Biorefining of wood: combined production of ethanol and xylanase from waste fiber sludge

The possibility to utilize fiber sludge, waste fibers from pulp mills and lignocellulose-based biorefineries, for combined production of liquid biofuel and biocatalysts was investigated. Without pretreatment, fiber sludge was hydrolyzed enzymatically to monosaccharides, mainly glucose and xylose. In the first of two sequential fermentation steps, the fiber sludge hydrolysate was fermented to cellulosic ethanol with the yeast Saccharomyces cerevisiae. Although the final ethanol yields were similar, the ethanol productivity after 9.5?h was 3.3?g/l/h for the fiber sludge hydrolysate compared with only 2.2?g/l/h for a reference fermentation with similar sugar content. In the second fermentation step, the spent fiber sludge hydrolysate (the stillage obtained after distillation) was used as growth medium for recombinant Aspergillus niger expressing the xylanase-encoding Trichoderma reesei (Hypocrea jecorina) xyn2 gene. The xylanase activity obtained with the spent fiber sludge hydrolysate (8,500?nkat/ml) was higher than that obtained in a standard medium with similar monosaccharide content (1,400?nkat/ml). Analyses based on deglycosylation with N-glycosidase?F suggest that the main part of the recombinant xylanase was unglycosylated and had molecular mass of 20.7?kDa, while a minor part had N-linked glycosylation and molecular mass of 23.6?kDa. Chemical analyses of the growth medium showed that important carbon sources in the spent fiber sludge hydrolysate included xylose, small aliphatic acids, and oligosaccharides. The results show the potential of converting waste fiber sludge to liquid biofuel and enzymes as coproducts in lignocellulose-based biorefineries.     

Inhibitory effects of HgCl2 on excitation-secretion coupling at the motor nerve terminal and excitation-contraction coupling in the muscle cell

Summary 1. Indirect and direct twitch (0.1-Hz) stimulation of the rat phrenic nerve-diaphragm disclosed that the inhibitory effect of HgCl₂, 3.7 × 10^–5 M, on the neuromuscular transmission and in the muscle cell, was accelerated by 10-sec periods of 50-Hz tetanic stimulation every 10 min. This activity-dependent enhancement suggested an inhibitory mechanism of HgCl₂ related to the development of fatigue, like membrane depolarization or decreased excitability, decreased availability of transmitter, or interference with the factors controlling excitation-secretion coupling of the nerve terminal, i.e. (Ca²⁺)₀ or (Ca²⁺)_i, and excitation-contraction coupling in the muscle cell, i.e., (Ca²⁺)_i.2. During both indirect and direct stimulation, HgCl₂-induced inhibition was enhanced markedly by pretreatment with caffeine, which releases Ca²⁺ from endoplasmic and sarcoplasmic reticulum in the nerve terminal and muscle cell, respectively. This caffeine-induced enhancement was completely antagonized by dantrolene, which inhibits the caffeine-induced release. However, dantrolene alone did not antagonize the HgCl₂-induced inhibition.3. Since caffeine depletes the intracellular Ca²⁺ stores of the smooth endoplasmic reticulum, HgCl₂ probably inhibits by binding to SH groups of transport proteins conveying the messenger function of (Ca²⁺)_i. In the muscle cell this leads to inhibition of contraction. In the nerve terminal, an additional enhancement of the HgCl₂-induced inhibition, by inhibiting reuptake of choline by TEA and tetanic stimulation, suggested that HgCl₂ inhibited a (Ca²⁺)_i signal necessary for this limiting factor in resynthesis of acetylcholine.4. The (Ca²⁺)₀ signal necessary for stimulus-induced release of acetylcholine was not affected by HgCl₂. Hyperpolarization in K⁺-free solution antagonized the inhibitory effect of HgCl₂ at indirect stimulation, and Ca²⁺-free solution enhanced the inhibitory effect at direct stimulation. K⁺ depolarization, membrane electric field increase with high Ca²⁺, membrane stabilization with lidocaine, and half-threshold stimulation, did not change the inhibitory effect of HgCl CH₃HgCl, 1.85 × 10^–5 M, disclosed a synergistic interaction with caffeine during direct, but not during indirect, stimulation.     

Additions to the lichen family Pannariaceae in Ecuador

Collections from high altitudes (3840–5100 m a.s.l.) have added several species to the Pannariaceae of Ecuador: Fuscopannaria praetermissa, Pannaria hookeri, Psoroma paleaceum and Psoroma tenue are new to the Andes, and phytogeographic consequences of these are discussed. The following new taxa are described: Parmeliella corallina P. M. Jørg. & Palice, Parmeliella psoromoides P. M. Jørg. & Palice, Psoroma cinnamomeum ssp. andinum P. M. Jørg. & Palice and Santessoniella macrospora P. M. Jørg. & Palice. They are part of a high‐andean element which is not well known and is indicative of a migration route between the polar regions. In addition, two species with associated cyanobacteria, as the primary or secondary symbiont, are reported for the first time from South America: Thelignya lignyota (Wahlenb.) P. M. Jørg. & Henssen and Pilophorus cereolus (Ach.) Th. Fr, both originating from the Northern Hemisphere.     

The opioid systems - Panacea and nemesis

This mini-review outlines the opioid systems and their roles primarily as related to reward and compulsive drug/alcohol intake. The central role is taken by the mu-opioid receptor, target for opiate analgesics and also a central target in compulsive alcohol abuse, alcoholism. The mu-opioid receptor and the cognate opioid neuropeptides from proenkephalin and proopiomelancortin are members of a superfamily of opioid systems, each with unique and still to be defined roles in the central nervous system.     

