The overarching goal of this dissertation is to refine methods employed for assessing neuromuscular changes and associated power/force outputs during various perturbations of fatigue, direct or perceived, induced by either exercise or nutritional interventions, with associated performance outcomes. To address this goal, we collected physiological and biomechanical data from subjects across a set of experiments designed to induce different levels of fatigue by the implementation of various exercise and nutritional interventions to cause various levels of fatigue in an ecologically valid manner. The data sets were collected during a single joint task and during cycling trials. During these experimental trials, we collected measures of kinetics (force and cycling power) as well as muscle activation (EMG) and physiological measures (heart rate, rating of perceived exertion, blood lactate, blood glucose, ventilation, oxygen uptake and carbon dioxide production) to investigate the overall performance, as well as potential mechanisms for improved performance related to the exercise and nutritional interventions. In order to substantially enhance the collection of cycling kinetics and kinematics, we have developed an innovative sensor that improved the measurement resolution (temporal and spatial) of a commercial research grade power meter. Using these improved measures alongside advanced muscle activity analysis, we could ameliorate an experimental framework that could be used to investigate changes in fatigue and coordination pattern associated with exercise and nutritional interventions. Investigation of the effects of a CHO mouth rinse vs. placebo on force and muscle activity during a very short (<3 min) neuromuscular demanding fatiguing trial demonstrated a consistent change in EMG median frequency related to increased fatigue in both experimental conditions, providing little evidence of change in neuromuscular strategy associated with CHO mouth rinse. Further investigation explored the effects of a CHO mouth rinse vs. placebo using fundamental physiological measures of neuromuscular activation and overall performance measures during an ecologically valid late endurance cycling time trial. Our results demonstrated that while there was no overall effect noticed for time to completion, there was a significant decrease in performance in the time to complete various components of the time trial during the placebo trial only. Muscle activity of the lower leg (MG and SOL) demonstrated a modification in frequency only evident during the placebo condition. Application of principal component analysis to power output and the EMG intensity profiles of the muscles of the lower leg during the pedal cycle revealed a more detailed understanding of the effect of CHO mouth rinse on performance during cycling. The average power output profile in WASH showed an earlier onset in the pedal cycle, greater duration and higher amplitude versus PLA during the TT. Additionally, only the PLA condition showed a significant increase in muscle activation throughout the time trial, which could be evidence of fatigue. This dissertation shows for the first time that CHO mouth rinse may have a substantial effect on the maintenance of power while mitigating the impact of neuromuscular fatigue, in late endurance performance, further strengthen our assertion that CHO may, in fact, minimize the changes in performance that are associated with fatigue during late endurance fatiguing events. / Graduate

The topic of my master thesis is monitoring of carbohydrate intake in sportsmen's diet. In the thesis, intake of carbohydrates as well as other macronutrients is monitored in order to evaluate, if athletes prefer other energy sources to carbohydrates and if their total energy intake is sufficient. If athletes prefer easily digestible carbohydrates or if they follow recommendation for healthy diet and increase their intake of whole foods, legumes, fruits and vegetables can be evaluated by monitoring fiber and sugar intake. All the monitored data are compared with non-athletes in order to see, in which field their diets vary. In the theoretical part, principles of sports nutrition are defined, basics of population recommendations are mentioned and methods of nutritional assessment and energy expenditure estimation are explained. In the practical part, five-day food-diaries of athletes and non-athletes are evaluated by NutriPro Expert software. Five-day physical activity diaries recorded during the same days are evaluated by using the Compendium of Physical Activities. All the studied values (energy intake, energy expenditure, carbohydrate, fat, protein, sugar, fiber and alcohol intake) were compared to recommended values for athletes and non- athletes and further compared among these two groups. The...

