In this work, we developed a synthetic route for obtaining β-aryl-chalcogenoazide and β-aryl-chalcogenotriazole (S, Se and Te) derived from L-amino acids (L-phenylalanine, L-valine and L-leucine). We obtained a compounds library due to the structural diversity of amino acids, organochalcogen compounds and terminal alkynes used in this synthesis. For the synthesis of chalcogenides, L-amino acids have been submitted to reduction, protection and mesylation processes for obtain protected aminomesylate. With this requerid mesylate in hand, we turned our attention to the introduction of the organochalcogenide moiety in the mesylate through the nucleophilic substitution reaction of the OMs leaving group, obtaining compound. Subsequently, the β-chiral amines were prepared after removal of the Boc protecting group with trifluoracetic acid. Further, with the chiral β-chalcogenoamines in hand, we moved our attention to the synthesis of β-aryl-chalcogenoazide via diazotransfer reaction with triflyl azide (TfN3) as a diazo donor group. This procedure has several advantages such as high yield, complete retention of configuration, mild reaction conditions, and good compatibility with most functional groups. Firstly we carry out the synthesis of TfN3 with sodium azide, triflic anhydride and acetonitrile as solvent. After, it was added to the chiral β-chalcogen amines solution for subsequent diazotransfer reaction getting β-chalcogenazides. In view of the possibility of the synthesis of triazoles due to the presence of azido group, we decided to synthesize 1,2,3-triazoles. For this synthesis we used copper (II) as a catalyst and the product is only the regioisomer 1,4-substituted 1,2,3-triazole. The β-aryl-chalcogenoazide and β-aryl-chalcogenotriazole were submited to analysis of antioxidant properties by in vitro tests. The cyclic voltammetry technique was used to study the redox potential of the synthesized compounds. / No presente trabalho, desenvolveu-se uma rota sintética para a obtenção de calcogenilazidas e calcogeniltriazóis (S, Se e Te) derivados de L-aminoácidos (L-fenilalanina, L-valina e L-leucina). Uma variedade de compostos pode ser obtida devido à diversidade estrutural dos aminoácidos, organocalcogênios e alquinos terminais utilizados. Para a síntese das calcogenilazidas, os L-aminoácidos foram submetidos aos processos de redução, proteção e mesilação para a obtenção dos β-aminomesilatos protegidos. De posse destes aminomesilatos realizou-se a síntese dos calcogenetos utilizando Reação de Substituição Nucleofílica no mesilato, OMs. Após foi realizada a desproteção do calcogenetos obtendo-se as β-calcogenilaminas quirais para posterior síntese das β-calcogenilazidas. A reação para a síntese das calcogenilazidas está baseada na diazotransferência. Neste tipo de metodologia a quiralidade da molécula é preservada, não havendo racemização e nem inversão de configuração do carbono assimétrico, além da obtenção de rendimentos elevados e da boa compatibilidade química com outros grupos funcionais. Dessa forma, primeiramente realizou-se a síntese do trifluorometanossulfônico azida (TfN3), chave desta etapa reacional. Após, foi procedida a síntese das β-calcogenilazidas a partir das β-calcogenilaminas. Com as calcogenilazidas em mãos e tendo em vista a possibilidade da síntese de triazóis, devido ao grupo azida ser um excelente bloco construtor de moléculas contendo heterociclos, realizou-se a síntese de calcogeniltriazóis. Para esta síntese foi utilizado uma metodologia com cobre (II) como catalisador obtendo-se apenas o regioisômero 1,2,3-triazol 1,4 substituído. Os respectivos compostos, β-calcogenilazidas e β-calcogeniltriazóis, foram avaliados com relação a sua atividade antioxidante in vitro e forneceram resultados bastante promissores com relação a esta atividade. Além disso, a técnica de Voltametria Cíclica também foi utilizada para avaliar o potencial redox desses compostos.

