178 resultados para Metabolic flux analysis
Resumo:
An improved scaling analysis and direct numerical simulations are performed for the unsteady natural convection boundary layer adjacent to a downward facing inclined plate with uniform heat flux. The development of the thermal or viscous boundary layers may be classified into three distinct stages: a start-up stage, a transitional stage and a steady stage, which can be clearly identified in the analytical as well as the numerical results. Previous scaling shows that the existing scaling laws of the boundary layer thickness, velocity and steady state time scale for the natural convection flow on a heated plate of uniform heat flux provide a very poor prediction of the Prandtl number dependency of the flow. However, those scalings perform very well with Rayleigh number and aspect ratio dependency. In this study, a modified Prandtl number scaling is developed using a triple layer integral approach for Pr > 1. It is seen that in comparison to the direct numerical simulations, the modified scaling performs considerably better than the previous scaling.
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It is found in the literature that the existing scaling results for the boundary layer thickness, velocity and steady state time for the natural convection flow over an evenly heated plate provide a very poor prediction of the Prandtl number dependency of the flow. However, those scalings provide a good prediction of two other governing parameters’ dependency, the Rayleigh number and the aspect ratio. Therefore, an improved scaling analysis using a triple-layer integral approach and direct numerical simulations have been performed for the natural convection boundary layer along a semi-infinite flat plate with uniform surface heat flux. This heat flux is a ramp function of time, where the temperature gradient on the surface increases with time up to some specific time and then remains constant. The growth of the boundary layer strongly depends on the ramp time. If the ramp time is sufficiently long, the boundary layer reaches a quasi steady mode before the growth of the temperature gradient is completed. In this mode, the thermal boundary layer at first grows in thickness and then contracts with increasing time. However, if the ramp time is sufficiently short, the boundary layer develops differently, but after the wall temperature gradient growth is completed, the boundary layer develops as though the startup had been instantaneous.
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A new scaling analysis has been performed for the unsteady natural convection boundary layer under a downward facing inclined plate with uniform heat flux. The development of the thermal or viscous boundary layers may be classified into three distinct stages including an early stage, a transitional stage and a steady stage, which can be clearly identified in the analytical as well as numerical results. Earlier scaling shows that the existing scaling laws of the boundary layer thickness, velocity and steady state time scales for the natural convection flow on a heated plate of uniform heat flux provide a very poor prediction of the Prandtl number dependency. However, those scalings performed very well with Rayleigh number and aspect ratio dependency. In this study, a modifed Prandtl number scaling has been developed using a triple-layer integral approach for Pr > 1. It is seen that in comparison to the direct numerical simulations, the new scaling performs considerably better than the previous scaling.
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Aims: This study investigated the association between the basal (rest) insulin-signaling proteins, Akt, and the Akt substrate AS160, metabolic risk factors, inflammatory markers and aerobic fitness, in middle-aged women with varying numbers of metabolic risk factors for type 2 diabetes. Methods: Sixteen women (n = 16) aged 51.3+/-5.1 (mean +/-SD) years provided muscle biopsies and blood samples at rest. In addition, anthropometric characteristics and aerobic power were assessed and the number of metabolic risk factors for each participant was determined (IDF criteria). Results: The mean number of metabolic risk factors was 1.6+/-1.2. Total Akt was negatively correlated with IL-1 beta (r = -0.45, p = 0.046), IL-6 (r = -0.44, p = 0.052) and TNF-alpha (r = -0.51, p = 0.025). Phosphorylated AS160 was positively correlated with HDL (r = 0.58, p = 0.024) and aerobic fitness (r = 0.51, p = 0.047). Furthermore, a multiple regression analysis revealed that both HDL (t = 2.5, p = 0.032) and VO(2peak) (t = 2.4, p = 0.037) were better predictors for phosphorylated AS160 than TNF-alpha or IL-6 (p>0.05). Conclusions: Elevated inflammatory markers and increased metabolic risk factors may inhibit insulin-signaling protein phosphorylation in middle-aged women, thereby increasing insulin resistance under basal conditions. Furthermore, higher HDL and fitness levels are associated with an increased AS160 phosphorylation, which may in turn reduce insulin resistance.