The conceptually simple process of linking carbohydrate units by glycosylation has proven to be one of the most difficult synthetic processes to control from a stereochemical perspective. In particular it is the stereocontrolled synthesis of 1,2-cis glycosyl linkages (e.g. α-glucosides, β-mannosides) which poses the most difficult challenge. The research presented in this thesis describes new ways in which stereocontrol in glycosylation reactions can be achieved. New methods of neighbouring group participation have been explored, utilising novel protecting groups at the 2-postion of a series of glycosyl donors. In particular the use of glycosyl donors bearing a (thiophen-2-yl)methyl protecting group at the 2-hydroxyl have shown exceptional α-selectivity especially when used in conjunction with a sterically hindered glycosyl acceptor. Work within this thesis also describes the first use of chiral Brønsted acid catalysts in the activation of glycosyl donors. It has been clearly demonstrated that not only can such catalysts be used in glycosylation reactions, but also that the chirality of the catalyst can dictate the stereochemical outcome of the reaction. The preliminary studies presented demonstrate that this methodology warrants further investigation.

This thesis investigates the ability of rainbow trout to modulate hepatic glucose production (Ra) and disposal (Rd). My goals were to determine: (1) if resting trout can modulate fluxes to cope with exogenous glucose; (2) how fluxes change during graded swimming; (3) how exogenous glucose affects swimming kinetics; and (4) if exogenous glucose affects cost of transport or performance. Results show that resting trout suppress Ra completely and stimulate Rd from 10.6 to 27.6 μmol kg-1 min-1. During swimming, fluxes increase from 15.6 to 21.9 μmol kg-1 min-1, but only at speeds >2.4 BL s-1. When given glucose, trout suppress Ra from 16.4 to 4.1 μmol kg-1 min-1 and stimulate Rd from 16.4 to 40.1 μmol kg-1 min-1. Glucose lowers metabolic rate but does not affect critical swimming speed. Therefore, this research shows that rainbow trout have a much better capacity for glucoregulation than generally suggested by current literature.

Digestible carbohydrate is commonly encountered by chinook salmon {Oncorhynchus tshawytscha) in practical culture diets, although little is known regarding its utilization. This study was undertaken to examine (1) the effects of a high carbohydrate diet and (2) glucose tolerance in chinook salmon of selected British Columbia strains. Yearling chinook salmon of three strains were fed to satiation either a high or a low carbohydrate diet for 63 days. The diets were isonitrogenous, and contained respectively 30 % gelatinized wheat starch or an equicaloric amount of herring oil. There was an overall reduction in growth of chinook fed the high-carbohydrate diet over the 63-day feeding period. Although specific growth rates declined initially in the high carbohydrate-fed groups, they were comparable to those of control groups in the final third of the trial, indicating an adaptation response. Chinook fed the high carbohydrate diet had increased carcass protein and ash, and decreased carcass fat levels relative to controls. Feed intake was generally lower in these groups, and differences in feeding response were observed between diets and strains. Although feed and energy efficiencies were reduced in chinook fed the high carbohydrate diet, protein utilization was comparable on the two diets, indicating a protein-sparing effect of the carbohydrate. Consumption of the high carbohydrate diet led to significant elevations in hepatosomatic indices (HSI) and liver glycogen (LG) concentrations. In Quesnel chinook, LG levels exceeding 10 % did not appear to have any detrimental effects on feeding, growth or health. LG concentrations and HSI fell to basal levels in all groups 21 days after feed withdrawal. Some strain differences were evident. For example, Big Qualicum chinook fed the high carbohydrate diet exhibited the lowest liver glycogen accumulation, highest rate of carcass fat deposition, and best energy efficiency ratios relative to control groups, suggesting a difference in carbohydrate metabolism in this strain. On the other hand, Quesnel chinook exhibited the highest relative growth on the high carbohydrate diet. Mortality, although unaffected by diet in the Quesnel and Robertson Creek chinook, appeared to be higher in high carbohydrate-fed Big Qualicum chinook. In the second part of the study, chinook salmon subjected to an oral glucose tolerance test displayed pronounced and persistent hyperglycaemia, indicative of poor glucose tolerance. Strain differences were evident in the magnitude of response. Acclimation to a high carbohydrate diet prior to testing resulted in a significantly reduced elevation of blood glucose, indicating an adaptation response. While plasma glucose concentrations approached 500 mg/dl in some trials, plasma insulin concentrations exhibited a two-fold rise, with indistinct peaks. Plasma glucose and plasma insulin concentrations were poorly correlated, indicating that glucose is a poor insulin secretagogue in chinook salmon. / Land and Food Systems, Faculty of / Graduate