Il est admis que le Transforming Growth Factor-bêta (TGF-ß) est athéroprotecteur. Son apport bénéfique dans l'athérosclérose semble être contrarié par l'Interleukine-1-bêta (IL-1ß) qui est l'archétype des cytokines pro-inflammatoires. Notre but est, d'une part, de comprendre la régulation transcriptionnelle du TGF-ß dans les cellules musculaires lisses vasculaires (CMLv) humaines, dans l'athérome carotidien humain à partir des données du transcriptome obtenues sur 32 patients à la fois dans les plaques athéromateuses (ATH) et dans le tissu avoisinant macroscopiquement sain (TMS) et, d'autre part, d'étudier l'antagonisme des effets du TGF-ß et de l'IL-1ß sur des CMLv en culture provenant du TMS. Nous avons abordé nos problématiques par deux approches : (1) une analyse bioinformatique de données transcriptomiques issues de biopuces. (2) une étude in vitro sur des CMLv de la carotide humaine. in vitro. Grace à l'analyse in silico nous avons montré que l'implication de KLF6 dans la transcription du TGFB1 était assez spécifique des cellules musculaires lises vasculaires (CMLv). Nos résultats permettent également de proposer de nouveaux FT potentiels, spécifiques des CMLv (SLC2A4RG, GABP et SALL2), qui pourraient favoriser le rôle athéroprotecteur de TGFB1 dans les CMLv de la carotide. Enfin, il semble qu'il existe un équilibre entre les FT activateurs ou inhibiteurs de l'expression de TGFB1 qui permet d'en moduler finement la transcription. Notre approche in vitro, à montre que l'IL-1ß induisait un phénotype inflammatoire associé à une activité élastolytique consécutive, pour l'essentiel, à l'augmentation de l'expression de la cathépsine S (CTSS). La neutralisation du TGF-ß endogène dé-réprime l'expression de la CTSS et exerce un effet additif à celui de l'IL-1ß sur l'expression de l'enzyme. En conclusion, l'expression du TGFB1 dans les CMLv étant soumise à un contrôle transcriptionnel spécifique du type cellulaire, il est envisageable de développer des approches pharmacologiques qui permettent de maintenir l'expression du TGFB1 dans la paroi artérielle au niveau requis pour qu'il y exerce ses effets athéroprotecteurs / It is accepted that the TGF-β is atheroprotective. Its beneficial contribution atherosclerosis seems to be opposed by IL -1β that is the archetype of the pro-inflammatory cytokines. Our goal is, first, to understand the transcriptional regulation of TGF-β in human vSMCs in carotid atherosclerosis from the transcriptomic data obtained from 32 patients in both atherosclerotic plaques (ATM) and in the adjacent macroscopically intact tissue (MIT) and, secondly, to study the antagonism of the effects of TGF-β and IL-1β on vSMCs cultured from MIT samples. We followed two approaches: (1) a bioinformatic analysis of transcriptomic data from the microarrays; (2) an in vitro study of the human vSMCs. In silico, we have shown that the involvement of KLF6 in the transcription of TGFB1 was specific to the vSMCs. Our results also identify potential new transcription factors (SLC2A4RG, GABP and SALL2) that are vSMC-specific and could promote the atheroprotective role of TGFB1 in carotid vSMCs. The balance between the FT activating or inhibiting the expression of TGFB1 allows the fine tuning of its transcription. Our in vitro approach showed that IL-1β induces in the vSMCs an inflammatory phenotype associated with an elastolytic activity resulting mainly from the increase in the expression of cathepsin S (CTSS). Neutralization of endogenous TGF-β in the vSMCs de-represses the expression of the CTSS and exerts an additive effect to that of IL-1β on the expression of the enzyme. In conclusion, the expression of TGFB1 in the vSMCs is submitted to a cell-specific transcriptional control. It is possible to develop pharmacological approaches that maintain the expression the expression of TGFB1 in the arterial wall at the level required allowing TGF-β1 to exert its atheroprotective effects

