GWAS of butyrylcholinesterase activity identifies four novel loci, independent effects within BCHE and secondary associations with metabolic risk factors

Serum butyrylcholinesterase (BCHE) activity is associated with obesity, blood pressure and biomarkers of cardiovascular and diabetes risk. We have conducted a genome-wide association scan to discover genetic variants affecting BCHE activity, and to clarify whether the associations between BCHE activity and cardiometabolic risk factors are caused by variation in BCHE or whether BCHE variation is secondary to the metabolic abnormalities. We measured serum BCHE in adolescents and adults from three cohorts of Australian twin and family studies. The genotypes from approximately 2.4 million single-nucleotide polymorphisms (SNPs) were available in 8791 participants with BCHE measurements. We detected significant associations with BCHE activity at three independent groups of SNPs at the BCHE locus (P = 5.8 x 10(-262), 7.8 x 10(-47), 2.9 x 10(-12)) and at four other loci: RNPEP (P = 9.4 x 10(-16)), RAPH1-ABI2 (P = 4.1 x 10(-18)), UGT1A1 (P = 4.0 x 10(-8)) and an intergenic region on chromosome 8 (P = 1.4 x 10(-8)). These loci affecting BCHE activity were not associated with metabolic risk factors. On the other hand, SNPs in genes previously associated with metabolic risk had effects on BCHE activity more often than can be explained by chance. In particular, SNPs within FTO and GCKR were associated with BCHE activity, but their effects were partly mediated by body mass index and triglycerides, respectively. We conclude that variation in BCHE activity is due to multiple variants across the spectrum from uncommon/large effect to common/small effect, and partly results from (rather than causes) metabolic abnormalities.

Genetics of Cardiovascular Disease: A Candidate Gene Study of USF1

Cardiovascular diseases (CVD) are major contributors to morbidity and mortality worldwide. Several interacting environmental, biochemical, and genetic risk factors can increase disease susceptibility. While some of the genes involved in the etiology of CVD are known, many are yet to be discovered. During the last few decades, scientists have searched for these genes with genome-wide linkage and association methods, and with more targeted candidate gene studies. This thesis investigates variation within the upstream transcription factor 1 (USF1) gene locus in relation to CVD risk factors, atherosclerosis, and incidence and prevalence of CVD. This candidate gene was first identified in Finnish families ascertained for familial combined hyperlipidemia, a common dyslipidemia predisposing to coronary heart disease. The gene is a ubiquitously expressed transcription factor regulating expression of several genes from lipid and glucose metabolism, inflammation, and endothelial function. First, we examined association between USF1 variants and several CVD risk factors, such as lipid phenotypes, body composition measures, and metabolic syndrome, in two prospective population cohorts. Our data suggested that USF1 contributes to these CVD risk factors at the population level. Notably, the associations with quantitative measurements were mostly detected among study subjects with CVD or metabolic syndrome, suggesting complex interactions between USF1 effects and the pathophysiological state of an individual. Second, we investigated how variation at the USF1 locus contributes to atherosclerotic lesions of the coronary arteries and abdominal aorta. For this, we used two study samples of middle-aged men with detailed measurements of atherosclerosis obtained in autopsy. USF1 variation significantly associated with areas of several types of lesions, especially with calcification of the arteries. Next, we tested what effect the USF1 risk variants have on sudden cardiac death and incidence of CVD. The atherosclerosis-associated risk variant increased the risk of sudden cardiac death of the same study subjects. Furthermore, USF1 alleles associated with incidence of CVD in the Finnish population follow-up cohorts. These associations were especially prominent among women, suggesting a sex specific effect, which has also been detected in subsequent studies. Finally, as some of the low-yield DNA samples of the Finnish follow-up study cohort needed to be whole-genome amplified (WGA) prior to genotyping, we evaluated whether the produced WGA genotypes were of good quality. Although the samples giving genotype discrepancies could not be detected before genotyping with standard laboratory quality control methods, our results suggested that enhanced quality control at the time of the genotyping could identify such samples. In addition, combining two WGA reactions into one pooled DNA sample for genotyping markedly reduced the number of discrepancies and samples showing them. In conclusion, USF1 seems to have a role in the etiology of CVD. Additional studies are warranted to identify functional variants and to study interactions between USF1 and other genetic or environmental factors. This USF1 study, and other studies with low DNA yield of some samples, can benefit from whole genome amplification of the low-yield samples prior to genotyping. Careful quality control procedures are, however, needed in WGA genotyping.

