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Forschungszentrum Karlsruhe has developed a concept for the utilization of cereal straw and other thin-walled biomass with high ash content. The concept consists of a regional step (drying, chopping, flash-pyrolysis, and mixing) and a central one (pressurized entrained-flow gasification, gas cleaning, synthesis of fuel, and production of byproducts). The purpose of the regional plant is to prepare the biomass by minimizing its volume and producing a stable and safe storage and transport form. In the central gasifier, the pyrolysis products are converted into syngas. The syngas is tar-free and can be used for Fischer-Tropsch synthesis after gas cleaning.

Fatty acid methyl esters, also known as biodiesel, have been shown to have a great deal of potential as petro-diesel substitutes. Biodiesel comprise a renewable alternative energy source, the development of which would clearly reduce global dependence on petroleum, and would also help to reduce air pollution. This paper analyzes the fuel properties of rapeseed biodiesel and its blend with petro-diesel, as well as the emission profiles of a diesel engine on these fuels. Fuels performance studies were conducted in order to acquire comparative data regarding specific fuel consumption and exhaust emissions, including levels of carbon monoxide (CO), carbon dioxide (CO_2), smoke density, and NO_X, in an effort to assess the performance of these biodiesel and blend. The fuel consumption amount of oil operations at high loads was similar or greater than that observed during petro-diesel operation. The use of biodiesel is associated with lower smoke density than would be seen with petro-diesel. However, biodiesel and its blend increased the emission of CO, CO_2, and nitrogen oxides, to a greater degree than was seen with petro-diesel. The above results indicate that rapeseed biodiesel can be partially substituted for petro-diesel under most operating conditions, regarding both performance parameters and exhaust, without any modifications having to be made to the engine.

Enzymes are clearly recognized as a keystone technology for the production of fuels and chemicals from renewable feedstocks. Their specificity, performance under mild reaction conditions, and biodegradability make them ideally suited to widespread use in biorefineries around the world, and as the world puts greater and greater value on sustainable processes and environmentally friendly production methods, the further the development of enzyme technology grows in importance. This session focuses on the discovery, production, modification, and use of enzymes by bringing together 6 oral and 64 poster presentations describing the state of the art in enzyme technology.

Studies were conducted on maltodextrin saccharification and on simultaneous saccharification and fermentation (SSF) with various commercial glucoamylases. In kinetics studies, none of the glucoamylases were able to completely convert maltodextrin into glucose. Typically, about 85% conversion was obtained, and glucose yields were about 75%. Typically, the kinetics were biphasic, with 1 h of rapid conversion, then a significant reduction in rate. Data were consistent with strong product inhibition and/or enzyme inactivation. Some glucoamylases followed first-order kinetics, initially slower at dextrin conversion, but eventually achieving comparable conversion and glucose concentrations. Most of the glucoamylases were more active at 55°C than at 35°C, but pH had little effect on activity. Screening studies in an SSF system demonstrated little difference between the glucoamylases, with a few exceptions. Subsequent targeted studies showed clear differences in performance, depending on the fermentation temperature and yeast used, suggesting that these are key parameters that would guide the selection of a glucoamylase.

Convenient expression systems for efficient heterologous production of different laccases are needed for their characterization and application. The laccase cDNAs lcc1 and lcc2 from Trametes versicolor were expressed in Pichia pastoris and Aspergillus niger under control of their respective glyceraldehyde-3-phosphate dehydrogenase promoters and with the native secretion signal directing catalytically active laccase to the medium. P. pastoris batch cultures in shake-flasks gave higher volumetric activity (1.3 U/L) and a better activity to biomass ratio with glucose than with glycerol or maltose as carbon source. Preliminary experiments with fed-batch cultures of P. pastoris in bioreactors yielded higher activity (2.8 U/L) than the shake-flask experiments, although the levels remained moderate and useful primarily for screening purposes. With A. niger , high levels of laccase (2700 U/L) were produced using a minimal medium containing sucrose and yeast extract. Recombinant laccase from A. niger harboring the lcc2 cDNA was purified to homogeneity and it was found to be a 70-kDa homogeneous enzyme with biochemical and catalytic properties similar to those of native T. versicolor laccase A.