BACKGROUND AND AIMS OF THE THESIS: Group A β-haemolytic Streptococcus (GAS) also known as Streptococcus pyogenes, is responsible for a wide range of invasive and non-invasive GAS diseases. Prevalence and incidence data on GAS from developing countries are largely lacking when compared with industrialised nations. This thesis sought to (1) establish the South African arm of the AFROStrep biorepository and clinical database for patients with invasive and non-invasive GAS infection, (2) identify and summarize all published studies of laboratory-confirmed GAS infection in Africa, (3) describe, from national laboratory data, the incidence of invasive and non-invasive GAS in South Africa and, (4) conduct a prospective, surveillance study in order to determine the molecular epidemiology of GAS in Cape Town, South Africa over a 12-month period. METHODS: A systematic review was conducted on population-based studies reporting on the prevalence of laboratory-confirmed GAS infection among patients living in Africa (Study 1). A retrospective study of the incidence of GAS infection was conducted on data obtained from the National Health Laboratory Service between 2003 – 2015 (Study 2). The AFROStrep registry and biorepository (based in Cape Town) was established and through passive surveillance, laboratory confirmed invasive and non-invasive GAS cases were collected over a 12-month period. The molecular analysis of invasive and non-invasive infection was determined using emm type sequencing to provide insight into vaccine development (Study 3). RESULTS AND DISCUSSION: The pooled prevalence of GAS pharyngitis in Africa was determined to be 21% (95% CI, 17% to 26%). The incidence rates of laboratory-confirmed non-invasive GAS infection in the South African public sector appears to have declined over the last 13 years. Given the possibility that the lower incidence of invasive and non-invasive GAS infection found in our study is due to infrequent submission of specimens for microbiological culture by health practitioners, our findings may be an underestimate of the true burden of disease in South Africa. In our prospective surveillance study, 46 different emm types were identified. The most prevalent emm types were M76 (16% of isolates), M81 (10%), M80 (6%), M43 (6%), and M183 (6%) and were almost evenly distributed between invasive and non-invasive GAS isolates. When compared against the putative 30-valent vaccine under development, four of our most prevalent emm types are not included; vaccine coverage (i.e. vaccine type and non-vaccine type-killing) for non-invasive and invasive GAS infection in our setting was 60% and 59% respectively, notably lower than coverage in developed countries. CONCLUSION: This work provides evidence for a significantly high prevalence of GAS pharyngitis in Africa. While GAS surveillance in South Africa indicates a declining incidence of GAS disease in parts of the country over the last thirteen years, the findings may be an underestimate of the true burden of disease, demonstrating the need for accurate and comprehensive surveillance of GAS in South Africa. Finally, this research showed a low potential vaccine coverage in our setting and thus, emphasises the need for a reworking of the potential vaccine formulation to improve coverage in areas where the burden of disease is high.

<p> The products of the reaction between oxomethoxodichlorobistriphenylphosphinerhenium(V) and acetylacetone are determined, characterized and correlated in terms of a reaction scheme. A series of rhenium(III) compounds, characteristic of reactions analogous to the one mentioned above, with other β-ketoenols is prepared and characterized. The syntheses and properties of tris-1,1,1-5,5,5-hexafluoropentane-2,4-dionatorhenium(III) and trispentane-2,4-dionatorhenium(III) are described. A new nitrido-rhenium complex, nitridochloro-1,1,1-5,5,5-hexafluoropentane-2,4-dionatobistriphenylphosphinerhenium(V) complex is prepared and characterized. A synthetic route to pyridine-acetylacetonate complexes of rhenium is described. </p> / Thesis / Doctor of Philosophy (PhD)