Round scallops and square meshes: a comparison of four codend types on the catch rates of target species and by-catch in the Queensland (Australia) saucer scallop (Amusium balloti) trawl fishery

Concern over the amount of by-catch from benthic trawl fisheries and research into the problem have increased in recent years. The present paper demonstrated that by-catch rates in the Queensland (Australia) saucer scallop (Amusium balloti) trawl fishery can be reduced by 77% (by weight) using nets fitted with a turtle excluder device (TED) and a square-mesh codend, compared with a standard diamond-mesh codend with no TED. This large reduction was achieved with no significant effect on the legal size scallop catch rate and 39% fewer undersize scallops were caught. In total, 382 taxa were recorded in the by-catch, which was dominated by sponges, portunid crabs, small demersal and benthic fish (e.g. leatherjackets, stingerfish, bearded ghouls, nemipterids, longspine emperors, lizard fish, triggerfish, flounders and rabbitfish), elasmobranchs (e.g. mainly rays) and invertebrates (e.g. sea stars, sea urchins, sea cucumbers and bivalve molluscs). Extremely high reductions in catch rate (i.e. ≥85%) were demonstrated for several by-catch species owing to the square-mesh codend. Square-mesh codends show potential as a means of greatly reducing by-catch and lowering the incidental capture and mortality of undersize scallops and Moreton Bay bugs (Thenus australiensis) in this fishery

Diversity and abundance of fungivorous thrips (Thysanoptera) associated with leaf-litter and bark across forest types and two tree genera in subtropical Australia

Leaf-litter thrips were much more common and diverse in dry sclerophyll forest than in wetter forest types in subtropical southeast Queensland, Australia. In dry sclerophyll forest, the species composition of thrips in leaf-litter was strongly differentiated from the thrips fauna associated with bark of the trees Eucalyptus major and Acacia melanoxylon (4 of 34 species in common). The species composition of bark-dwelling thrips was similar across the two tree species and also across two eucalypts with different bark types, Eucalyptus major (flaky) and Eucalyptus siderophloia (rough). The diversity of thrips from the leaf-litter was not differentiated across all of these tree species. Virtually all thrips collected were Phlaeothripidae, subfamilies Idolothripinae and Phlaeothripinae. Idolothripinae were associated almost exclusively with leaf-litter, but Phlaeothripinae were in leaf-litter and bark. The association of fungal-feeding thrips with dry sclerophyll forest raises questions about their ecological requirements and the role they play in nutrient cycling. © 2012 Copyright Taylor and Francis Group, LLC.

A core metabolic enzyme mediates resistance to phosphine gas

Phosphine is a small redox-active gas that is used to protect global grain reserves, which are threatened by the emergence of phosphine resistance in pest insects. We find that polymorphisms responsible for genetic resistance cluster around the redox-active catalytic disulfide or the dimerization interface of dihydrolipoamide dehydrogenase (DLD) in insects (Rhyzopertha dominica and Tribolium castaneum) and nematodes (Caenorhabditis elegans). DLD is a core metabolic enzyme representing a new class of resistance factor for a redox-active metabolic toxin. It participates in four key steps of core metabolism, and metabolite profiles indicate that phosphine exposure in mutant and wild-type animals affects these steps differently. Mutation of DLD in C. elegans increases arsenite sensitivity. This specific vulnerability may be exploited to control phosphine-resistant insects and safeguard food security.

Influence of oxygen content of the certain types of biodiesels on particulate oxidative potential

Oxidative potential (OP) is related to the organic phase, specifically to its oxygenated organic fraction (OOA). Furthermore, the oxygen content of fuel molecules has significant influence on particulate OP. Thus, this study aimed to explore the actual dependency of the OOA and ROS to the oxygen content of the fuel. In order to reach the goal, different biodiesels blends, with various ranges of oxygen content; have been employed. The compact time of flight aerosol mass spectrometer (c-ToF AMS) enabled better identification of OOA. ROS monitored by using two assays: DTT and BPEA-nit. Despite emitting lower mass, both assays agreed that oxygen content of a biodiesel is directly correlated with its OOA, and highly related to its OP. Hence, the more oxygen included in the considered biodiesels, the higher the OP of PM emissions. This highlights the importance of taking oxygen content into account while assessing emissions from new fuel types, which is relevant from a health effects standpoint.

Structures of some unusual types of organic compounds

Structures of a variety of compounds isolated in reactions and elucidated with the help of spectral (uv,ir,nmr and mass) data, have been discussed. In a few cases, the assigned structures were confirmed by x-ray crystal structure analysis.

