This diploma thesis deals with application of evolutionary engineering on PHA producing bacterial strains. Two bacterial strains, Cupriavidus necator H16 and Halomonas halophila, were chosen for the evolutionary experiments. Copper cations (Cu2+) and sodium chloride (NaCl) were chosen as the selective pressure for C. necator H16; acetic acid (AA) and levulinic acid (LA) for Halomonas halophila. The adapted strains were during long-time evolutionary experiments characterized by GC-FID and SEC-MALS. The growth of the adapted strains was studied by the mean of optical density measurement. The amount of viable cells was determined by spectral FC after their expositon to selected stress factors. Specific enzyme activities of enzymes involved in citrate and glyoxalate cycle, enzymes generating NADPH, LA metabolism enzyme and PHA biosynthesis enzymes were determined. The adapted strains were compared with the wild-type of strains. The successfull adaptation of C. necator H16 adapted to Cu2+ was detected. Biomass and PHA production of both wild and adapted H. halophila strains cultivated in lignocellulosis waste were determined. It was found out that H. halophila adapted to the LA is capable of producing more PHA than the wild strain of this bacteria.

Submitted by Ricardo Cedraz Duque Moliterno (ricardo.moliterno@uefs.br) on 2018-01-30T22:06:24Z No. of bitstreams: 1 Disserta??o Mileide Coutinho_P?s_Defesa_COMPLETO_VERSAO_FINAL.pdf: 2079669 bytes, checksum: 16f275e93cd170ec8dbe5989847d2cf3 (MD5) / Made available in DSpace on 2018-01-30T22:06:24Z (GMT). No. of bitstreams: 1 Disserta??o Mileide Coutinho_P?s_Defesa_COMPLETO_VERSAO_FINAL.pdf: 2079669 bytes, checksum: 16f275e93cd170ec8dbe5989847d2cf3 (MD5) Previous issue date: 2017-06-20 / Coordena??o de Aperfei?oamento de Pessoal de N?vel Superior - CAPES / The cultivation of potential fruit species in the semiarid region, using a hydroponic system, arises in an attempt to diversify and boost the economy, especially small and medium producers in this region. However, studies are needed to evaluate the growth, productivity and adaptation of these species. The objective of this study was to evaluate the growth and nutritional profile of Physalis peruviana in a hydroponic floating system, considering different levels of salinity of the water artificially produced with NaCl. The experiment was carried out in a protected environment (greenhouse), where a randomized block design (RBD) was used, with four blocks, being tested five concentrations of salinity: 0; 20; 30; 40 and 60 mM NaCl. At each interval of 10 days, the following measurements were obtained: stem and root lengths, total leaf area and neck diameter and dry matter masses of plants. After 48 days of treatment, in addition to the growth analyzes, the nutrient contents of the leaves were quantified. Data were submitted to analysis of variance and analyzed by means test and regression curves. Leaf nutrient contents were only descriptive. Within the expected, the salinity of the medium restricted the growth of plants of fis?lis. The variables total dry mass (TDM), absolute growth rate (AGR) and number of leaves (NL) were those that suffered the greatest reductions with the increase of the salinity of the medium. In relation to the leaf nutrient profile, it was observed that the higher the Na + and Cl- concentration in the leaves, the lower the K +, Ca2 + and Mg2+ concentrations. According to the data of total dry mass, it was possible to determine the P. peruviana salinity tolerance index (STI), which presented mean values> 70% in most of the experimental period at concentrations of 20 and 30 mM, demonstrating that P. peruviana species is moderately tolerant to salinity. In addition, it was considered in this work that high NaCl levels led to a nutritional imbalance, where sodium ion was the main cause of interferences during growth. / O cultivo de esp?cies frut?feras potenciais na regi?o semi?rida, utilizando sistema hidrop?nico surge na tentativa de diversificar e impulsionar a economia, principalmente de pequenos e m?dios produtores desta regi?o. No entanto s?o necess?rios estudos que avaliem o crescimento, produtividade e adapta??o destas esp?cies. O presente trabalho teve como objetivo avaliar o crescimento e o perfil nutricional da Physalis peruviana em sistema hidrop?nico do tipo floating, considerando diferentes n?veis de salinidade da ?gua produzidos artificialmente com NaCl. O experimento foi realizado em ambiente protegido (estufa), onde foi empregado o delineamento em blocos casualisados (DBC), com 4 blocos, sendo testadas cinco concentra??es de salinidade: 0; 20; 30; 40 e 60 mM de NaCl. A cada intervalo de 10 dias foram obtidas as seguintes medidas: comprimentos do caule e da raiz, ?rea foliar total e di?metro do colo e as massas de mat?ria seca das plantas. Ap?s 48 dias de tratamento al?m das an?lises de crescimento foram quantificados os conte?dos de nutrientes minerais das folhas. Os dados foram submetidos a an?lise de vari?ncia e analisados mediante teste de m?dias e por curvas de regress?o. Os teores dos nutrientes foliares foram apenas descritivos. Dentro do esperado, a salinidade do meio restringiu o crescimento de plantas de fis?lis. As var?veis massa seca total (MST), taxa de crescimento absoluto (TCA) e n?mero de folhas (NF) foram aquelas que sofreram as maiores redu??es com o incremento da salinidade do meio. Em rela??o ao perfil de nutrientes foliares, constatou-se que quanto maior a concentra??o de Na+ e Cl- nas folhas, menores as de K+, Ca2+ e Mg2+. De acordo com os dados de massa seca total foi poss?vel determinar o ?ndice de toler?ncia ? salinidade (ITS) de P. peruviana, que apresentou valores m?dios >70% na maior parte do per?odo experimental nas concentra??es de 20 e 30 mM, demonstrando que a esp?cie P. peruviana ? moderadamente tolerante a salinidade. Al?m disso, considerou-se neste trabalho que altos n?veis de NaCl levaram a um desbalan?o nutricional nas plantas, onde o ?on s?dio foi o principal causador das interfer?ncias durante o crescimento.