β-glucosidases catalyze the hydrolysis of aryl and alkyl β-D-glucosides as well as glucosides with a carbohydrate moiety. The maize β-glucosidase isozymes Glu1and Glu2 hydrolyze a broad spectrum of substrates in addition to its natural substrate DIMBOAGlc, while the sorghum β-glucosidase Dhr1 (dhurrinase-1) hydrolyzes exclusively its natural substrate dhurrin. For the expression of mature β-glucosidase isozymes Glu1 and Glu2 of maize and Dhr1 of sorghum, complementary DNAs were amplified by PCR and cloned into the expression vector pET-21a. Recombinant Glu1, Glu2 and Dhr1 enzymes were found to display activity towards the physiological substrates DIMBOAGlc and dhurrin, respectively, at levels similar to their native counterparts. It has been a subject of the subsequent studies by our lab and others to investigate what determines the aglycone specificity in β-glucosidases, and how β-glucosidases catalyze the hydrolysis of β-glycosidic bond between sugar and aglycone moieties. Molecular modeling techniques allowed to predict the substrate binding sites in Glu1 and Dhr1. Based on structural analysis of Glu1 and Dhr1, chimeric β-glucosidases (Glu1/Dhr1) were constructed by shuffling the C-terminal amino acids of Glu1 with the homologous region of Dhr1 to study the mechanism of substrate specificity. The resulting chimeric enzymes were characterized with respect to substrate specificity as well as kinetic, immunological, and electrophoretic properties. Shuffling a small portion of the C-terminal region altered the substrate specificity and improved by 2-4 fold the catalytic efficiency on other substrates in the chimeric β-glucosidases. These experiments showed that one or more of the 10 amino acid substitutions in the 30 amino acid long Dhr1 subdomain, 462SSGYTERFGIVYVDRENGCERTMKRSARWL491, plays a key role in dhurrin recognition and hydrolysis. To further investigate dhurrin recognition within this peptide region, two chimeric enzymes containing ⁴⁶²SSGYTERF⁴⁶⁹ and ⁴⁶⁶FAGFTERY⁴⁷³ Dhr1 peptides, respectively, were generated. The kinetic parameters indicated that Dhr1 peptide, ⁴⁶²SSGYTERF⁴⁶⁹, alone is sufficient to convert Glu1 to Dhr1 substrate specificity when it replaces the homologous peptide, ⁴⁶⁶FAGFTERY⁴⁷³, of Glu1. Maize β-glucosidases share high sequence similarities with Family 1 O-glucosidases. Therefore, these proteins are classified as retaining glycosyl hydrolases whose active site contains two glutamic acids (E) as the key catalytic residues, one as a general acid/base catalyst (E191) and the other as a nucleophile (E406). To confirm the identity and function of the acid/base catalyst E191, we have changed this residue to isosteric glutamine (Q) and aspartic acid (D) in both Glu1 and Glu2 isozymes by site-directed mutagenesis. The resulting mutant proteins were purified and their kinetic parameters (K_m, k_cat and k_i) were determined. The replacement of the acid/base catalyst E191 in the active site of maize β-glucosidase by Q and D resulted in inactivation of the enzyme. The kinetic analysis of the E191Q mutants showed that catalytic activity was reduced 200- and 110-fold towards ortho- and para-nitrophenyl- β-D-glucosides, respectively, when compared with the wild type enzyme. The E191D mutants showed no detectable activity towards any of the substrates tested. The back mutation of the E191Q mutants of the Glu1 and Glu2 isozymes to wild type restored full catalytic activity in both cases. These data indicate that E191 in maize β-glucosidases functions as an acid/base catalyst, and its function in catalysis cannot be performed by an isosteric residue such as glutamine or by a carboxyl group on a shorter side chain such as in aspartic acid. / Ph. D.

En este artículo usamos la definición de los conjuntos (α,β)-sg-abiertos para definir la (α,β) -sg-compacidad y la (α,β)-sg-conexidad de un espacio topológico (X, T) sobre el cual se tienen operadores α ,β asociados a T. Se estudian y se caracterizan los espacios (α,β)-sg-compactos y los espacios (α,β)-sg-conexos además buscamos condiciones bajo el cual se preserva la imagen de espacios (α,β)-sg-compactos y (α,β)-sg-conexos mediante funciones.