Novel α-L-Fucosidases from a Soil Metagenome for Production of Fucosylated Human Milk Oligosaccharides

This paper describes the discovery of novel α-L-fucosidases and evaluation of their potential to catalyse the transglycosylation reaction leading to production of fucosylated human milk oligosaccharides. Seven novel α-L-fucosidase-encoding genes were identified by functional screening of a soil-derived metagenome library and expressed in E. coli as recombinant 6xHis-tagged proteins. All seven fucosidases belong to glycosyl hydrolase family 29 (GH 29). Six of the seven α-L-fucosidases were substrate-inhibited, moderately thermostable and most hydrolytically active in the pH range 6–7, when tested with para-nitrophenyl-α-L-fucopyranoside (pNP-Fuc) as the substrate. In contrast, one fucosidase (Mfuc6) exhibited a high pH optimum and an unusual sigmoidal kinetics towards pNP-Fuc substrate. When tested for trans-fucosylation activity using pNP-Fuc as donor, most of the enzymes were able to transfer fucose to pNP-Fuc (self-condensation) or to lactose. With the α-L-fucosidase from Thermotoga maritima and the metagenome-derived Mfuc5, different fucosyllactose variants including the principal fucosylated HMO 2’-fucosyllactose were synthesised in yields of up to ~6.4%. Mfuc5 was able to release fucose from xyloglucan and could also use it as a fucosyl-donor for synthesis of fucosyllactose. This is the first study describing the use of glycosyl hydrolases for the synthesis of genuine fucosylated human milk oligosaccharides.     

Children’s Phthalate Intakes and Resultant Cumulative Exposures Estimated from Urine Compared with Estimates from Dust Ingestion,Inhalation and Dermal Absorption in Their Homes and Daycare Centers

Total daily intakes of diethyl phthalate (DEP), di(n-butyl) phthalate (DnBP), di(isobutyl) phthalate (DiBP), butyl benzyl phthalate (BBzP) and di(2-ethylhexyl) phthalate (DEHP) were calculated from phthalate metabolite levels measured in the urine of 431 Danish children between 3 and 6 years of age. For each child the intake attributable to exposures in the indoor environment via dust ingestion, inhalation and dermal absorption were estimated from the phthalate levels in the dust collected from the child’s home and daycare center. Based on the urine samples, DEHP had the highest total daily intake (median: 4.42 µg/d/kg-bw) and BBzP the lowest (median: 0.49 µg/d/kg-bw). For DEP, DnBP and DiBP, exposures to air and dust in the indoor environment accounted for approximately 100%, 15% and 50% of the total intake, respectively, with dermal absorption from the gas-phase being the major exposure pathway. More than 90% of the total intake of BBzP and DEHP came from sources other than indoor air and dust. Daily intake of DnBP and DiBP from all exposure pathways, based on levels of metabolites in urine samples, exceeded the Tolerable Daily Intake (TDI) for 22 and 23 children, respectively. Indoor exposures resulted in an average daily DiBP intake that exceeded the TDI for 14 children. Using the concept of relative cumulative Tolerable Daily Intake (TDI_cum), which is applicable for phthalates that have established TDIs based on the same health endpoint, we examined the cumulative total exposure to DnBP, DiBP and DEHP from all pathways; it exceeded the tolerable levels for 30% of the children. From the three indoor pathways alone, several children had a cumulative intake that exceeded TDI_cum. Exposures to phthalates present in the air and dust indoors meaningfully contribute to a child’s total intake of certain phthalates. Such exposures, by themselves, may lead to intakes exceeding current limit values.     

Stronger dung removal in forests compared with grassland is driven by trait composition and biomass of dung beetles

1. Ecosystem processes depend on the biomass of the involved organisms, but their functional diversity may play an additional role. In particular, the exclusion of key functional groups through habitat disturbance may lead to the breakdown of ecosystem functions. Dung removal is an important process contributing to nutrient cycling and thus productivity in grazed ecosystems. 2. This study investigated the role of different functional groups of dung beetles in dung removal in different habitats within a wood-pasture in two different seasons. An experimental setting with 12 blocks and 108 dung pads was used to investigate short-term dung removal over 1 week of exposure. 3. Dung removal was most strongly affected by habitat type, with almost 40% lower levels in grassland than in adjacent forest and forest gaps. Of all assemblage characteristics, total biomass of tunneller species was the strongest predictor of dung removal, whereas functional diversity showed no significant effect. In accordance with the dung removal pattern at habitat type level, densities of large tunnellers were suppressed in grassland compared with forest. 4. It is concluded that dung removal is habitat-specific and large tunnellers play a disproportionate role in this important ecosystem function in temperate forests.     

SuperSAGE analysis of the <Emphasis Type="Italic">Nicotiana attenuata</Emphasis> transcriptome after fatty acid-amino acid elicitation (FAC): identification of early mediators of insect responses

Background  

Plants trigger and tailor defense responses after perception of the oral secretions (OS) of attacking specialist lepidopteran larvae. Fatty acid-amino acid conjugates (FACs) in the OS of the Manduca sexta larvae are necessary and sufficient to elicit the herbivory-specific responses in Nicotiana attenuata, an annual wild tobacco species. How FACs are perceived and activate signal transduction mechanisms is unknown.