<p>The tubuloglomerular feedback mechanism is an important regulator in the juxtaglomerular apparatus and it detects flow dependent alterations in luminal NaCl concentration ([NaCl]) at the macula densa (MD) cell site via a Na+-K+-2Cl cotransporter. Signals are sent by the MD to adjust the afferent arteriole tone and altering release of renin. This signaling mechanism is unclear but MD cell calcium concentration, release of ATP and nitric oxide (NO) might be important.</p><p>In cultured rat glomerular mesangial cells the NO production was measured using confocal microscopy and calcium responses to ATP was measured with fura-2 using imaging techniques. NO from spermine-NONOate and L-arginine could resensitize, desensitized ATP receptors in a cGMP independent way. In mesangial cells from spontaneously hypertensive rats (SHR) less NO effect was found on ATP receptor de/resensitization indicating an impaired NO release or effect.</p><p>The macula densa cells were studied using microperfusion techniques with confocal and video imaging systems. Changes in [Ca2+]i from exposed macula densa plaques were assessed upon addition of agonists added to bath. The order of efficacy of agonists was UTP = ATP >> 2MesATP = ADP. Dose response curve for ATP added in bath showed an EC50 of 15 μM. Macula densa cell volume and NO concentration increased considerably with increasing luminal [NaCl] indicating an important role for NO in the signaling process to counteract a vasoconstrictor response and reset the sensitivity of the tubuloglomerular feedback mechanism. </p><p>In conclusion, the results showed 1). NO can increase the P2Y receptor resensitization in rat glomerular mesangial cells, acting through a cGMP-independent pathway. 2) An impaired NO generation/effect on P2Y receptors in mesangial cells from SHR rats. 3) Macula densa cells possess P2Y2, purinergic receptors on basolateral and that activation of these receptors results in the mobilization of Ca2+. 4) Increased luniinal [NaCl] delivery increased cell volume and the NO productions in the macula densa cells. </p>

The tubuloglomerular feedback mechanism is an important regulator in the juxtaglomerular apparatus and it detects flow dependent alterations in luminal NaCl concentration ([NaCl]) at the macula densa (MD) cell site via a Na+-K+-2Cl cotransporter. Signals are sent by the MD to adjust the afferent arteriole tone and altering release of renin. This signaling mechanism is unclear but MD cell calcium concentration, release of ATP and nitric oxide (NO) might be important. In cultured rat glomerular mesangial cells the NO production was measured using confocal microscopy and calcium responses to ATP was measured with fura-2 using imaging techniques. NO from spermine-NONOate and L-arginine could resensitize, desensitized ATP receptors in a cGMP independent way. In mesangial cells from spontaneously hypertensive rats (SHR) less NO effect was found on ATP receptor de/resensitization indicating an impaired NO release or effect. The macula densa cells were studied using microperfusion techniques with confocal and video imaging systems. Changes in [Ca2+]i from exposed macula densa plaques were assessed upon addition of agonists added to bath. The order of efficacy of agonists was UTP = ATP &gt;&gt; 2MesATP = ADP. Dose response curve for ATP added in bath showed an EC50 of 15 μM. Macula densa cell volume and NO concentration increased considerably with increasing luminal [NaCl] indicating an important role for NO in the signaling process to counteract a vasoconstrictor response and reset the sensitivity of the tubuloglomerular feedback mechanism. In conclusion, the results showed 1). NO can increase the P2Y receptor resensitization in rat glomerular mesangial cells, acting through a cGMP-independent pathway. 2) An impaired NO generation/effect on P2Y receptors in mesangial cells from SHR rats. 3) Macula densa cells possess P2Y2, purinergic receptors on basolateral and that activation of these receptors results in the mobilization of Ca2+. 4) Increased luniinal [NaCl] delivery increased cell volume and the NO productions in the macula densa cells.