The cryopreservation of cells and tissues requires the addition of a cryoprotectant in order to prevent cellular damage caused by ice. Unfortunately, common cryoprotectants such as DMSO and glycerol exhibit significant toxicity which makes their use unfeasible for many clinical procedures. Our laboratory is interested in the development of alternative, non-toxic cryoprotectants which possess ice recrystallization inhibition (IRI) activity. Potent IRI activity has recently been discovered in certain small molecules, but the structural features required for this process are unclear. Herein we report the development of a library of O-aryl glycosides in order to probe the importance of electron density and hydrophobic moieties for IRI activity. It was found that the degree of electron density at the anomeric oxygen does not correlate with IRI ability in para-substituted aryl glycosides, nor does changing the position of the aryl substituent impart a predictable effect on activity. However, the addition of hydrophobic alkyl or acyl chains was beneficial for IRI activity; generally, increasing chain length was found to correlate with increasing activity. In some instances, an optimal alkyl chain length was identified, after which continued lengthening results in a loss of potency. We conclude from this study that a certain extent of hydrophobic character is beneficial for the IRI activity of aryl glycosides, and that a balance between hydrophobicity and hydrophilicity is required for optimum IRI ability. It is hoped that these findings will aid future efforts towards the rational design of novel cryoprotectants.

La société Adisseo produit un cocktail d’enzymes hydrolytiques appelé Rovabio® Excel sécrété par un champignon filamenteux Talaromyces versatilis. Ce cocktail est utilisé comme additif alimentaire pour augmenter la digestibilité de complexes polysaccharidiques en nutrition animale et ainsi augmenter la valeur nutritionnelle des matières premières agricoles. Une récente étude protéomique de ce champignon (Guais et al., 2008) a révélé la présence d’un grand nombre d’arabinofuranosidases (ABFs) appartenant à différentes familles de glycosides hydrolases : 5 ABFs de la famille GH54, 3 ABFs de la famille GH62 et enfin une de la famille GH51. Un des objectifs de mes travaux de thèse a été le clonage, l’expression hétérologue (hôte lévurien Pichia pastoris) des 9 gènes codant pour ces 9 enzymes et la caractérisation complète de la famille GH62. La caractérisation des capacités d’hydrolyse des ABFs 54 et 62 a également été étudiée grâce à une technique d’empreintes d’hydrolyse enzymatique sur arabinoxylane de blé. Enfin, la dernière partie de mes travaux consistait à confectionner des mélanges d’enzymes hydrolytiques des différentes familles d’ABFs associés à des xylanases et de suivre l’efficacité de la dégradation de l’arabinoxylane insoluble grâce à l’utilisation d’un réacteur torique permettant l’acquisition d’images et l’analyse en ligne de la dégradation. Ces travaux sur le réacteur ont permis de mettre en évidence une synergie entre Abfs et Xylanases. / Adisseo produce and commercialize a hydrolytic enzymatic cocktail termed Rovabio and secreted by a filamentous fungus Talaromyces versatilis. This cocktail is used as feed additive for increased digestibility of complex polysaccharides in animal nutrition. A recent genomic study of this fungus revealed the presence of 5 arabinofuranosidases (Abfs) to family GH 54, 3 of GH 62 and 1 of GH51. The first aim of my thesis works was about cloning, heterologous overexpression (in pichia pastoris yeast) of this 9 genes encoding for this 9 enzymes and characterization of the family GH 62. Mode of action of ABFs 54 and 62s has been characterized by enzymatic fingerprinting analysis on wheat arabinoxylan. Then, last part was to design enzymatic cocktail with differents families of ABFs and Xylanases and test their impact on insoluble arabinoxylan hydrolysis with toric reactor. These works on reactor have bringing to light a synergy between ABFs and Xylanases