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Siamese mud carp (Henichorynchus siamensis) is a freshwater teleost of high economic importance in the Mekong River Basin. However, genetic data relevant for delineating wild stocks for management purposes currently are limited for this species. Here, we used 454 pyrosequencing to generate a partial genome survey sequence (GSS) dataset to develop simple sequence repeat (SSR) markers from H. siamensis genomic DNA. Data generated included a total of 65,954 sequence reads with average length of 264 nucleotides, of which 2.79% contain SSR motifs. Based on GSS-BLASTx results, 10.5% of contigs and 8.1% singletons possessed significant similarity (E value < 10–5) with the majority matching well to reported fish sequences. KEGG analysis identified several metabolic pathways that provide insights into specific potential roles and functions of sequences involved in molecular processes in H. siamensis. Top protein domains detected included reverse transcriptase and the top putative functional transcript identified was an ORF2-encoded protein. One thousand eight hundred and thirty seven sequences containing SSR motifs were identified, of which 422 qualified for primer design and eight polymorphic loci have been tested with average observed and expected heterozygosity estimated at 0.75 and 0.83, respectively. Regardless of their relative levels of polymorphism and heterozygosity, microsatellite loci developed here are suitable for further population genetic studies in H. siamensis and may also be applicable to other related taxa.
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The development and design of electric high power devices with electromagnetic computer-aided engineering (EM-CAE) software such as the Finite Element Method (FEM) and Boundary Element Method (BEM) has been widely adopted. This paper presents the analysis of a Fault Current Limiter (FCL), which acts as a high-voltage surge protector for power grids. A prototype FCL was built. The magnetic flux in the core and the resulting electromagnetic forces in the winding of the FCL were analyzed using both FEM and BEM. An experiment on the prototype was conducted in a laboratory. The data obtained from the experiment is compared to the numerical solutions to determine the suitability and accuracy of the two methods.
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Lignocellulosic materials, such as sugar cane bagasse, a waste product of the sugarcane processing industry, agricultural residues and herbaceous crops, may serve as an abundant and comparatively cheap feedstock for largescale industrial fermentation, resulting in the production of marketable end-products. However, the complex structure of lignocellulosic materials, the presence of various hexose and pentose sugars in the hemicellulose component, and the presence of various compounds that inhibit the organisms selected for the fermentation process, all constitute barriers that add to the production costs and make full scale industrial production economically less feasible. The work presented in this thesis was conducted in order to screen microorganisms for ability to utilize pentose sugars derived from the sugar mill industrial waste. A large number of individual bacterial strains were investigated from hemi-cellulose rich material collected at the Proserpine and Maryborough sugar mills, notably soil samples from the mill sites. The research conducted to isolation of six pentose-capable Gram-positive organisms from the actinomycetes group by using pentose as a sole carbon source in the cultivation process. The isolates were identified as Corynebacterium glutamicum, Actinomyces odontolyticus, Nocardia elegans, and Propionibacterium freudenreichii all of which were isolated from the hemicellulose-enriched soil. Pentose degrading microbes are very rare in the environment, so this was a significant discovery. Previous research indicated that microbes could degrade pentose after genetic modification but the microbes discovered in this research were able to naturally utilize pentose. Six isolates, identified as four different genera, were investigated for their ability to utilize single sugars as substrates (glucose, xylose, arabinose or ribose), and also dual sugars as substrates (a hexose plus a pentose). The results demonstrated that C. glutamicum, A. odontolyticus, N. elegans, and P. freudenreichii were pentose-capable (able to grow using xylose or other pentose sugar), and also showed diauxie growth characteristics during the dual-sugar (glucose, in combination with xylose, arabinose or ribose) carbon source tests. In addition, it was shown that the isolates displayed very small differences in growth rates when grown on dual sugars as compared to single sugars, whether pentose or hexose in nature. The anabolic characteristics of C. glutamicum, A. odontolyticus, N. elegans and P. freudenreichii were subsequently investigated by qualitative analysis of their end-products, using high performance liquid chromatography (HPLC). All of the organisms produced arginine and cysteine after utilization of the pentose substrates alone. In addition, P. freudenreichii produced alanine and glycine. The end-product profile arising from culture with dual carbon sources was also tested. Interestingly, this time the product was different. All of them produced the amino acid glycine, when grown on a combination substrate-mix of glucose with xylose, and also glucose with arabinose. Only N. elegans was able to break down ribose, either singly or in combination with glucose, and the end-product of metabolism of the glucose plus ribose substrate combination was glutamic acid. The ecological analysis of microbial abundance in sugar mill waste was performed using denaturing gradient gel electrophoresis (DGGE) and also the metagenomic