This thesis presents a strategy designed to determine how Thi1 and Thi5 may differentially regulate thiamine production and the vegetative and sexual life cycles in fission yeast. Fission yeast cells regulate the cell cycle and development in response to available nutrients. Thiamine, vitamin B1, is an essential vitamin and its active form, TDP (thiamine diphosphate), is an important co-factor for many metabolic enzymes. It also acts as an inhibitor of meiosis and zygote formation in fission yeast. The thiamine biosynthetic pathway in fission yeast is repressed by the presence of thiamine. The transcription factors, Thi1 and Thi5, are independently capable of positively regulating the nmt1 (no message in thiamine) promoter in fission yeast and both factors are required for wildtype nmt1 expression. Although Thi1 and Thi5 regulate the same promoter in thiamine biosynthesis, it has been previously found that Thi1 and Thi5 antagonistically regulate different aspects of meiosis. Thi1 and Thi5 are both Zn(II) 2Cys6 transcription factors. Since zinc finger transcription factors are known to homo- and hetero-dimerize, I hypothesize that the synergistic and antagonistic attributes of Thi1 and Thi5 may be due to this process. To examine their potential interaction in vegetative and meiotic cells using fluorescence resonance energy transfer (FRET) as well as co-immunoprecipitation, tagged versions of these proteins have been constructed. For FRET analysis Thi1 and Thi5 have been successfully tagged with CFP and YFP and are able to rescue the thiamine auxotrophy of strains lacking thi1 and thi5. To perform co-immunoprecipitation Thi1 and Thi5 have been tagged with His6. I was able to detect the tagged versions of Thi1 and Thi5 using commercially available antibodies against the fluorescent tags CFP and YFP and the His6 tag. The use of these constructs to address the interaction between these proteins is currently underway. / Thesis (Master, Biology) -- Queen's University, 2012-06-29 09:18:42.349

Thesis (M.Sc. (Pharmacology))--Mahidol University, 1983.

Thesis (M.Sc. (Tropical Medicine))--Mahidol University, 1979.

No description available.

Thiamine (vitamin B1) is involved in several basal metabolic processes. It is an essential compound for many organisms and in aquatic systems it is mainly produced by phytoplankton and prokaryotes and transferred to higher trophic levels through grazing and predation. The occurrence of thiamine deficiency in top predators has been reported from several aquatic systems. In the Baltic populations of the Atlantic salmon (Salmo salar) this has been observed since 1974 and recently thiamine deficiency has also been reported for Baltic sea birds. This thesis aims at investigating what processes that governs the flow of thiamine from the primary producers to top predators via zooplankton grazers and planktivoric fish. Paper I showed that abiotic stress factors such as salinity, temperature and light conditions can alter the thiamine content of phytoplankton. Paper II showed that abiotic factors indirectly can affect the stress resistance of zooplankton grazers by changing the nutritional quality of their food. In Paper III we found that the in situ thiamine content of zooplankton grazers was directly affected by that of the phytoplankton diet. In Paper IV we found a similar connection between the thiamine contents of Baltic salmon and herring, one of the major salmon prey species. In Paper V we looked at the thiamine content of the pelagic food web of the Baltic Sea as a whole and found a pattern of trophic dilution; the higher the trophic level of an organism (i.e. the further away from the source of thiamine in the food web), the lower was its thiamine content. In all, the results of this thesis suggests a bottom up effect on the thiamine status of the higher trophic levels of  the Baltic Sea and that external factors, both natural and man-made, have the capability to affect the thiamine status of the plankton communities and thereby the whole Baltic ecosystem. Thiamine and other micronutrients are not something generally considered in the environmental management of aquatic systems but the results of this thesis suggest that ecological disturbances indirectly can have negative effects on top predators via a disrupted supply of essential substances. / <p>At the time of the doctoral defense, the following papers were unpublished and had a status as follows: Paper 2: Manuscript. Paper 3: Manuscript. Paper 4: Manuscript. Paper 5: Manuscript.</p>