Crinkles in connectivity: combining genetics and other types of biological data to estimate movement and interbreeding between populations

Marine species generally have large population sizes, continuous distributions and high dispersal capacity. Despite this, they are often subdivided into separate populations, which are the basic units of fisheries management. For example, populations of some fisheries species across the deep water of the Timor Trench are genetically different, inferring minimal movement and interbreeding. When connectivity is higher than the Timor Trench example, but not so high that the populations become one, connectivity between populations is crinkled. Crinkled connectivity occurs when migration is above the threshold required to link populations genetically, but below the threshold for demographic links. In future, genetic estimates of connectivity over crinkled links could be uniquely combined with other data, such as estimates of population size and tagging and tracking data, to quantify demographic connectedness between these types of populations. Elasmobranch species may be ideal targets for this research because connectivity between populations is more likely to be crinkled than for finfish species. Fisheries stock-assessment models could be strengthened with estimates of connectivity to improve the strategic and sustainable harvesting of biological resources.

On various types of decoupling for time-varying system

This paper deals with the problem of decoupling a class of linear time-varying multi-variable systems, based on the defining property that the impulse response matrix of a decoupled system is diagonal. Depending on the properties of the coefficient matrices of the vector differential equation of the open-loop system, the system may be uniformly or totally decoupled. The necessary and sufficient conditions that permit a system to be uniformly or totally decoupled by state variable feedback are given. The main contribution of this paper is the precise definition of these two classes of decoupling and a rigorous derivation of the necessary and sufficient conditions which show the necessity of requiring that the system be of constant ordered rank with respect to observability. A simple example illustrates the importance of having several definitions of decoupling. Finally, the results are specialized to the case of time invariant systems.

Effect of biochar amendment on yield and photosynthesis of peanut on two types of soils

Abstract Biochar has significant potential to improve crop performance. This study examined the effect of biochar application on the photosynthesis and yield of peanut crop grown on two soil types. The commercial peanut cultivar Middleton was grown on red ferrosol and redoxi-hydrosol (Queensland, Australia) amended with a peanut shell biochar gradient (0, 0.375, 0.750, 1.50, 3.00 and 6.00 %, w/w, equivalent up to 85 t ha−1) in a glasshouse pot experiment. Biomass and pod yield, photosynthesis-[CO2] response parameters, leaf characteristics and soil properties (carbon, nitrogen (N) and nutrients) were quantified. Biochar significantly improved peanut biomass and pod yield up to 2- and 3-folds respectively in red ferrosol and redoxi-hydrosol. A modest (but significant) biochar-induced improvement of the maximumelectron transport rate and saturating photosynthetic rate was observed for red ferrosol. This response was correlated to increased leaf N and accompanied with improved soil available N and biological N fixation. Biochar application also improved the availability of other soil nutrients, which appeared critical in improving peanut performance, especially on infertile redoxihydrosol. Our study suggests that application of peanut shell derived biochar has strong potential to improve peanut yield on red ferrosol and redoxi-hydrosol. Biochar soil amendment can affect leaf N status and photosynthesis, but the effect varied with soil type.

Precipitation-induced runoff and leaching from milled peat mining mires by peat types : a comparative method for estimating the loading of water bodies during peat pruduction