microarray PhyloChip method. Eleven solid samples and seven liquid samples were investigated. A very complex bacterial ecosystem was demonstrated in the seven liquid samples after testing with the PhyloChip method. It was also shown that bagasse leachate was the most different, compared to all of the other samples, by virtue of its richness in variety of taxa and the complexity of its bacterial community. The bacterial community in solid samples from Proserpine, Mackay and Maryborough sugar mills showed huge diversity. The information found from 16S rDNA sequencing results was that the bacterial genera Brevibacillus, Rhodospirillaceae, Bacillus, Vibrio and Pseudomonas were present in greatest abundance. In addition, Corynebacterium was also found in the soil samples. The metagenomic studies of the sugar mill samples demonstrate two important outcomes: firstly that the bagasse leachate, as potentially the most pentose-rich sample tested, had the most complex and diverse bacterial community; and secondly that the pentose-capable isolates that were initially discovered at the beginning of this study, were not amongst the most abundant taxonomic groups discovered in the sugar mill samples, and in fact were, as suspected, very rare. As a bioprospecting exercise, therefore, the study has discovered organisms that are naturally present, but in very small numbers, in the appropriate natural environment. This has implications for the industrial application of E-PUB, in that a seeding process using a starter culture will be necessary for industrial purposes, rather than simply assuming that natural fermentation might occur.
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In this study, a tandem LC-MS (Waters Xevo TQ) MRM-based MS method was developed for rapid, broad profiling of hydrophilic metabolites from biological samples, in either positive or negative ion modes without the need for an ion pairing reagent, using a reversed-phase pentafluorophenylpropyl (PFPP) column. The developed method was successfully applied to analyze various biological samples from C57BL/6 mice, including urine, duodenum, liver, plasma, kidney, heart, and skeletal muscle. As result, a total 112 of hydrophilic metabolites were detected within 8 min of running time to obtain a metabolite profile of the biological samples. The analysis of this number of hydrophilic metabolites is significantly faster than previous studies. Classification separation for metabolites from different tissues was globally analyzed by PCA, PLS-DA and HCA biostatistical methods. Overall, most of the hydrophilic metabolites were found to have a "fingerprint" characteristic of tissue dependency. In general, a higher level of most metabolites was found in urine, duodenum, and kidney. Altogether, these results suggest that this method has potential application for targeted metabolomic analyzes of hydrophilic metabolites in a wide ranges of biological samples.
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The promise of metabonomics, a new "omics" technique, to validate Chinese medicines and the compatibility of Chinese formulas has been appreciated. The present study was undertaken to explore the excretion pattern of low molecular mass metabolites in the male Wistar-derived rat model of kidney yin deficiency induced with thyroxine and reserpine as well as the therapeutic effect of Liu Wei Di Huang Wan (LW) and its separated prescriptions, a classic traditional Chinese medicine formula for treating kidney yin deficiency in China. The study utilized ultra-performance liquid chromatography/electrospray ionization synapt high definition mass spectrometry (UPLC/ESI-SYNAPT-HDMS) in both negative and positive electrospray ionization (ESI). At the same time, blood biochemistry was examined to identify specific changes in the kidney yin deficiency. Distinct changes in the pattern of metabolites, as a result of daily administration of thyroxine and reserpine, were observed by UPLC-HDMS combined with a principal component analysis (PCA). The changes in metabolic profiling were restored to their baseline values after treatment with LW according to the PCA score plots. Altogether, the current metabonomic approach based on UPLC-HDMS and orthogonal projection to latent structures discriminate analysis (OPLS-DA) indicated 20 ions (14 in the negative mode, 8 in the positive mode, and 2 in both) as "differentiating metabolites".
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This paper was designed to study metabonomic characters of the hepatotoxicity induced by alcohol and the intervention effects of Yin Chen Hao Tang (YCHT), a classic traditional Chinese medicine formula for treatment of jaundice and liver disorders in China. Urinary samples from control, alcohol- and YCHT-treated rats were analyzed by ultra-performance liquid chromatography/electrospray ionization quadruple time-of-flight mass spectrometry (UPLC/ESI-QTOF-MS) in positive ionization mode. The total ion chromatograms obtained from the control, alcohol- and YCHT-treated rats were easily distinguishable using a multivariate statistical analysis method such as the principal components analysis (PCA). The greatest difference in metabolic profiling was observed from alcohol-treated rats compared with the control and YCHT-treated rats. The positive ions m/z 664.3126 (9.00 min) was elevated in urine of alcohol-treated rats, whereas, ions m/z 155.3547 (10.96 min) and 708.2932 (9.01 min) were at a lower concentration compared with that in urine of control rats, however, these ions did not indicate a statistical difference between control rats and YCHT-treated rats. The ion m/z 664.3126 was found to correspond to ceramide (d18:1/25:0), providing further support for an involvement of the sphingomyelin signaling pathway in alcohol hepatotoxicity and the intervention effects of YCHT.