Thiamine-phosphate kinase (or ATP:thiamine-phosphate phosphotransferase, ThiL) and homoserine (ThrB) kinases are essential to metabolism in Mycobacterium tuberculosis (Mtb). ThiL and ThrB respectively phosphorylate thiamine monophosphate (TMP) to thiamine diphosphate (TDP), the active form of vitamin B1, and L-homoserine to Ophosphohomoserine, critical to aspartate biosynthesis. In this study, ThiL and ThrB from Mtb were characterised structurally and functionally by producing the proteins recombinantly in E. coli. Proteins were purified by affinity, anion exchange and size exclusion chromatographies and purity checked by SDS-PAGE. ThiL and ThrB enzyme activities were confirmed and reaction products verified by high pressure liquid chromatography (HPLC). The crystal structure of ThiL was solved by molecular replacement using X-ray diffraction data. Functionally active ThiL, 36 kDa, produced hexagonal crystals belonging to space group P6122 with one monomer per asymmetric unit. Structurally it is related to ThiL from other organisms with minor structural deviations. Enzymatically active ThrB, 33 kDa, was crystallised. However, crystals failed to diffract Xrays to a suitable resolution. ThiL and ThrB could act as possible anti-TB drug targets against Mtb. / Dissertation (MSc)--University of Pretoria, 2017. / Biochemistry / MSc / Unrestricted

Alfalfa has long been recognized as an outstanding hay plant in term. of both yield and feeding value. It is high in digestible protein, minerals, and vitamins. Because of its long life it can be economically produced. It is of exceptional value in maintaining soil fertility by helping to control erosion, by improving the physical condition of the soil, and by accumulating large amount of nitrogen.

The majority of water soluble vitamin and organic acid food additives are distributed in their crystalline forms. However, when they are combined with water and other food ingredients and then exposed to a variety of unit operations, there is potential to solidify these initially crystalline ingredients in the amorphous state. Amorphous solids are generally less chemically and physically stable than their crystalline counterparts. To ensure nutrient delivery to the consumer and fulfill labeling laws, deterioration of nutrients due to unintentional amorphization is undesirable. Additionally, the potential for recrystallization of an amorphous ingredient may alter texture and redistribute water. Hence, solid state form is a critical factor dictating the stability of food formulations. Building on earlier work from my M.S. degree that demonstrated thiamine chloride hydrochloride could solidify in the amorphous state in the presence of a variety of polymers (Arioglu-Tuncil et al., 2017), a major goal of this study was to develop a comprehensive understanding of the physical and chemical stability of amorphous forms of two thiamine salts, thiamine chloride hydrochloride (TClHCl) and thiamine mononitrate (TMN), in comparison to their crystalline counterparts and each other. The objectives for this part of the work were to investigate amorphization/recrystallization tendencies of TMN and TClHCl in solid dispersions, as well as chemical stability of thiamine in the solid dispersions to understand the impact of vitamin form, physical state (amorphous vs. crystalline), polymer type and features (Tg, hygroscopicity, and ability for intermolecular interactions), storage conditions, proportion of vitamin to polymer,and pre-lyophilized solution pHs on thiamine degradation and the physical stability of dispersions. Thiamine degraded more when in the amorphous form compared to in the crystalline state. Additionally, polymer type and vitamin proportion influenced thiamine degradation, where thiamine degraded more when it was present in lower concentrations (in dispersions that had higher Tgs), and it was chemically more stable when a polymer with greater intermolecular interactions with the vitamin was used. As storage RH increased, variably hygroscopicities of the polymers resulted in different thiamine degradation rates. The pre-lyophilization pHs of the solutions had a significant impact on thiamine stability in the solid dispersions. Similar to thiamine salts, citric acid is a commonly used food ingredient with a high crystallization tendency. Following similar experimental designs for documenting the recrystallization tendencies of citric acid in amorphous solid dispersions to those used in the thiamine studies, hydrogen bonding and/or ionic interactions between polymer and citric acid were found to be the main stabilizing factor for delaying recrystallization, more than polymer Tg and hygroscopicity. The findings of this dissertation provide a powerful prediction approach to physically and chemically stabilize the small compounds in the complex food matrices for the production of high quality food products and ensuring nutrient delivery to target populations.<br>