京都大学 / 新制・論文博士 / 博士(工学) / 乙第13581号 / 論工博第4212号 / 新制||工||1990(附属図書館) / (主査)教授 松原 誠二郎, 教授 中尾 佳亮, 教授 浦山 健治 / 学位規則第4条第2項該当 / Doctor of Philosophy (Engineering) / Kyoto University / DFAM

Les antibiotiques β-lactamines sont parmi les médicaments antimicrobiens les plus anciens et les plus utilisés. L'enzyme bactérienne β-lactamase hydrolyse l'antibiotique β-lactame en cassant la structure de base "anneau β-lactame". Pour identifier les nouvelles β-lactamases, une étude complète a été réalisée dans diverses bases de données biologiques telles que Human Microbiome Project, env_nr et NCBI nr. L'analyse a révélé que les séquences ancestrales putatives et les recherches de profil HMM jouaient un rôle important dans l'identification de la base de données homologue et métagénomique à distance dans l'enzyme β-lactamase existante comme matière noire. Les larges analyses phylogénétiques des β-lactamases existantes et nouvellement identifiées représentent les nouveaux clades dans les arbres. En outre, l'activité d'hydrolyse des antibiotiques β-lactamines de séquences nouvellement identifiées (provenant d'archées et d'humains) a été étudiée en laboratoire, ce qui montre l'activité de la β-lactamase. La deuxième phase de l'étude a été entreprise pour examiner l'évolution fonctionnelle des β-lactamases. Premièrement, des séquences de protéines ß-lactamase 1155 ont été extraites de la base de données ARG-ANNOT et des valeurs CMI la littérature correspondante. Les résultats ont révélé que l'activité fonctionnelle de la β-lactamase évoluait de manière convergente au sein de la classe moléculaire. La troisième phase de cette thèse représente le développement d'une base de données intégrative de β-lactamases. La base de données publique actuelle de β-lactamases a des informations limitées, par conséquence, une base de données intégrative a été développée. / The β-lactam antibiotics are one of the oldest and widely used antimicrobial drugs. The bacterial enzyme β-lactamase hydrolyzes the β-lactam antibiotic by breaking the core structure “β-lactam ring”. To identify the novel β-lactamases a comprehensive investigation was performed in different biological databases such as Human Microbiome Project, env_nr, and NCBI nr. The analysis revealed that putative ancestral sequences and HMM profile searches played a significant role in the identification of remote homologous and uncovered the existing β-lactamase enzyme in the metagenomic database as dark-matter. The comprehensive phylogenetic analyses of extant and newly identified β-lactamase represent the novel clades in the trees. Further, the β-lactam antibiotic hydrolysis activity of newly identified sequences (from archaea and human) was investigated in laboratory, which shows β-lactamase activity.The second phase of the investigation was undertaken to examine the functional evolution of β-lactamases. First, 1155 β-lactamase protein sequences were retrieved from ARG-ANNOT database and MIC values from the corresponding literature. The results revealed that the functional activity of β-lactamase evolved convergently within the molecular class.The third phase of this thesis presents development of an integrative β-lactamase database. The existing public database of β-lactamase has limited information, therefore, an integrative database was developed.

<p>У раду је главни циљ истраживања био да се утврди утицај додатог &beta;-каротина и витамина А на оваријалну активност у току периовулаторног периода код млечних крава са хроничним функционалним стерилитетом. За потребе овог истраживања одабрано је 46 негравидних крава расе Холштајн (Holstein) са функционалним стерилитетом, старости од 5-7 година. Животиње су подељене у три експерименталне групе, са комбинацијом &beta;-каротина и витамина А, само &beta;-каротина и само витамина А. Контролну групу сачињавале су краве без додатне терапије. Свим животињама на почетку истраживања извршен је гинеколошки преглед, синхронизација еструса и вештачко осемењавање. Мерен је почетни ниво &beta;-каротина у крви, ниво у време оплодње и током лутеинске фазе, затим жуто тело (corpus luteum) и ниво P4 као и проценат концепције. Резултати су показали постојање средње корелације r = 0,4 (p&lt;0,01) и r = 0,36 (p&lt;0,05) између