Cytochrome P450 3A isozymes (CYP3A) metabolize approximately 50% of therapeutic drugs. It has recently been discovered that human CYP3A mRNA levels can be induced by hypertonicity; a physiological state not previously linked to its regulation. The osmosensitive transcription factor, Nuclear Factor of Activated T-Cells 5 (NFAT5), regulates multiple genes that restore osmolyte homeostasis and promote cell protection during osmotic stress. In silico examinations and in vitro experiments using reporters, knockdown and binding assays in the human intestinal cell line C2bbe1 have revealed an active tonicity-responsive enhancer (TonE) within CYP3A7 intron (+5417/+5427 from CYP3A7 transcriptional start site) that is responsible for NFAT5 binding and NFAT5-dependent regulation of CYP3A isoforms. In addition, hypertonicity-mediated CYP3A induction is also observed in both hepatic and intestinal cell lines. Effects of tonicity changes on in vivo CYP3A expression and function were examined in a humanized CYP3A transgenic mouse with similar tissue expression in humans. More specifically, intervention with prolonged dehydration involving alternating between 24-hour cycles of water-deprivation and water ad lib for 1 week (cyclic water-deprivation; four 24-hour water-deprivation and three 24-hour water ad lib periods), increased expression of NFAT5 target genes Slc6a12 in the liver and kidney (2.5 ± 0.6-fold over water ad lib, n = 14, p = 0.04; and 3.1 ± 0.6-fold, n = 10, p = 0.02, respectively), Akr1b3 in the liver, and Slc5a3 in the kidney. Immunofluorescent microscopy revealed an increase of nuclear-distributed mouse NFAT5 in cyclic water-deprived animals, consistent with NFAT5 activation. Most importantly, CYP3A4 mRNA levels were noted to be elevated in the liver and kidney (11.8 ± 4.8-fold over water ad lib, n = 14, p = 0.04 and 2.2 ± 0.4-fold, n = 9, p = 0.02, respectively), with concurrent CYP3A protein and activity increase. Localized hypertonic environment in the gut was simulated by providing animals with a week-long high-salt diet. The effects of high-salt diet in the gut were similar to those of cyclic water-deprivation in the liver and kidney; where NFAT5 showed nuclear distribution and NFAT5 target gene expression (Slc6a12; 20.5 ± 6.7-fold over a week-long low-salt diet, n = 8, p = 0.02 and Slc6a6; 3.2 ± 0.7-fold, n = 10, p < 0.01, in the duodenum). Furthermore, an increase of CYP3A4 mRNA was observed (2.6 ± 0.5-fold over a week-long low-salt diet, n = 14, p = 0.03), with a corresponding rise in protein expression and activity levels. In summary, increased expression of in vitro and in vivo human CYP3A was achieved using a hypertonic stimulus; concurrent NFAT5 activation and NFAT5 target gene expression were observed. These results suggested a possible binding of activated NFAT5 to CYP3A TonE situated within the intronic region of CYP3A7. It could be further concluded that NFAT5 may be responsible for the hypertonic induction of human CYP3A.