Depuis peu, des travaux ont montré que chez l’Homme, la transcétolase (TK, EC 2.2.1.1.) dont le cofacteur est la thiamine diphosphate (forme active de la vitamine B1), est une enzyme impliquée dans de nombreuses maladies telles que, le diabète, certains cancers, ou encore des maladies neurologiques, comme le syndrome de Wernicke-Korsakoff et la maladie d’Alzheimer. Pour des applications thérapeutiques, des inhibiteurs spécifiques de cette enzyme sont actuellement conçus et synthétisés dans les milieux académiques et industriels. Afin de déterminer l’activité de la TK (dans un but de diagnostic) d’une part, et de détecter des inhibiteurs potentiels de cette enzyme (dans un but thérapeutique) d’autre part, il est nécessaire de disposer de tests alliant rapidité, sensibilité et faible coût. Nous avons envisagé d’utiliser des biocapteurs ampérométriques qui combinent l’ensemble de ces avantages, et qui, de plus, n’ont jamais été mis en oeuvre avec la TK. Pour la détermination de l’activité des TK d’E. coli et humaine libres en solution, nous avons tout d’abord élaboré un premier biocapteur à galactose oxydase (GAOx, EC 1.1.3.9), dans lequel cette enzyme est immobilisée sur la laponite. Puis, dans le but de detecter des inhibiteurs de la TK, avec un système réutilisable, nous avons developpé un biocapteur à GAOx-TK d’E. coli, les deux enzymes étant co-immobilisées à la surface de l’électrode. Pour cela la TK a été immobilisée dans des Hydroxydes Doubles Lamellaires (HDL). Ce biocapteur bicouche et bi-enzymatique GAOx-TK, nous a permis d’évaluer l’effet d’inhibiteurs, tels que différents analogues du cofacteur et de substrats pris comme modèles. / Some recent studies have shown that human transketolase (TK, EC 2.2.1.1.), which thiamine diphosphate (active form of vitamin B1) is the cofactor, is involved in numerous disease such as diabete, some cancers and neurodegenerative diseases as Alzheimer’s disease and Wernicke-Korsakoff syndrome. For therapeutic purposes, TK inhibitors have been designed and synthesized in both academic and industrial fields. To determine TK activity (diagnostic) on the one hand, and to detect potential inhibitors of this enzyme (therapeutic) on the other hand, it is necessary to develop fast, sensitive and low cost assays. In this context, we designed some original amperometric biosensors that combine these advantages and were never studied with TK from now. We performed a first galactose oxidase (GAOx, EC 1.1.3.9) biosensor for E. coli and human TK activities detection. For that purpose, GAOx was immobilized on laponite matrix. Then, we designed a GAOx-TK biosensor by co-immobilization of GAOX and TK on the electrode surface that enabled the detection TK inhibitors with a reusable system. Thence, TK was immobilized in Layered Double Hydroxides (HDL). This bilayer and bi-enzymic biosensors, allowed us to determine the inhibitor potencies of several cofactors and substrates analogues as model compounds.