иницијалне серумске концентрације &beta;-каротина и концентрација &beta;-каротина на дан овулације и у лутеалној фази, висок степен корелације између вредности &beta;-каротина на дан овулације и у лутеалној фази r = 0,83 (p&lt;0,01). Исто тако, резултати нису показали сигнификантну разлику у величини фоликула у групи са додатком &beta;-каротина и витамина А и контролној групи. Међутим, постоје значајне разлике између група са додатком &beta;-каротина и витамина А, витамина А и контролне групе. Позитивну колерацију налазимо између серумске концентрације &beta;-каротина током оплодње и концентрације прогестерона r = 0,33 (p&lt;0,05), као и процента концепције r = 0,39 (p&lt;0,01), те између серумске концентације &beta;-каротина 7. дана (у лутеалној фази) и концентрације прогестерона r = 0,51 (p&lt;0,01). Из анализе узајамне повезаности неопходно је да се истакну и корелације између дијаметара предовулаторних фоликула и процента правовремених овулација и концентрације естрадиола (r = 0,74; p&lt;0,01 и r = 0,57; p&lt;0,01). Исто тако, утврђена је корелација између концентрације естрадиола и концентрације прогестерона r = 0,39 (p&lt;0,05), као и висока корелација између величине функционалног жутог тела и концентрације прогестерона r = 0,82 (p&lt;0,01). Да би се остварио пуни потенцијал овог истраживања у пракси, неопходна су даља истраживања као и бољи менаџмент производње сточне хране и генерално исхране, на фармама млечних крава.</p> / <p>U radu je glavni cilj istraživanja bio da se utvrdi uticaj dodatog &beta;-karotina i vitamina A na ovarijalnu aktivnost u toku periovulatornog perioda kod mlečnih krava sa hroničnim funkcionalnim sterilitetom. Za potrebe ovog istraživanja odabrano je 46 negravidnih krava rase Holštajn (Holstein) sa funkcionalnim sterilitetom, starosti od 5-7 godina. Životinje su podeljene u tri eksperimentalne grupe, sa kombinacijom &beta;-karotina i vitamina A, samo &beta;-karotina i samo vitamina A. Kontrolnu grupu sačinjavale su krave bez dodatne terapije. Svim životinjama na početku istraživanja izvršen je ginekološki pregled, sinhronizacija estrusa i veštačko osemenjavanje. Meren je početni nivo &beta;-karotina u krvi, nivo u vreme oplodnje i tokom luteinske faze, zatim žuto telo (corpus luteum) i nivo P4 kao i procenat koncepcije. Rezultati su pokazali postojanje srednje korelacije r = 0,4 (p&lt;0,01) i r = 0,36 (p&lt;0,05) između inicijalne serumske koncentracije &beta;-karotina i koncentracija &beta;-karotina na dan ovulacije i u lutealnoj fazi, visok stepen korelacije između vrednosti &beta;-karotina na dan ovulacije i u lutealnoj fazi r = 0,83 (p&lt;0,01). Isto tako, rezultati nisu pokazali signifikantnu razliku u veličini folikula u grupi sa dodatkom &beta;-karotina i vitamina A i kontrolnoj grupi. Međutim, postoje značajne razlike između grupa sa dodatkom &beta;-karotina i vitamina A, vitamina A i kontrolne grupe. Pozitivnu koleraciju nalazimo između serumske koncentracije &beta;-karotina tokom oplodnje i koncentracije progesterona r = 0,33 (p&lt;0,05), kao i procenta koncepcije r = 0,39 (p&lt;0,01), te između serumske koncentacije &beta;-karotina 7. dana (u lutealnoj fazi) i koncentracije progesterona r = 0,51 (p&lt;0,01). Iz analize uzajamne povezanosti neophodno je da se istaknu i korelacije između dijametara predovulatornih folikula i procenta pravovremenih ovulacija i koncentracije estradiola (r = 0,74; p&lt;0,01 i r = 0,57; p&lt;0,01). Isto tako, utvrđena je korelacija između koncentracije estradiola i koncentracije progesterona r = 0,39 (p&lt;0,05), kao i visoka korelacija između veličine funkcionalnog žutog tela i koncentracije progesterona r = 0,82 (p&lt;0,01). Da bi se ostvario puni potencijal ovog istraživanja u praksi, neophodna su dalja istraživanja kao i bolji menadžment proizvodnje stočne hrane i generalno ishrane, na farmama mlečnih krava.