Precipitation-induced runoff and leaching from milled peat mining mires by peat types: a comparative method for estimating the loading of water bodies during peat production. This research project in environmental geology has arisen out of an observed need to be able to predict more accurately the loading of watercourses with detrimental organic substances and nutrients from already existing and planned peat production areas, since the authorities capacity for insisting on such predictions covering the whole duration of peat production in connection with evaluations of environmental impact is at present highly limited. National and international decisions regarding monitoring of the condition of watercourses and their improvement and restoration require more sophisticated evaluation methods in order to be able to forecast watercourse loading and its environmental impacts at the stage of land-use planning and preparations for peat production.The present project thus set out from the premise that it would be possible on the basis of existing mire and peat data properties to construct estimates for the typical loading from production mires over the whole duration of their exploitation. Finland has some 10 million hectares of peatland, accounting for almost a third of its total area. Macroclimatic conditions have varied in the course of the Holocene growth and development of this peatland, and with them the habitats of the peat-forming plants. Temperatures and moisture conditions have played a significant role in determining the dominant species of mire plants growing there at any particular time, the resulting mire types and the accumulation and deposition of plant remains to form the peat. The above climatic, environmental and mire development factors, together with ditching, have contributed, and continue to contribute, to the existence of peat horizons that differ in their physical and chemical properties, leading to differences in material transport between peatlands in a natural state and mires that have been ditched or prepared for forestry and peat production. Watercourse loading from the ditching of mires or their use for peat production can have detrimental effects on river and lake environments and their recreational use, especially where oxygen-consuming organic solids and soluble organic substances and nutrients are concerned. It has not previously been possible, however, to estimate in advance the watercourse loading likely to arise from ditching and peat production on the basis of the characteristics of the peat in a mire, although earlier observations have indicated that watercourse loading from peat production can vary greatly and it has been suggested that differences in peat properties may be of significance in this. Sprinkling is used here in combination with simulations of conditions in a milled peat production area to determine the influence of the physical and chemical properties of milled peats in production mires on surface runoff into the drainage ditches and the concentrations of material in the runoff water. Sprinkling and extraction experiments were carried out on 25 samples of milled Carex (C) and Sphagnum (S) peat of humification grades H 2.5 8.5 with moisture content in the range 23.4 89% on commencement of the first sprinkling, which was followed by a second sprinkling 24 hours later. The water retention capacity of the peat was best, and surface runoff lowest, with Sphagnum and Carex peat samples of humification grades H 2.5 6 in the moisture content class 56 75%. On account of the hydrophobicity of dry peat, runoff increased in a fairly regular manner with drying of the sample from 55% to 24 30%. Runoff from the samples with an original moisture content over 55% increased by 63% in the second round of sprinkling relative to the first, as they had practically reached saturation point on the first occasion, while those with an original moisture content below 55% retained their high runoff in the second round, due to continued hydrophobicity. The well-humified samples (H 6.5 8.5) with a moisture content over 80% showed a low water retention capacity and high runoff in both rounds of sprinkling. Loading of the runoff water with suspended solids, total phosphorus and total nitrogen, and also the chemical oxygen demand (CODMn O2), varied greatly in the sprinkling experiment, depending on the peat type and degree of humification, but concentrations of the same substances in the two sprinklings were closely or moderately closely correlated and these correlations were significant. The concentrations of suspended solids in the runoff water observed in the simulations of a peat production area and the direct surface runoff from it into the drainage ditch system in response to rain (sprinkling intensity 1.27 mm/min) varied c. 60-fold between the degrees of humification in the case of the Carex peats and c. 150-fold for the Sphagnum peats, while chemical oxygen demand varied c. 30-fold and c. 50-fold, respectively, total phosphorus c. 60-fold and c. 66-fold, total nitrogen c. 65-fold and c. 195-fold and ammonium nitrogen c. 90-fold and c. 30-fold. The increases in concentrations in the runoff water were very closely correlated with increases in humification of the peat. The correlations of the concentrations measured in extraction experiments (48 h) with peat type and degree of humification corresponded to those observed in the sprinkler experiments. The resulting figures for the surface runoff from a peat production area into the drainage ditches simulated by means of sprinkling and material concentrations in the runoff water were combined with statistics on the mean extent of daily rainfall (0 67 mm) during the frost-free period of the year (May October) over an observation period of 30 years to yield typical annual loading figures (kg/ha) for suspended solids (SS), chemical oxygen demand of organic matter (CODmn O2), total phosphorus (tot. P) and total nitrogen (tot. N) entering the ditches with respect to milled Carex (C) and Sphagnum (S) peats of humification grades H 2.5 8.5. In order to calculate the loading of drainage ditches from a milled peat production mire with the aid of these annual comparative values (in kg/ha), information is required on the properties of the intended production mire and its peat. Once data are available on the area of the mire, its peat depth, peat types and their degrees of humification, dry matter content, calorific value and corresponding energy content, it is possible to produce mutually comparable estimates for individual mires with respect to the annual loading of the drainage ditch system and the surrounding watercourse for the whole service life of the production area, the duration of this service life, determinations of energy content and the amount of loading per unit of energy generated (kg/MWh). In the 8 mires in the Köyhäjoki basin, Central Ostrobothnia, taken as an example, the loading of suspended solids (SS) in the drainage ditch networks calculated on the basis of the typical values obtained here and existing mire and peat data and expressed per unit of energy generated varied between the mires and horizons in the range 0.9 16.5 kg/MWh. One of the aims of this work was to develop means of making better use of existing mire and peat data and the results of corings and other field investigations. In this respect combination of the typical loading values (kg/ha) obtained here for S, SC, CS and C peats and the various degrees of humification (H 2.5 8.5) with the above mire and peat data by means of a computer program for the acquisition and handling of such data would enable all the information currently available and that deposited in the system in the future to be used for defining watercourse loading estimates for mires and comparing them with the corresponding estimates of energy content. The intention behind this work has been to respond to the challenge facing the energy generation industry to find larger peat production areas that exert less loading on the environment and to that facing the environmental authorities to improve the means available for estimating watercourse loading from peat production and its environmental impacts in advance. The results conform well to the initial hypothesis and to the goals laid down for the research and should enable watercourse loading from existing and planned peat production to be evaluated better in the future and the resulting impacts to be taken into account when planning land use and energy generation. The advance loading information available in this way would be of value in the selection of individual peat production areas, the planning of their exploitation, the introduction of water protection measures and the planning of loading inspections, in order to achieve controlled peat production that pays due attention to environmental considerations.