Cytochrome P450 3A isozymes (CYP3A) metabolize approximately 50% of therapeutic drugs. It has recently been discovered that human CYP3A mRNA levels can be induced by hypertonicity; a physiological state not previously linked to its regulation. The osmosensitive transcription factor, Nuclear Factor of Activated T-Cells 5 (NFAT5), regulates multiple genes that restore osmolyte homeostasis and promote cell protection during osmotic stress. In silico examinations and in vitro experiments using reporters, knockdown and binding assays in the human intestinal cell line C2bbe1 have revealed an active tonicity-responsive enhancer (TonE) within CYP3A7 intron (+5417/+5427 from CYP3A7 transcriptional start site) that is responsible for NFAT5 binding and NFAT5-dependent regulation of CYP3A isoforms. In addition, hypertonicity-mediated CYP3A induction is also observed in both hepatic and intestinal cell lines. Effects of tonicity changes on in vivo CYP3A expression and function were examined in a humanized CYP3A transgenic mouse with similar tissue expression in humans. More specifically, intervention with prolonged dehydration involving alternating between 24-hour cycles of water-deprivation and water ad lib for 1 week (cyclic water-deprivation; four 24-hour water-deprivation and three 24-hour water ad lib periods), increased expression of NFAT5 target genes Slc6a12 in the liver and kidney (2.5 ± 0.6-fold over water ad lib, n = 14, p = 0.04; and 3.1 ± 0.6-fold, n = 10, p = 0.02, respectively), Akr1b3 in the liver, and Slc5a3 in the kidney. Immunofluorescent microscopy revealed an increase of nuclear-distributed mouse NFAT5 in cyclic water-deprived animals, consistent with NFAT5 activation. Most importantly, CYP3A4 mRNA levels were noted to be elevated in the liver and kidney (11.8 ± 4.8-fold over water ad lib, n = 14, p = 0.04 and 2.2 ± 0.4-fold, n = 9, p = 0.02, respectively), with concurrent CYP3A protein and activity increase. Localized hypertonic environment in the gut was simulated by providing animals with a week-long high-salt diet. The effects of high-salt diet in the gut were similar to those of cyclic water-deprivation in the liver and kidney; where NFAT5 showed nuclear distribution and NFAT5 target gene expression (Slc6a12; 20.5 ± 6.7-fold over a week-long low-salt diet, n = 8, p = 0.02 and Slc6a6; 3.2 ± 0.7-fold, n = 10, p < 0.01, in the duodenum). Furthermore, an increase of CYP3A4 mRNA was observed (2.6 ± 0.5-fold over a week-long low-salt diet, n = 14, p = 0.03), with a corresponding rise in protein expression and activity levels. In summary, increased expression of in vitro and in vivo human CYP3A was achieved using a hypertonic stimulus; concurrent NFAT5 activation and NFAT5 target gene expression were observed. These results suggested a possible binding of activated NFAT5 to CYP3A TonE situated within the intronic region of CYP3A7. It could be further concluded that NFAT5 may be responsible for the hypertonic induction of human CYP3A.

Die therapeutische Beeinflußbarkeit einer Leberverfettung gilt weltweit als unbefriedigend ge-löst, so daß ein letaler Ausgang besonders bei Wiederkäuern teilweise nicht zu verhindern ist. Die Nutzung von beta-Rezeptorenblockern hat bisher mit dieser Indikation keinen Eingang in die Veterinärmedizin gefunden. In den vorliegenden Untersuchungen wurden deshalb die Auswirkungen einer unspezifischen Blockade der beta-adrenergen Rezeptoren auf die Lipolyse, die klinischen und hämatologi-schen Funktionen sowie Leber-, Eiweiß- und Mineralstoffwechsel bei fastenden Schafen ge-prüft. Insbesondere wurde dabei die Wirkung einer Propranololapplikation auf die Lipolyse in der frühen Phase des Fastens untersucht. Zu diesem Zweck wurden insgesamt 15 weiblichen, klinisch gesunden, güsten Schafen der Rasse Merino-Fleisch während eines dreitägigen Futterentzuges mit Hilfe einer Dauertropfinfu-sion mit zwei 8stündigen Pausen 0,5 bzw. 1 mg Propranolol/kg KM/d bzw. den Tieren der Kontrollgruppe ein vergleichbares Volumen einer NaCl-Lösung appliziert. Neben der klini-schen Kontrolle von Puls- und Atemfrequenz, Körpertemperatur und Pansenaktivität erfolgte über die wiederholte Gewinnung von Blutproben aus der Vena jugularis externa eine Erfassung der Konzentrationen von Glucose, FFS, Bilirubin, BHB, K, Na, Mg, Gesamteiweiß und Albu-min im Blutserum. Zur Kontrolle der Leberfunktion wurden die Aktivitäten der GLDH und ASAT bestimmt. Die Wirkung des beta-Rezeptor-Antagonisten Propranolol auf die hämatolo-gischen Parameter wurde durch die Kontrolle der Leukozyten-, Erythrozyten- und Hämoglo-binkonzentrationen und den Hämatokrit der Schafe im Versuchsverlauf überprüft. In Übereinstimmung mit bisherigen Untersuchungen an Wiederkäuern und Nichtwiederkäuern kam es aufgrund der dreitägigen Futterdeprivation in allen Tiergruppen zu einer signifikanten Verminderung des Körpergewichtes um bis zu 9,8 %. Die Zahl der Pansenbewegungen redu-zierte sich bei allen Tieren signifikant bereits innerhalb der ersten 48 Stunden der Futterdepri-vation. Der stärkste Abfall der Pansenaktivität ließ sich bei den Schafen, denen 1 mg Propra-nolol/kg KM/d infundiert wurde, nachweisen. Puls- und