</p> / <p>The aim of this study was to determine the influence of supplemented &beta;-carotene and vitamin A on the ovarian activity during the periovulatory period in dairy cows with chronic fertility impairment. A total of 46 non pregnant Holstein cows with fertility impairment, at the age of 5-7 years, were selected for this study. The animals in three experimental groups were supplemented with either a combination of &beta;-carotene and vitamin A, &beta;-carotene only or vitamin A only. Non supplemented animals served as controls. All animals included in the survey were gynecologically examined; the estrus was synchronized and inseminated. Initial blood levels of &beta;-carotene as well as the levels at the time of insemination and in the luteal phase as well as the ovarian structures during ovulation and formation of corpus luteum and P4 and conception rate, were monitored.<br />The results have shown an existence of medium correlation r = 0,4 (p&lt;0,01) and r = 0,36 (p&lt;0,05) between the initial serum concentration of &beta;-carotene and the concentration of &beta;-carotene on the day of ovulation, and on the luteal phase, high degree of correlation between the value of &beta;-carotene on the day of ovulation, and in the luteal phase r = 0,83 (p&lt;0,01). Besides, the results haven&rsquo;t shown a significant difference in the size of the follicle in the group with added &beta;-carotene and vitamin A, and the control group. However, there are significant differences between the groups with added &beta;-carotene and vitamin A, vitamin A, and the control group. There is a positive correlation between the serum concentrations of &beta;-carotene during fertilization, and the concentration of progesterone r = 0,33 (p&lt;0,05), the same as the percentage of conception r = 0,39 (p&lt;0,01); and between serum concentration of &beta;-carotene on the 7th day (in the luteal phase) and the concentration of progesterone r = 0,51 (p&lt;0,01). From the mutually connected analyzes the correlation between the diameters of the pre ovulation follicles and the percentage of the on time ovulations and the concentration of estradiol (r = 0,74; p&lt;0,01 и r = 0,57; p&lt;0,01 respectively), is also important to be emphasized. Besides, the correlation between estradiol concentration and progesterone concentration r = 0,39 (p&lt;0,05) was determined, as the high correlation between the size of the functional yellow body and the progesterone concentrations r = 0,82 (p&lt;0,01). For the fulfillment of this research&rsquo;s potential in practice, further research is necessary, as well as better management of the fodder and the feeding in general on the farms with milk cows.</p>

Endo-β-mannanase is an endohydrolase enzyme responsible for the breakdown of mannan-containing polysaccharides common in the cell walls of many plants. The action of endo-β-mannanase in barley, its optimum temperature and pH for action, temporal and spatial localization, activity in the presence of hormones and sugars and its effect on the seed's mechanical strength were assayed. The development of a spectrophotometric assay for endo-β-mannanase detection was also trialed. The optimum temperature and pH for these experiments were found to be 37℃ and pH 7. Using these parameters, the endo-β-mannanase enzyme was found to be initially localized in the seed coat and moved through to the endosperm over time. The detected level of enzyme activity increased in the presence of gibberellic acid and glucose, or decreased when abscisic acid was added. Similar results were seen when the embryo was removed and the endosperm and seed coat were incubated in hormone- and sugar-containing media. The presence of exogenous endo-β-mannanase did not affect the mechanical strength of the seed but there was a strong correlation between increasing endo-β-mannanase activity and decreasing mechanical strength over time. The spectrophotometric assay for quantifying endo-β-mannanase in extracts showed promise but did not reach fruition due to unexplained sources of variation. The localization and regulation of endo-β-mannanase in barley were similar to those seen in other plants, such as tomato, lettuce and coffee. These findings have biotechnological applications within the brewery industry. By increasing the mobilization of reserves such as mannan, it is thought that the seedling can utilize this secondary carbohydrate source instead of, or at least supplementing, glucose which was mobilized from starch. This will theoretically reduce the starch and glucose lost during the malting period leaving a higher sugar content free for fermentation.