH 4.8 using 1 % Avicel aqueous solution as substrate. The K _m and V _max were 4.34 mM and 7.29 μM/min, respectively. Calcium, magnesium, and zinc ions have positive effect on exoglucanase activity.     

Effect of rapid batch decompression on hydrolysate quality after hydrothermal pretreatment of wheat straw

The subject of this paper was to study the effect of rapid batch decompression on hydrolysate quality and on biogas yield after the hydrothermal pretreatment of wheat straw. An aqueous batch containing 5 mass % total solids of wheat straw was thermally and thermally-expansionary treated in parallel at the process temperature of 170–200°C and the residence time of 0–60 min. An analysis of the thermal and thermal-expansionary hydrolysate provided identical results in the dependences and values of chemical oxygen demand, acidities, and glucose yields of both treatments based on severity factors including the combined effects of temperature and residence time. Increases in the methane content of 33 % for thermally and of 34 % for thermally-expansionary treated wheat straw were reached in comparison to the methane yield from an untreated sample. This means that the polysaccharide cell wall was dissolved because of the high process temperature and residence time. From this it follows that all its nutrients were subsequently washed out of the cell into liquid where they caused changes in its chemical oxygen demand, glucose content, and acidities. There was therefore no rapid decompression effect on the hydrothermally treated wheat straw.     

Preliminary results on optimization of pilot scale pretreatment of wheat straw used in coproduction of bioethanol and electricity

The overall objective in this European Union-project is to develop cost and energy effective production systems for coproduction of bioethanol and electricity based on integrated biomass utilization. A pilot plan reactor for hydrothermal pretreatment (including weak acid hydrolysis, wet oxidation, and steam pretreatment) with a capacity of 100 kg/h was constructed and tested for pretreatment of wheat straw for ethanol production. Highest hemicellulose (C5 sugar) recovery and extraction of hemicellulose sugars was obtained at 190°C whereas highest C6 sugar yield was obtained at 200°C. Lowest toxicity of hydrolysates was observed at 190°C; however, addition of H₂O₂ improved the fermentability and sugar recoveries at the higher temperatures. The estimated total ethanol production was 223 kg/t straw assuming utilisation of both C6 and C5 during fermentation, and 0.5 g ethanol/g sugar.     

Ab initio studies of structural features not easily amenable to experiment. 38. Structural and conformational investigations of propanoic, 2-methylpropanoic,and butanoic acid

The structures and conformational energies of several conformations of propanoic acid, 2-methylpropanoic acid, and butanoic acid were determined by geometrically unconstrained ab initio gradient geometry refinement on the 4-21G level. The O?C? C? C torsional potentials of propanoic acid and butanoic acid are found to be practically identical. There are energy minima at 0° and 120°, and maxima in the 60° region and at 180°. In 2-methylpropanoic acid there are energy minima at H? C? C?O dihedral angles of 0° and 120°, and maxima at 60° and 180°. The exact positions of the maxima and minima of the H? C? C?O torsional potential of 2-methylpropanoic acid are found to be predictable from propanoic acid rotational-potential parameters. Some conformationally dependent, local geometry trends are discussed.     

Enhancing the Cellulose-Degrading Activity of Cellulolytic Bacteria CTL-6 (Clostridium thermocellum) by Co-Culture with Non-cellulolytic Bacteria W2-10 (Geobacillus sp.)

The effect of a non-cellulolytic bacterium W2-10 (Geobacillus sp.) on the cellulose-degrading activity of a cellulolytic bacterium CTL-6 (Clostridium thermocellum) was determined using cellulose materials (paper and straw) in peptone cellulose solution (PCS) medium under aerobic conditions. The results indicated that in the co-culture, addition of W2-10 resulted in a balanced medium pH, and may provide the required anaerobic environment for CTL-6. Overall, addition of W2-10 was beneficial to CTL-6 growth in the adverse environment of the PCS medium. In co-culture with W2-10, the CTL-6 cellulose degradation efficiency of filter paper and alkaline-treated wheat straw significantly increased up to 72.45 and 37.79 %, respectively. The CMCase activity and biomass of CTL-6 also increased from 0.23 U ml^?1 and 45.1 μg ml^?1 (DNA content) up to 0.47 U ml^?1 and 112.2 μg ml^?1, respectively. In addition, co-culture resulted in accumulation of acetate and propionate up to 4.26 and 2.76 mg ml^?1. This was a respective increase of 2.58 and 4.45 times, in comparison to the monoculture with CTL-6.     