Humic substances, such as the closely related humic and fulvic acids are ubiquitous, naturally occurring organic macromolecules of complex but undefined structure. These compounds are known complexing agents due to their supramolecular like structures and are capable of binding a wide variety of compounds. Numerous studies have confirmed that humic and fulvic acids exhibit diverse medicinal and therapeutic properties. For this reason, alternative or “natural” medicinal preparations rich in these substances are being self-administered, often concomitantly with conventional drugs. The possibility exists that these humic substances, found in the alternative medicinal products, may result in drug-drug interactions and bind to simultaneously ingested drugs. Complex formation may affect absorption and alter overall bioavailability. Changes in these parameters may lead to reduced therapeutic effect or toxic side effects of prescribed drugs in patients. Similarly, these humic substances may bind to and alter the uptake of ingested nutrients, such as vitamins and minerals, obtained from food sources as well as dietary supplements. Changes in absorption may result in a loss of proper physiological functioning in the body or in unwanted effects of overdose. This study investigated the effect of fulvic and humic acids on the absorption of commonly administered classes of drugs, vitamins and minerals using the everted mouse gut model that was successfully used to assess the membrane transport of the test compounds. This model made use of everted segments of excised intestinal tissue placed in Krebs Ringer Buffer (pH7.4), where physiological functioning of the tissue is maintained for up to two hours after excision. The amount of test compound which crossed through the intestinal membrane without and in the presence of each humic substance was quantified using LC-MS/MS methods developed for each of the drugs and vitamins, and ICP-MS, in the case of the minerals. The amount of test compound absorbed alone was compared to the amount absorbed when in the presence of each humic substance. Changes in the uptake, for each test compound was noted, the extent of the absorption increase or decrease was compound specific. The changes in absorption observed could be attributed to changes in compound solubility and mechanism of transport across the intestinal membrane once in complex. Drugs and vitamins were seen to be more prone to decreases in absorption in the presence of the humic substances, whereas the majority of the minerals showed significantly increased absorption. Binding of the minerals to the humic substances through chelation, and not complex formation, could have a greater effect on compound solubility. Health care professionals, as well as individuals ingesting these and other substances concurrently, should be aware of the potential effects on absorption that may occur due to drug-drug interactions in order to avoid a loss of therapeutic/physiological activity or negative toxic symptoms. / Dissertation (MSc)--University of Pretoria, 2015. / tm2015 / Pharmacology / MSc / Unrestricted

Orientador: Paula Teixeira Fernandes / Dissertação (mestrado) - Universidade Estadual de Campinas, Faculdade de Educação Física / Made available in DSpace on 2018-08-24T10:11:07Z (GMT). No. of bitstreams: 1 Castanho_GabrielaKaiserFullin_M.pdf: 2680275 bytes, checksum: d8146be4deb23a380916dab5e59d52c2 (MD5) Previous issue date: 2014 / Resumo: O consumo de carboidrato (CHO) está relacionado com melhor desempenho esportivo. Por vias diretas ou indiretas o substrato mantém a glicemia, poupa glicogênio, repõe glicogênio muscular e, ainda, exerce papel importante no cérebro como fonte de energia, podendo melhorar a função neural durante exercício físico. Com o consumo de CHO é possível que haja diferenciação na ativação de áreas cerebrais relacionadas ao desempenho no exercício. Com isso, esse estudo objetivou avaliar o efeito do consumo de CHO na ativação cerebral durante exercício físico realizado durante aquisição de ressonância magnética funcional (RMf). Foram voluntários 10 ciclistas do sexo masculino que realizaram um protocolo de pedalada em cicloergômetro acoplado à RMf, consumindo de 50g de carboidrato ou solução placebo no intervalo de duas séries de exercício. Os resultados mostraram que o CHO apresentou um importante papel nas áreas cerebrais durante o exercício, ativando áreas relacionadas à tomada de decisões (insula) e motivação (sistema límbico) e desativando principalmente áreas dos lobos frontal e parietal. Com o consumo de placebo também houve ativação de áreas importantes no psicológico do individuo (cíngulo