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BACKGROUND & AIMS Metabolomics is comprehensive analysis of low-molecular-weight endogenous metabolites in a biological sample. It could enable mapping of perturbations of early biochemical changes in diseases and hence provide an opportunity to develop predictive biomarkers that could provide valuable insights into the mechanisms of diseases. The aim of this study was to elucidate the changes in endogenous metabolites and to phenotype the metabolic profiling of d-galactosamine (GalN)-inducing acute hepatitis in rats by UPLC-ESI MS. METHODS The systemic biochemical actions of GalN administration (ip, 400 mg/kg) have been investigated in male wistar rats using conventional clinical chemistry, liver histopathology and metabolomic analysis of UPLC- ESI MS of urine. The urine was collected predose (-24 to 0 h) and 0-24, 24-48, 48-72, 72-96 h post-dose. Mass spectrometry of the urine was analysed visually and via conjunction with multivariate data analysis. RESULTS Results demonstrated that there was a time-dependent biochemical effect of GalN dosed on the levels of a range of low-molecular-weight metabolites in urine, which was correlated with developing phase of the GalN-inducing acute hepatitis. Urinary excretion of beta-hydroxybutanoic acid and citric acid was decreased following GalN dosing, whereas that of glycocholic acid, indole-3-acetic acid, sphinganine, n-acetyl-l-phenylalanine, cholic acid and creatinine excretion was increased, which suggests that several key metabolic pathways such as energy metabolism, lipid metabolism and amino acid metabolism were perturbed by GalN. CONCLUSION This metabolomic investigation demonstrates that this robust non-invasive tool offers insight into the metabolic states of diseases.
De Novo Transcriptome Sequence Assembly and Analysis of RNA Silencing Genes of Nicotiana benthamiana
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Background: Nicotiana benthamiana has been widely used for transient gene expression assays and as a model plant in the study of plant-microbe interactions, lipid engineering and RNA silencing pathways. Assembling the sequence of its transcriptome provides information that, in conjunction with the genome sequence, will facilitate gaining insight into the plant's capacity for high-level transient transgene expression, generation of mobile gene silencing signals, and hyper-susceptibility to viral infection. Methodology/Results: RNA-seq libraries from 9 different tissues were deep sequenced and assembled, de novo, into a representation of the transcriptome. The assembly, of16GB of sequence, yielded 237,340 contigs, clustering into 119,014 transcripts (unigenes). Between 80 and 85% of reads from all tissues could be mapped back to the full transcriptome. Approximately 63% of the unigenes exhibited a match to the Solgenomics tomato predicted proteins database. Approximately 94% of the Solgenomics N. benthamiana unigene set (16,024 sequences) matched our unigene set (119,014 sequences). Using homology searches we identified 31 homologues that are involved in RNAi-associated pathways in Arabidopsis thaliana, and show that they possess the domains characteristic of these proteins. Of these genes, the RNA dependent RNA polymerase gene, Rdr1, is transcribed but has a 72 nt insertion in exon1 that would cause premature termination of translation. Dicer-like 3 (DCL3) appears to lack both the DEAD helicase motif and second dsRNA binding motif, and DCL2 and AGO4b have unexpectedly high levels of transcription. Conclusions: The assembled and annotated representation of the transcriptome and list of RNAi-associated sequences are accessible at www.benthgenome.com alongside a draft genome assembly. These genomic resources will be very useful for further study of the developmental, metabolic and defense pathways of N. benthamiana and in understanding the mechanisms behind the features which have made it such a well-used model plant. © 2013 Nakasugi et al.