Cyanidin 3-glucoside improves diet-induced metabolic syndrome in rats

Increased consumption of dark-coloured fruits and vegetables may mitigate metabolic syndrome. This study has determined the changes in metabolic parameters, and in cardiovascular and liver structure and function, following chronic administration of either cyanidin 3-glucoside (CG) or Queen Garnet plum juice (QG) containing cyanidin glycosides to rats fed either a corn starch (C) or a high-carbohydrate, high-fat (H) diet. Eight to nine-week-old male Wistar rats were randomly divided into six groups for 16-week feeding with C, C with CG or QG, H or H with CG or QG. C or H were supplemented with CG or QG at a dose of ∼8 mg/kg/day cyanidin glycosides from week 8 to 16. H rats developed signs of metabolic syndrome including visceral adiposity, impaired glucose tolerance, hypertension, cardiovascular remodelling, increased collagen depots in left ventricle, non-alcoholic fatty liver disease, increased plasma liver enzymes and increased inflammatory cell infiltration in the heart and liver. Both CG and QG reversed these cardiovascular, liver and metabolic signs. However, no intact anthocyanins or common methylated/conjugated metabolites could be detected in the plasma samples and plasma hippuric acid concentrations were unchanged. Our results suggest CG is the most likely mediator of the responses to QG but that further investigation of the pharmacokinetics of oral CG in rats is required.

Stable expression of silencing-suppressor protein enhances the performance and longevity of an engineered metabolic pathway

Transgenic engineering of plants is important in both basic and applied research. However, the expression of a transgene can dwindle over time as the plant's small (s)RNA-guided silencing pathways shut it down. The silencing pathways have evolved as antiviral defence mechanisms, and viruses have co-evolved viral silencing-suppressor proteins (VSPs) to block them. Therefore, VSPs have been routinely used alongside desired transgene constructs to enhance their expression in transient assays. However, constitutive, stable expression of a VSP in a plant usually causes pronounced developmental abnormalities, as their actions interfere with endogenous microRNA-regulated processes, and has largely precluded the use of VSPs as an aid to stable transgene expression. In an attempt to avoid the deleterious effects but obtain the enhancing effect, a number of different VSPs were expressed exclusively in the seeds of Arabidopsis thaliana alongside a three-step transgenic pathway for the synthesis of arachidonic acid (AA), an ω-6 long chain polyunsaturated fatty acid. Results from independent transgenic events, maintained for four generations, showed that the VSP-AA-transformed plants were developmentally normal, apart from minor phenotypes at the cotyledon stage, and could produce 40% more AA than plants transformed with the AA transgene cassette alone. Intriguingly, a geminivirus VSP, V2, was constitutively expressed without causing developmental defects, as it acts on the siRNA amplification step that is not part of the miRNA pathway, and gave strong transgene enhancement. These results demonstrate that VSP expression can be used to protect and enhance stable transgene performance and has significant biotechnological application.