posterior). Independente da substância consumida, a ativação após o consumo de bebida mostrou importante relação com áreas relacionadas à continuidade do exercício como giro do cíngulo quando comparado ao controle. Durante todo o estudo as áreas responsáveis pela iniciação e manutenção dos movimentos, localizadas principalmente no lobo frontal e cerebelo, apresentaram-se ativas. Os resultados entre o consumo do placebo e o momento controle mostraram que a substância placebo não trouxe diferença de ativação neural, sendo que o inverso ocorreu na comparação entre CHO e controle. O CHO ativou áreas de extrema importância na continuidade do exercício como o giro do cíngulo e cíngulo anterior, mostrando que a suplementação pode ter influências no cérebro durante o exercício que melhore o rendimento. Portanto, o CHO mostrou efetividade como suplemento esportivo também na atividade cerebral, mostrando importância para a prática de exercício e a melhora do rendimento. Assim conseguimos aprofundar o conhecimento da atuação de nutrientes no cérebro durante o exercício / Abstract: The consumption of carbohydrate (CH) is related to better sports performance. By direct or indirect way the substrate maintains blood glucose, glycogen saves, restores muscle glycogen, and also plays an important role in the brain as an energy source, thus improving neural function during exercise. With the consumption of CH is possible that there is differentiation in the activation of brain areas related to exercise performance. The study aimed to evaluate the effect of CH consumption in brain activation during exercise performed during acquisition of functional magnetic resonance imaging (fMRI). Volunteers were six male cyclists who performed a protocol cycleergometer coupled to fMRI, consuming 50g of carbohydrate or placebo solution in the range of two series of exercise. The results showed that the CHO had an important role in brain areas during exercise, activating related decision making (insula) and motivation (limbic system) areas and mostly disabling areas of the frontal and parietal lobes. With the use of placebo was also significant activation in areas of individual psychological (posterior cingulate). Independent of substance, activation after beverage consumption showed a significant relationship with the continuity of exercise as the cingulate gyrus area, when compared to the control. Throughout the study the areas responsible for the initiation and maintenance of motion, located mainly in the cerebellum and frontal lobe had become active. The results between the placebo and the control group showed that the difference did not had neural activation, and the opposite occurred when comparing CHO and control. The CHO activated areas of extreme importance in the continuity of the company as the cingulate gyrus and anterior cingulate cortex, showing that supplementation may have influences in the brain during exercise to improve performance. Therefore, the CHO has shown effectiveness as a sports supplement also in brain activity important for exercise training results and greater efficiency. Thus we could deepen the understanding of the action of nutrients in the brain during exercise / Mestrado / Atividade Fisica, Adaptação e Saude / Mestra em Educação Física

&#946;-1,3-glucanases são enzimas encontradas em muitos organismos, como fungos, bactéria e plantas. Suas funções incluem remodelamento de parede celular, defesa e digestão. É um alvo interessante para o controle populacional de insetos-praga, porque é ausente em vertebrados. Em insetos, é encontrada no intestino de muitas ordens diferentes, e hidrolizam &#946;-1,3 ou &#946;-1,3(4)-glucanas ingeridas, mas pouco se sabe sobre as propriedades e a função dessas enzimas. Nós estudamos três espécies de insetos, de três ordens diferentes. Spodoptera frugiperda (Lepidoptera), Tenebrio molitor (Coleoptera) e Abracris flavolineata (Orthoptera) são insetos herbívoros e pestes de plantações, mas suas beta-1,3-glucanases diferem significativamente. A &#946;- 1,3-glucanase de S. frugiperda (SLAM) foi purificada do intestino médio da larva. Ela apresenta uma massa molecular de 37,5 kDa, um pH ótimo alcalino de 9,0, é ativa contra &#946;-1,3-glucana (laminarina), mas é incapaz de hidrolizar &#946;-1,3-1,6-glucana de levedura ou outros polisacarídeos. SLAM é não-processiva (0,4), e não é inibida por altas concentrações de substrato. Diferente de outras &#946;-1,3-glucanases digestivas de insetos, SLAM é incapaz de lisar células de Saccharomyces cerevisae. O cDNA correspondente à SLAM foi clonado e sequenciado, demonstrando que a proteína pertence à família 16 das