Atemfrequenz, die Konzentrationen von Na, K, Mg, Gesamteiweiß und Albumin sowie die Aktivitäten von GLDH und ASAT im Blutserum blieben im Versuchsverlauf ohne signifikante Veränderungen. Eine Beeinflussung der hämatologischen Parameter ließ sich we-der bei den Schafen der Kontrollgruppe noch bei denen der Versuchsgruppen nachweisen. Während der Futterentzug in allen Tiergruppen zu einer tendenziellen Abnahme der Blut-serumkonzentration an Glucose führte, stiegen die Konzentrationen an FFS, Bilirubin und BHB im Blutserum aller Tiere an. Damit weisen die Veränderungen in den Konzentrationen von FFS, BHB und Bilirubin im Blutserum der Schafe der Kontrollgruppe die typischen Merkmale einer Fastenstoffwechsellage auf. In beiden Versuchsgruppen fielen diese Konzen-trationserhöhungen gegenüber denen der Kontrollgruppe statistisch gesichert niedriger aus. Unter Berücksichtigung der dabei erreichten Niveaus ließ sich für die Versuchsgruppen eine geringere Belastung von Energie- und Leberstoffwechsel als in der Kontrollgruppe feststellen. Der Übergang von der ersten in die zweite Phase des Fastenstoffwechsels ist in allen Tiergrup-pen, besonders deutlich in beiden Versuchsgruppen, zum Zeitpunkt um 48 h nach Versuchsbe-ginn an der Erhöhung der Körpertemperatur sowie den stärkeren Anstiegen der FFS-Konzentrationen erkennbar. Die besondere klinische und labordiagnostische Bedeutung der Änderungen der FFS-Konzentration im Blutserum zeigte sich in der zeitlich früheren und ausgeprägteren Reaktion als die Konzentrationsänderungen von Bilirubin und BHB. Bereits nach 24stündigem Fasten waren in allen Schafgruppen signifikant gegenüber den Ausgangswerten erhöhte FFS-Konzentrationen nachweisbar. In beiden Versuchsgruppen war bis zum Erreichen der Maxi-malwerte 48 h nach Beginn des Fastens ein geringerer Anstieg der FFS-Konzentration im Blut-serum als in der Kontrollgruppe nachzuweisen. Nach dem Erreichen der Maximalkonzentration kam es unter dem Propranololeinfluß in beiden Versuchsgruppen zu einem raschen, signifi-kantem Abfall der FFS-Konzentration um 44,7 bzw. 63,5 %. Die Abnahme der FFS-Konzentration im Blutserum vom Maximalwert bis zum Versuchsende betrug in der Kontroll-gruppe lediglich 8,6 %. und lag damit signifikant unter den Vergleichswerten der Versuchstie-re. Für die Konzentration der FFS fanden sich zum Versuchsende zwischen allen Gruppen si-gnifikante Unterschiede. Damit läßt sich eine dosisabhängige Wirkung einer Propranololappli-kation auf die Freisetzung von FFS aus den körpereigenen Fettdepots ableiten. Als Besonderheit war zu beobachten, daß sich die Konzentration der FFS in der Kontrollgrup-pe in den Zeiträumen der Infusion der NaCl-Lösung vermindert. Möglicherweise spielt hierbei der säuernde Einfluß des NaCl auf den pH-Wert im Blut eine Rolle. Unter dem Einfluß einer pH-Verminderung kommt es dabei zu einer Absenkung der Lipolyserate. Der zwischen den Infusionszeiten starke Konzentrationsanstieg der FFS in der Kontrollgruppe führt in dieser zu signifikant höheren FFS-Konzentrationen als in den Versuchsgruppen. Die Konzentrationsänderungen von direkt reagierendem und Gesamtbilirubin fielen bei den Schafen der Kontrollgruppe höher aus als bei den Tieren der Versuchsgruppen. Während die Maximalkonzentrationen für das Gesamtbilirubin in den Versuchsgruppen mit Dosierungen von 0,5 bzw. 1 mg Propranolol/kg KM/d 48 h nach Versuchsbeginn erreicht werden, ließen sich die maximalen Gesamtbilirubinkonzentrationen in der Kontrollgruppe erst 56 h nach Versuchsbe-ginn nachweisen. Die dabei vorhandenen Konzentrationsunterschiede zwischen den einzelnen Gruppen weisen auf eine geringere Belastung der Leber bei den Schafen der Versuchsgruppen hin. Auch die BHB-Konzentrationen im Blutserum der Schafe der Versuchsgruppen lagen zum Versuchsende unter denen der Tiere in der Kontrollgruppe. Damit liegt ein weiterer Indikator auf eine geringere Leberbelastung der Tiere in den Versuchsgruppen gegenüber den Schafen der Kontrollgruppe vor. Die absolut niedrigsten BHB-Konzentrationen waren bei den Schafen der Versuchsgruppe mit einer Propranololgabe von 1 mg/kg KM/d nachweisbar. In dieser Gruppe wurde die maximale BHB-Konzentration 32 h nach Fastenbeginn erreicht. Die vorliegenden Ergebnisse zeigen, daß sich mit Propranololgaben in Höhe von 0,5 bzw. 1 mg/kg KM/d beim Schaf eine Hemmung der Lipolyse innerhalb der ersten 64 Stunden eines Futterentzuges erreichen läßt, ohne dabei nachweisbaren Einfluß auf hämatologische Parameter auszuüben. Insbesondere weisen die zwischen Versuchs- und Kontrollgruppe vergleichbaren Anstiege der FFS-Konzentrationen in den infusionsfreien Zeiträumen auf den Einfluß des beta-adrenergen Antagonisten auf die Lipolyse während der Infusion hin. Die Konzentrationsände-rungen von Bilirubin und BHB in den Versuchsgruppen erfolgen in deutlich geringerem Um-fang als in der Kontrollgruppe. Damit läst sich auf eine geringere Belastung von Energie- und Leberstoffwechsel bei den Versuchstieren schließen. Die nicht signifikanten Veränderungen der Enzymaktivitäten von GLDH und ASAT bestätigen, daß durch die Anwendung von Propra-nolol keine negative Beeinflussung der