Investigation on catalyzed combustion of wheat straw by thermal analysis

Combustion of wheat straw incorporating TiO₂, CuO and MnO₂ was investigated by means of thermal analysis carried out at 20 °C/min in the temperature range from 50 °C to 900 °C. Combustion characteristic indexes had been put forward to describe wheat straw combustion characteristics. All the results showed that the catalysis of the catalysts to the wheat straw combustion had been embodied in facilitation of the volatile matters release from wheat straw, which reduced the temperature of the maximum combustion rate, and the relative active sequence of catalysts to the ignition characteristic could be improved remarkably. The catalysis of different catalysts to the Devolatilization Index could be described as follows: MnO₂ > TiO₂ > CuO, and the relative active sequence of catalysts to the Combustion Characteristic Index could be described as follows: CuO > TiO₂ > MnO₂.     

Polymerization of coordinated monomers. V. Polymerization of methyl methacrylate–Lewis acid complexes

The polymerization of the complex of methyl methacrylate with stannic chloride, aluminum trichloride, or boron trifluoride was carried out in toluene solution at several temperatures in the range of 60° to ?78°C by initiation of α,α′-azobisisobutyronicrile or by irradiation with ultraviolet rays. The tacticities of the resulting polymers were determined by NMR spectroscopy. Both the 1:1 and the 2:1 methyl methacrylate–SnCl₄ complexes gave polymers with similar tacticities at the polymerization temperatures above ?60°C. With decreasing temperature below ?60°C, the isotacticity was more favored for the 2:1 complex, whereas the tacticities did not change for the 1:1 complex. On the ESR spectroscopy of the polymerization solution under the irradiation of ultraviolet rays at ?120°C, the 1:1 SnCl₄ complex gave a quintet, while the 2:1 SnCl₄ complex gave both a quintet and a sextet. The sextet became weaker with increasing temperature and disappeared at ?60°C. This behavior of the sextet corresponds to the change of the tacticities of polymer for the 2:1 SnCl₄ complex. An intra–intercomplex addition was suggested for the polymerization of the 2:1 complex, which took a cis-configuration on the basis of its infrared spectra. The sextet can be ascribed to the radical formed by the intracomplex addition reaction, while the quintet can correspond to that formed by the intercomplex addition reaction. The proportion of the intracomplex reaction was estimated to be about 0.25 at ?75°C, and the calculated value of the probability of isotactic diad addition of the intracomplex reaction was found to be almost unity.     

Fermentation of Biologically Pretreated Wheat Straw for Ethanol Production: Comparison of Fermentative Microorganisms and Process Configurations

The pretreatment of lignocellulosic biomass with white-rot fungi to produce bioethanol is an environmentally friendly alternative to the commonly used physico-chemical processes. After biological pretreatment, a solid substrate composed of cellulose, hemicellulose and lignin, the two latter with a composition lower than that of the initial substrate, is obtained. In this study, six microorganisms and four process configurations were utilised to ferment a hydrolysate obtained from wheat straw pretreated with the white-rot fungus Irpex lacteus. To enhance total sugars utilisation, five of these microorganisms are able to metabolise, in addition to glucose, most of the pentoses obtained after the hydrolysis of wheat straw by the application of a mixture of hemicellulolytic and cellulolytic enzymes. The highest overall ethanol yield was obtained with the yeast Pachysolen tannophilus. Its application in combination with the best process configuration yielded 163 mg ethanol per gram of raw wheat straw, which was between 23 and 35 % greater than the yields typically obtained with a conventional bioethanol process, in which wheat straw is pretreated using steam explosion and fermented with the yeast Saccharomyces cerevisiae.     

Characterization of Xylanase and Cellulase Produced by a Newly Isolated <Emphasis Type="Italic">Aspergillus fumigatus</Emphasis> N2 and Its Efficient Saccharification of Barley Straw

Aspergillus fumigatus N2 was isolated from decaying wood. This strain produces extracellular xylanases and cellulases. The highest xylanase (91.9 U/mL) and CMCase (5.61 U/mL) activity was produced when 1% barley straw was used as the carbon source. The optimum pH and temperature for xylanase activity were 6.0 and 65 °C, respectively. CMCase revealed maximum activity at pH 4.0 and in the range of 65 °C. The FPase was optimally active at pH 5.0 and 60 °C. The zymograms produced by the SDS-PAGE resolution of the crude enzymes indicated that multiple enzymes were secreted into the fermentation supernatant. Five bands of proteins with xylanase activity and four bands of proteins with endoglucanase were observed in the zymogram gel. The crude enzymes were used in the barley straw saccharification; an additive effect was observed when the commercial cellulase was added as supplement.