Glicosídeo-Hidrolases. A modelagem tridimensional de SLAM, feito com base em homologia de sequência, sugere que o resíduo E228 possa afetar a ionização dos resíduos catalíticos, causando o deslocamento do pH ótimo da enzima. Anti-corpos específicos para SLAM foram produzidos, e estes reagem com uma única proteína oriunda do intestino médio da larva, responsável pela atividade &#946;-1,3- glucanásica majoritária. A imunocitolocalização de SLAM demonstra que a enzima é encontrada em vesículas secretórias e no glicocálix das células colunares, e portanto não é originária de simbiontes. Nós clonamos e sequenciamos o cDNA correspondente à &#946;-1,3-glucanase majoritária presente no intestino médio da larva de T. molitor (TLAM). Ela pertence à família 16 das Glicosídeo-Hidrolases e está relacionada com proteínas ligantes de &#946; -glucanas, da mesma forma que a enzima de S. frugiperda. A modelagem tridimensional por homologia de sequência permitiu identificar alguns resíduos de amino-ácidos (E174, E179, H204, Y304, R127 e R2181) no sítio ativo da enzima, que podem ser importantes para a atividade de TLAM. A &#946;-1,3-glucanase digestiva do gafanhoto Abracris flavolineata (Orthoptera) é diferente das enzimas já estudadas em insetos. Ela apresenta uma estratégia catalítica processiva, liberando glicose como maior parte dos produtos, e é inibida por altas concentrações de substrato. Para estudar as bases estruturais desse mecanismo, nós procuramos obter a sequência de cDNA correspondente à enzima já caracterizada. O alinhamento múltiplo das &#946;-1,3- glucanases de insetos e proteínas ligantes de beta-glucanas indicou que uma duplicação gênica da enzima do ancestral comum de moluscos e artrópodes. Uma cópia originou as beta-1,3-glucanase de insetos, perdendo uma região N-Terminal com cerca de 100 pares de bases, enquanto a outra cópia originou as proteínas ligantes de beta-glucana, perdendo os resíduos catalíticos. / &#946;-1,3-glucanases are widespread enzymes, found in all major groups of invertebrates, fungi, bacteria and plants. Since this enzyme is absent in vertebrates, it constitutes an interesting target for control of insect pests population. Its functions range from cell wall remodeling, defense and digestion. In insects, it is found in the gut of many different orders, hydrolyzing &#946;-1,3 or &#946;-1,3(4)-glucanas, but little is known about the properties and function of these enzymes. We studied three insect species each pertaining to a different order. Spodoptera frugiperda (Lepidoptera), Tenebrio molitor (Coleoptera) and Abracris flavolineata are herbivores and crops pests, but their &#946;-1,3- glucanases differ significantly. S. frugiperda &#946; -1,3-glucanase (SLAM) was purified from the larval midgut. It has a molecular mass of 37.5 kDa, an alkaline optimum pH of 9.0, is active against &#946;-1,3-glucan (laminarin), but cannot hydrolyze yeast &#946;-1,3-1,6-glucan or other polysaccharides. The enzyme is an endoglucanase with low processivity (0.4), and is not inhibited by high concentrations of substrate. In contrast to other digestive &#946;-1,3- glucanases from insects, SLAM is unable to lyse Saccharomyces cerevisae cells. The cDNA encoding SLAM was cloned and sequenced, showing that the protein belongs to glycosyl hydrolase family 16 as other insect glucanases and glucan-binding proteins. SLAM homology modeling suggests that E228 may affect the ionization of the catalytic residues, thus displacing the enzyme pH optimum. SLAM antiserum reacts with a single protein in the insect midgut. Immunocytolocalization reveals the presence of the enzyme in secretory vesicles and glycocalyx from columnar cells. We cloned and sequenced the cDNA of T. molitor &#946;-1,3-glucanase. It belongs to glycoside hydrolase family 16, and is related to other insect glucanases and glucan-binding proteins. Sequence analysis and homology modeling allowed the identification of some residues (E174, E179, H204, Y304, R127 and R181) at the active site of the enzyme, which may be important for TLAM activity. The grasshopper A. flavolineata has a &#946;-1,3-glucanase with a processive catalytic strategy. To study the structural basis of this property, we aimed to obtain its encoding sequence to better understand this catalytic mechanism. Multiple sequence alignment of insects &#946;-1,3-glucanases and &#946;-glucan-binding protein indicates that the &#946;- 1,3-glucanase gene duplicated in the ancestor of mollusks and arthropods. One copy originated the insect &#946;-1,3-glucanases after losing a 100 bp N-terminal portion and the arthropode &#946;-glucan-binding proteins by the loss of the catalytic residues.