Leberfunktion erfolgt. Aufgrund der stärkeren Reduzie-rung der Zahl der Pansenbewegungen und des verminderten Konzentrationsanstieges von FFS im Blutserum der Schafe, denen 1 mg Propranolol/kg KM/d appliziert wurde, gegenüber denen die 0,5 mg Propranolol/kg KM/d erhielten, ist von einer Dosisabhängigkeit der Propranolol-wirkung auszugehen. Die Applikation von Propranolol in einer Dosis von 0,5 bzw. 1 mg/kg KM/d stellt beim Schaf eine geeignete Methode dar, frühzeitig eine Verminderung der fasten-induzierten Lipolyse zu erreichen. / The treatment of the fatty liver disease is world-wide regarded as unsatisfactory, so that death of the affected ruminants is partly unavoidable. The administration of beta receptor blockers in such cases has not found its way into veterinary medicine until now. In this study, the effects of a nonspecific blockade of the beta adrenoceptors on lipolysis, clinical and hematological parameters as well as on the metabolism of the liver, proteins and minerals in fasting sheep were therefore tested. Especially, the effect of a Propranolol administration on the lipolysis in the early stage of fasting was examined. During a three-day period of food deprivation, 15 female, clinically healthy and non-pregnant sheep (Merino-Fleisch) were given 0.5 or 1 mg Propranolol/kg body weight per day via a continuous infusion with two interruptions of eight hours. During the experiment, pulse, respiration rate, body temperature and rumen activity were checked. The serum concentrations of glucose, FFA, bilirubin, beta-hydroxybutyrate, potassium, sodium, magnesium, total protein and albumin were controlled by repeatedly taking blood samples from the vena jugularis externa. The activity of the enzymes GLDH and AST were determined to check the liver function. The effect of the beta-receptor antagonist Propranolol on the hematological parameters was checked by examining the WBC, RBC, the hemoglobin concentrations and the packed cell volume. All test groups showed significant decrease in body weigth of up to 9.8 %, due to the three-day food deprivation. Even within the first 48 hours of food deprivation, rumen motility of all animals was decreasing significantly. Those sheep which received 1 mg Propranolol/kg body weight per day showed the strongest decrease in rumen activity. Pulse and respiration rate, the concentrations of potassium, sodium, magnesium, total protein and albumin as well as the activity of GLDH and AST in blood serum remained without significant changes during the experiment. Neither the sheep of the control group nor those of the experimental groups showed any influence on the hematological parameters. While the blood glucose concentration tended to be lower during the food deprivation in all groups, the concentrations of FFA, bilirubin and beta-hydroxybutyrate in blood serum were increasing. Thus, the changes of FFA, bilirubin and beta-hydroxybutyrate concentrations in the control group showed the typical characteristics of fasting metabolism. These absolute increases in concentrations were significantly lower in both experimental groups than those in the control group. With regard to the levels reached and compared to the control group, a smaller load of energy and liver metabolism could be determined in the experimental groups. The transition from the first to the second stage of the fasting metabolism in all groups (but especially in the experimental groups) was clearly discernible in the rises of body temperature and in the stronger increases of the FFA concentrations approximately 48 hours after the experiment started. The special clinical and diagnostic importance of the FFA concentration was indicated in an earlier and stronger change of the FFA concentration in the blood serum, compared to the changes of bilirubin and beta-hydroxybutyrate. As early as 24 hours after the beginning of the fasting, a significant rise of the FFA concentration in comparison to the initial concentration could be proved. In contrast to the control group the two experimental groups showed a smaller increase in the FFA concentration. All groups reached their maxima of FFA concentration 48 hours after the beginning of the experiment. Afterwards FFA concentrations in the two experimental groups were sinking fast and significantly by 44.7 % and 63.5 % respectively. In the control group, the decrease of the FFA concentration from the maximum until the end of the experiment was only 8.6 % and thus significantly lower than the comparative results. At the end of the experiment, significant differences in FFA concentrations between all groups could be proved. So, it can be assumed that there is a dose-dependent effect of Propranolol on the FFA release from the bodyŽs own fat depots. The FFA concentration in the control group was decreasing during the NaCl-infusion. This could be due to NaClŽs influence on the pH value of the blood. The reduction of the pH causes a decreasing rate of the lipolysis. The strong rise of FFA concentration between the infusion in the control group, lead to significantly higher results compared to the experimental groups. The changes of total and