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The catalytic action of putrescine specific amine oxidases acting in tandem with 4-aminobutyraldehyde dehydrogenase is explored as a degradative pathway in Rhodococcus opacus. By limiting the nitrogen source, increased catalytic activity was induced leading to a coordinated response in the oxidative deamination of putrescine to 4-aminobutyraldehyde and subsequent dehydrogenation to 4-aminobutyrate. Isolating the dehydrogenase by ion exchange chromatography and gel filtration revealed that the enzyme acts principally on linear aliphatic aldehydes possessing an amino moiety. Michaelis-Menten kinetic analysis delivered a Michaelis constant (KM=0.014mM) and maximum rate (Vmax=11.2μmol/min/mg) for the conversion of 4-aminobutyraldehyde to 4-aminobutyrate. The dehydrogenase identified by MALDI-TOF mass spectrometric analysis (E value=0.031, 23% coverage) belongs to a functionally related genomic cluster that includes the amine oxidase, suggesting their association in a directed cell response. Key regulatory, stress and transport encoding genes have been identified, along with candidate dehydrogenases and transaminases for the further conversion of 4-aminobutyrate to succinate. Genomic analysis has revealed highly similar metabolic gene clustering among members of Actinobacteria, providing insight into putrescine degradation notably among Micrococcaceae, Rhodococci and Corynebacterium by a pathway that was previously uncharacterised in bacteria.
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The acyl composition of membrane phospholipids in kidney and brain of mammals of different body mass was examined. It was hypothesized that reduction in unsaturation index (number of double bonds per 100 acyl chains) of membrane phospholipids with increasing body mass in mammals would be made-up of similar changes in acyl composition across all phospholipid classes and that phospholipid class distribution would be regulated and similar in the same tissues of the different-sized mammals. The results of this study supported both hypotheses. Differences in membrane phospholipid acyl composition (i. e. decreased omega-3 fats, increased monounsaturated fats and decreased unsaturation index with increasing body size) were not restricted to any specific phospholipid molecule or to any specific phospholipid class but were observed in all phospholipid classes. With increase in body mass of mammals both monounsaturates and use of less unsaturated polyunsaturates increases at the expense of the long-chain highly unsaturated omega-3 and omega-6 polyunsaturates, producing decreases in membrane unsaturation. The distribution of membrane phospholipid classes was essentially the same in the different-sized mammals with phosphatidylcholine (PC) and phosphatidylethanolamine (PE) together constituting similar to 91% and similar to 88% of all phospholipids in kidney and brain, respectively. The lack of sphingomyelin in the mouse tissues and higher levels in larger mammals suggests an increased presence of membrane lipid rafts in larger mammals. The results of this study support the proposal that the physical properties of membranes are likely to be involved in changing metabolic rate.
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This study investigated the effects of high-intensity interval training (HIIT) vs. work-matched moderate-intensity continuous exercise (MOD) on metabolism and counterregulatory stress hormones. In a randomized and counterbalanced order, 10 well-trained male cyclists and triathletes completed a HIIT session [81.6 ± 3.7% maximum oxygen consumption (V̇o2 max); 72.0 ± 3.2% peak power output; 792 ± 95 kJ] and a MOD session (66.7 ± 3.5% V̇o2 max; 48.5 ± 3.1% peak power output; 797 ± 95 kJ). Blood samples were collected before, immediately after, and 1 and 2 h postexercise. Carbohydrate oxidation was higher (P = 0.037; 20%), whereas fat oxidation was lower (P = 0.037; −47%) during HIIT vs. MOD. Immediately after exercise, plasma glucose (P = 0.024; 20%) and lactate (P < 0.01; 5.4×) were higher in HIIT vs. MOD, whereas total serum free fatty acid concentration was not significantly different (P = 0.33). Targeted gas chromatography-mass spectromtery metabolomics analysis identified and quantified 49 metabolites in plasma, among which 11 changed after both HIIT and MOD, 13 changed only after HIIT, and 5 changed only after MOD. Notable changes included substantial increases in tricarboxylic acid intermediates and monounsaturated fatty acids after HIIT and marked decreases in amino acids during recovery from both trials. Plasma adrenocorticotrophic hormone (P = 0.019), cortisol (P < 0.01), and growth hormone (P < 0.01) were all higher immediately after HIIT. Plasma norepinephrine (P = 0.11) and interleukin-6 (P = 0.20) immediately after exercise were not significantly different between trials. Plasma insulin decreased during recovery from both HIIT and MOD (P < 0.01). These data indicate distinct differences in specific metabolites and counterregulatory hormones following HIIT vs. MOD and highlight the value of targeted metabolomic analysis to provide more detailed insights into the metabolic demands of exercise.