Computational Methods for Reconstruction and Analysis of Genome-Scale Metabolic Networks

Metabolism is the cellular subsystem responsible for generation of energy from nutrients and production of building blocks for larger macromolecules. Computational and statistical modeling of metabolism is vital to many disciplines including bioengineering, the study of diseases, drug target identification, and understanding the evolution of metabolism. In this thesis, we propose efficient computational methods for metabolic modeling. The techniques presented are targeted particularly at the analysis of large metabolic models encompassing the whole metabolism of one or several organisms. We concentrate on three major themes of metabolic modeling: metabolic pathway analysis, metabolic reconstruction and the study of evolution of metabolism. In the first part of this thesis, we study metabolic pathway analysis. We propose a novel modeling framework called gapless modeling to study biochemically viable metabolic networks and pathways. In addition, we investigate the utilization of atom-level information on metabolism to improve the quality of pathway analyses. We describe efficient algorithms for discovering both gapless and atom-level metabolic pathways, and conduct experiments with large-scale metabolic networks. The presented gapless approach offers a compromise in terms of complexity and feasibility between the previous graph-theoretic and stoichiometric approaches to metabolic modeling. Gapless pathway analysis shows that microbial metabolic networks are not as robust to random damage as suggested by previous studies. Furthermore the amino acid biosynthesis pathways of the fungal species Trichoderma reesei discovered from atom-level data are shown to closely correspond to those of Saccharomyces cerevisiae. In the second part, we propose computational methods for metabolic reconstruction in the gapless modeling framework. We study the task of reconstructing a metabolic network that does not suffer from connectivity problems. Such problems often limit the usability of reconstructed models, and typically require a significant amount of manual postprocessing. We formulate gapless metabolic reconstruction as an optimization problem and propose an efficient divide-and-conquer strategy to solve it with real-world instances. We also describe computational techniques for solving problems stemming from ambiguities in metabolite naming. These techniques have been implemented in a web-based sofware ReMatch intended for reconstruction of models for 13C metabolic flux analysis. In the third part, we extend our scope from single to multiple metabolic networks and propose an algorithm for inferring gapless metabolic networks of ancestral species from phylogenetic data. Experimenting with 16 fungal species, we show that the method is able to generate results that are easily interpretable and that provide hypotheses about the evolution of metabolism.

Algorithms for 13C metabolic flux analysis

The metabolism of an organism consists of a network of biochemical reactions that transform small molecules, or metabolites, into others in order to produce energy and building blocks for essential macromolecules. The goal of metabolic flux analysis is to uncover the rates, or the fluxes, of those biochemical reactions. In a steady state, the sum of the fluxes that produce an internal metabolite is equal to the sum of the fluxes that consume the same molecule. Thus the steady state imposes linear balance constraints to the fluxes. In general, the balance constraints imposed by the steady state are not sufficient to uncover all the fluxes of a metabolic network. The fluxes through cycles and alternative pathways between the same source and target metabolites remain unknown. More information about the fluxes can be obtained from isotopic labelling experiments, where a cell population is fed with labelled nutrients, such as glucose that contains 13C atoms. Labels are then transferred by biochemical reactions to other metabolites. The relative abundances of different labelling patterns in internal metabolites depend on the fluxes of pathways producing them. Thus, the relative abundances of different labelling patterns contain information about the fluxes that cannot be uncovered from the balance constraints derived from the steady state. The field of research that estimates the fluxes utilizing the measured constraints to the relative abundances of different labelling patterns induced by 13C labelled nutrients is called 13C metabolic flux analysis. There exist two approaches of 13C metabolic flux analysis. In the optimization approach, a non-linear optimization task, where candidate fluxes are iteratively generated until they fit to the measured abundances of different labelling patterns, is constructed. In the direct approach, linear balance constraints given by the steady state are augmented with linear constraints derived from the abundances of different labelling patterns of metabolites. Thus, mathematically involved non-linear optimization methods that can get stuck to the local optima can be avoided. On the other hand, the direct approach may require more measurement data than the optimization approach to obtain the same flux information. Furthermore, the optimization framework can easily be applied regardless of the labelling measurement technology and with all network topologies. In this thesis we present a formal computational framework for direct 13C metabolic flux analysis. The aim of our study is to construct as many linear constraints to the fluxes from the 13C labelling measurements using only computational methods that avoid non-linear techniques and are independent from the type of measurement data, the labelling of external nutrients and the topology of the metabolic network. The presented framework is the first representative of the direct approach for 13C metabolic flux analysis that is free from restricting assumptions made about these parameters.In our framework, measurement data is first propagated from the measured metabolites to other metabolites. The propagation is facilitated by the flow analysis of metabolite fragments in the network. Then new linear constraints to the fluxes are derived from the propagated data by applying the techniques of linear algebra.Based on the results of the fragment flow analysis, we also present an experiment planning method that selects sets of metabolites whose relative abundances of different labelling patterns are most useful for 13C metabolic flux analysis. Furthermore, we give computational tools to process raw 13C labelling data produced by tandem mass spectrometry to a form suitable for 13C metabolic flux analysis.