direct reacting bilirubin in the control group were higher than in the experimental groups. While maximum concentrations of total bilirubin acid in the experimental groups (which got 0.5 and 1 mg Propranolol/kg body weigth per day respectively) could be determined 48 hours after the experimentŽs start, maximum concentrations in the control group could be found only 56 hours after beginning of the fasting. Beta-hydroxybutyrate concentrations in the experimental groups were also lower than in the control group. These differences between the groups indicate a smaller strain of the liver in the experimental groups. The lowest beta-hydroxybutyrate concentrations could be proved in the experimental group with a dose of 1 mg Propranolol/kg body weight per day. In this group, the maximum beta-hydroxybutyrate concentration was reached 32 hours after the beginning of the fasting. These results suggest that by the administration of Propranolol at doses of 0.5 mg and 1 mg/kg body weigth per day respectively it is possible to inhibit the lipolysis within the first 64 hours after a food deprivation without effecting the hematological parameters. Particularly, the comparable increases in FFA concentrations in the experimental and control groups between the infusions indicate a direct effect of the beta-adrenergic antagonist on the lipolysis during the infusions. The changes of bilirubin and beta-hydroxybutyrate concentrations in the experimental groups were smaller than those in the control group. This suggests a smaller strain on the energy and liver metabolism in the experimental groups compared to the control group. The nonsignificant changes of the activity of GLDH and AST indicate no negative influence on the liver function after applicating Propranolol. The stronger reduction of rumen motility and the smaller increase of the FFA concentration in the group with 1 mg Propranolol/kg body weigth per day compared to the group with 0.5 mg Propranolol/kg body weigth per day shows the dose-dependent effect of Propranolol. The application of Propranolol at a dose of 0.5 mg or 1 mg/kg body weigth per day is a suitable method to early inhibit fasting-induced lipolysis in sheep.

Aerodynamically rough surfaces of forests provide for efficient air/ canopy exchange of mass, heat and momentum. In that context, the effects of forest edges come into focus, and therefore, coastal-zone forests constitute aparticular concern. Aerosol-sink modelling is of importance to the global-scalecontext, because sink strengths influence the concentration of aerosol particles in the atmosphere, and that concentration, in turn, influences climate. Dry deposition models are insufficient due to a lack of semi-empirical data and because of difficulties in parameterization of the efficiency (E) with which leaves capture aerosols. Quantifications of such parameters promote possibilities for modelling aerosol-sink processes within various canopy layers. This thesis focuses on studies of sea-salt aerosol dry deposition within models of oak canopies exposed to artificially generated aerosols in a wind tunnel. The overall goal is to advance the understanding of deposition processes in forest ecosystems. Aims are to determine capture efficiencies and deposition velocities (Vd) for oak (Quercus robur L.), to investigate E and Vd dependence on aerosol particle size, wind velocity and vegetation structural elements such as Leaf Area Index (LAI), to explore edge effects on deposition, to relate my results to natural situations in the field, and to address modelling applications. This thesis is a result of five studies. The first study is based on developing awind tunnel approach with a main focus on establishing reference conditions.The next step is to quantify E and provide estimates of how E, with respect toa well defined mass-vs-particle-size distribution, varies with wind speed. To that end, a special wash-off technique is developed. Finally, edge effects ondeposition processes are investigated. Results demonstrate that forest ecosystems would experience substantially increased deposition at edges. The findings suggest that field measurements of deposition in the interior of a forest “island” in an otherwise open landscape would underestimate the deposition to the entire forest. Results clearly indicate needs for further research on the effects of LAI on capture efficiency and deposition velocity. The obtained capture efficiencies can be translated into deposition velocities for trees with a specific leaf area. An increase of Vd with increasing wind speed is found, and is consistent with other studies. Results confirm advantages of the wind tunnel approach, including its ability to enable experiments under controlled conditions. However, several problems require that explicit sub-models be developed of wind-speed dependent effects on leaf posture in the aerosol flow field and that gradients in relative humidity close to leaf surfaces need further attention. The results also propose needs for a range of further experimental investigations regarding aerosol deposition across the complete sea-to-land aerodynamic transition.