Entende-se por pólen apícola o resultado da aglutinação do pólen das flores, efetuado pelas abelhas operárias, mediante néctar e substâncias salivares, o qual é recolhido no ingresso da colmeia. A literatura descreve que esse alimento contém proteínas, carboidratos, lipídeos, vitaminas e minerais. De acordo com estudo prévio, amostras de pólen apícola in natura e desidratado, da cidade de Pariquera-Açu (São Paulo), apresentaram teores significativos de vitamina B1(tiamina) e B2 (riboflavina), além da presença dos vitâmeros da vitamina B3 (ácido nicotínico e nicotinamida) e B6 (piridoxal, piridoxol e piridoxamina) em sua composição o que foi associado à flora local explorada pelas abelhas. A região Sul do Brasil possui clima, relevo e vegetação diferenciados de outras regiões, necessitando-se assim da verificação do potencial vitamínico deste produto local. Destaca-se, ainda, o fato de que nesta região encontra-se um dos dois maiores produtores nacionais de pólen apícola (estado de Santa Catarina). O presente trabalho teve como objetivo principal quantificar os teores das vitaminas do complexo B: vitaminas B1 e B2, assim como os vitâmeros das vitaminas B3 e B6. Foram coletados 28 lotes de pólen apícola desidratado de diferentes localidades da região Sul durante o período de agosto de 2011 a dezembro de 2012 que posteriormente foram armazenados, a -18 °C até o momento das análises. As vitaminas do complexo B foram analisadas por cromatografia liquida de alta eficiência (CLAE) na matriz pólen apícola desidratado e os resultados foram expressos em base seca. Entre as amostras analisadas foram verificados teores de vitamina B1 variando entre 0,46 e 1,83 mg / 100 g de pólen apícola; vitamina B2 de 0,40 à 1,86 mg / 100 g e quanto à vitamina B6 apenas os vitâmeros piridoxal e piridoxamina puderam ser quantificados em todos os lotes analisados. O piridoxal teve variação entre as amostras de 0,42 à 6,70 mg / 100 g e a piridoxamina de 0,26 à 0,95 mg / 100g. Em relação à vitamina B3, o vitâmero ácido nicotínico apresentou-se nos diferentes lotes variando de 0,68 à 3,93 mg / 100 g e a nicotinamida de 0,27 à 5,54 mg / 100 g de produto. Tomando-se como porção sugerida para consumo diário 25 g de pólen apícola, verificou-se que num total de 28 amostras, 15 foram consideradas fontes e 2 como ricas em tiamina; 19 lotes foram fontes e 3 ricos em riboflavina, e; 2 lotes foram fontes e 26 ricos em piridoxina segundo à Ingestão Diária Recomendada (IDR) para adultos como disponibilizado na Resolução de Diretoria Colegiada (RDC) nº. 269, de 22 de setembro de 2005. / Bee pollen is understood to be the result of agglutination of pollen from flowers, made by worker bees, and nectar through salivary substances, which is collected at the hive entrance. The literature describes that this product contains proteins, carbohydrates, lipids, vitamins, minerals. Previous study with fresh and dehydrated bee pollen, from the city of Pariquera-Açu (São Paulo) showed significant levels of vitamin B1 (thiamine), B2 (riboflavin), presence of B3 (nicotinic acid and nicotinamide) and B6 (pyridoxal, pyridoxamine, piridoxol) vitamins vitamers in its composition which was associated with the local flora explored by bees. Southern Brazil has a differentiated climate, topography and vegetation from other regions, thus requiring verification of vitamin potential of this local product. Also stands out the fact that this region is one of the two largest national producers of bee pollen (Santa Catarina state). This study aimed to quantify the levels of B complex vitamins: vitamins B1, B2, as well as the vitamers of vitamins B3 and B6. Thus, it was collected 28 batches of dehydrated bee pollen from different locations in the South during the period from August 2011 to December 2012. Samples were obtained and subsequently stored at -18 ° C until the analysis time. B vitamins were analyzed by high performance liquid chromatography (HPLC) in bee pollen dehydrated matrix and results were expressed on a dry basis. Among the samples it levels of vitamin B1 varied from 0.46 to 1.83 mg / 100 g; vitamin B2 from 0.40 to 1.86 mg / 100 g; and for vitamin B6, only the pyridoxal and pyridoxamine vitamers could be quantified in all analyzed batches. The pyridoxal had variation between samples from 0.42 to 6,70 mg / 100 g and pyridoxamine from 0.26 to 0.95 mg / 100g. Taking 25 g of bee pollen as suggested for daily intake portion, it was found in a total of 28 samples that 15 were considered sources and 2 rich in thiamine; 19 lots were sources and 3 rich in riboflavin, and; 2 lots were sources and 26 rich in pyridoxine in relation to the Reference Daily Intake (RDI) for adults as provided in Resolução de Diretoria Colegiada (RDC) nº 269, de setembro de 2005.