β-Galactosidase from the fungus Talaromyces thermophilus CBS 236.58 was immobilized by covalent attachment onto the insoluble carrier Eupergit C with a high binding efficiency of 95%. Immobilization increased both activity and stability at higher pH values and temperature when compared with the free enzyme. Especially the effect of immobilization on thermostability is notable. This is expressed by the half-lifetime of the activity at 50°C, which was determined to be 8 and 27 h for the free and immobilized enzymes, respectively. Although immobilization did not significantly change kinetic parameters for the substrate lactose, a considerable decrease in the maximum reaction velocity V _max was observed for the artificial substrate o-nitrophenyl-β- D -galactopyranoside (oNPG). The hydrolysis of both oNPG and lactose is competitively inhibited by the end products glucose and galactose. However, this inhibition is only very moderate as judged from kinetic analysis with glucose exerting a more pronounced inhibitory effect. It was evident from bioconversion experiments with 20% lactose as substrate, that the immobilized enzyme showed a strong transgalactosylation reaction, resulting in the formation of galactooligosaccharides (GalOS). The maximum yield of GalOS of 34% was obtained when the degree of lactose conversion was roughly 80%. Hence, this immobilized enzyme can be useful both for the cleavage of lactose at elevated temperatures, and the formation of GalOS, prebiotic sugars that have a number of interesting properties for food applications.

This work aims to evaluate cell recycle of a recombinant strain of Pichia pastoris GS115 on the Xylanase A (XynA) production of Thermomyces lanuginosus IOC-4145 in submerged fermentation. Fed-batch processes were carried out with methanol feeding at each 12 h and recycling cell at 24, 48, and 72 h. Additionally, the influence of the initial cell concentration was investigated. XynA production was not decreased with the recycling time, during four cell recycles, using an initial cell concentration of 2.5 g/L. The maximum activity was 14,050 U/L obtained in 24 h of expression. However, when the initial cell concentration of 0.25 g/L was investigated, the enzymatic activity was reduced by 30 and 75% after the third and fourth cycles, respectively. Finally, it could be concluded that the initial cell concentration influenced the process performance and the interval of cell recycle affected enzymatic production.

Cyclodextrin glycosyltransferase (CGTase) is an enzyme that produces cyclodextrins from starch by an intramolecular transglycosylation reaction. Cyclodextrins have been shown to have a number of applications in the food, cosmetic, pharmaceutical, and chemical industries. In the current study, the production of CGTase by Paenibacillus campinasensis strain H69-3 was examined in submerged and solid-state fermentations. P. campinasensis strain H69-3 was isolated from the soil, which grows at 45°C, and is a Gramvariable bacterium. Different substrate sources such as wheat bran, soybean bran, soybean extract, cassava solid residue, cassava starch, corn starch, and other combinations were used in the enzyme production. CGTase activity was highest in submerged fermentations with the greatest production observed at 48–72 h. The physical and chemical properties of CGTase were determined from the crude enzyme produced from submerged fermentations. The optimum temperature was found to be 70–75°C, and the activity was stable at 55°C for 1 h. The enzyme displayed two optimum pH values, 5.5 and 9.0 and was found to be stable between a pH of 4.5 and 11.0.

Foam fractionation has the potential to be a low-cost protein separation process; however, it may cause protein denaturation during the foaming process. In previous work with cellulase, artificial chaperones were integrated into the foam fractionation process in order to reduce the loss of enzymatic activity. In this study, other factors were introduced to further reduce the loss of cellulase activity: type of cyclodextrin, cyclodextrin concentration, dilution ratio cyclodextrin to the foamate and holding time. α-Cyclodextrin was almost as effective as β-cyclodextrin in refolding the foamed cellulase-Cetyltrimethylammonium bromide mixture. β-Cyclodextrin (6.5 m M ) was almost as effective as 13 m M β-cyclodextrin in refolding. The dilution ratio, seven parts foamate and three parts β-cyclodextrin solution, was found to be most effective among the three ratios tested (7:3, 1:1, and 3:7). The activity after refolding at this dilution ratio is around 0.14 unit/mL The refolding time study showed that the refolding process was found to be most effective for the short refolding times (within 1 h).

Biotechnology production of xylitol is an excellent alternative to the industrial chemical process for the production of this polyalcohol. In this work the behavior of Candida guilliermondii yeast was studied when crucial process variables were modified. The K _La (between 18 and 40/h) and the initial cell mass (between 4 and 10 g) were considered as control variables. A response surface methodology was applied to the experimental design to study the resulting effect when the control variables were modified. A regression model was developed and used to determine an optimal value that was further validated experimentally. The optimal values determined for K _La and X _O were 32.85/h and 9.86 g, respectively, leading to maximum values for productivity (1.628 g/h) and xylitol yield (0.708 g/g).