Resumo: A cenoura (Daucus carota L.) é uma hortaliça amplamente empregada na alimentação do brasileiro, constituindo uma rica fonte de cálcio e vitamina A, de fácil digestão e baixo teor de calorias. A cultivar Brasília foi desenvolvido para ser cultivada no verão, apresentando resistência ao calor, com produtividade média de 30 t ha-1 e ciclo médio de 85 a 100 dias. O presente trabalho teve por objetivo avaliar os possíveis efeitos do estresse hídrico e salino no crescimento e nas características morfológicas da cultivar Brasília. O experimento foi conduzido em casa de vegetação, no Departamento de Engenharia Rural da Faculdade de Ciências Agronômicas, Botucatu-SP, nos meses de setembro de 2009 e janeiro de 2010. O manejo hídrico foi estabelecido com base no potencial de água no solo e foram determinados três níveis de água (-0,01; -0,04 e -0,06 MPa), fazendo-se a reposição de água evapotranspirada por pesagem dos vasos. Foram utilizadas quatro doses de cloreto de sódio (1,41; 2,50; 4,50 e 6,45 dS m-1). O ensaio constituiu-se de um fatorial 3x4 (três níveis de água x quatro doses de salinidade). Os parâmetros morfológicos observados foram: altura da parte aérea, massas fresca e seca de parte aérea; massas fresca e seca da parte radicular (bulbo), comprimento da parte radicular (bulbo), diâmetro da parte radicular (bulbo) e produtividade. Foram realizadas três avaliações morfológicas durante os 100 dias após plantio e avaliações diárias de sobrevivência. A análise estatística dos dados foi feita pela análise de variância e teste de Tukey com 5% de probabilidade. Os resultados nas condições deste experimento permitiram as seguintes conclusões: Não foram observadas diferenças significativas... (Resumo completo, clicar acesso eletrônico abaixo) / Abstract: The carrot (Daucus carota L.) is a vegetable widely used in the Brazilian diet, constituting a rich source of calcium and vitamin A, digestible and low in calories. The cultivar Brasilia was designed to be cultivated in summer, showing resistance to heat, with an average yield of 30 t ha-1 and average cycle from 85 to 100 days. This study aimed to evaluate the possible effects of water stress and salinity on growth and morphological characteristics of the Brasilia cultivar. The experiment was conducted in a greenhouse at the Department of Agricultural Engineering, Faculty of Agronomic Sciences, Botucatu-SP, on September 2009 until January 2010. The water management was based on soil water potential determined and three water levels (-0.01, -0.04 and -0.06 MPa), but with the replacement of water transpired by weighing the pots. We used four doses of sodium chloride (1.41, 2.50, 4.50 and 6.45 dS m-1). The test consisted of a 3x4 factorial (three levels of water salinity x four doses). The morphological parameters were: shoot height, fresh and dry shoot, fresh and dry weight of roots (bulb), length of roots (bulb), diameter of the root (bulb) and productivity. They performed three morphological evaluations during the 100 days after planting and daily assessments of survival. The statistical analysis was performed by ANOVA and Tukey test at 5% probability. The results allowed the following conclusions: there are not observed significant differences on plant development subjected to different minmum soil water potentials (Ψs) = -0,01; -0,04 e -0,06 MPa; the treatments that did not received saline doses showed higher development in relation to aereal part height and bulb length on three available periods (60, 80 e 100 days after planting); the applied saline doses (1,41; 2,50; 4,50 e 6,45 dS m-1 NaCl) did not statistically affect plant development in relation to fresh and dry of aereal... (Complete abstract click electronic access below) / Orientador: Antonio Evaldo Klar / Coorientador: Antonio Ismael Inácio Cardoso / Banca: Hélio Grassi Filho / Banca: Elisandro Pires Frigo / Mestre

Made available in DSpace on 2016-06-02T19:22:52Z (GMT). No. of bitstreams: 1 2738.pdf: 452486 bytes, checksum: f00e2f08ad1170bdd3a4ce8e56fc0691 (MD5) Previous issue date: 2009-05-17 / Universidade Federal de Minas Gerais / The literature shows that gavage of 2 M NaCl, a method to induce cell dehydration, induces water and 0.15 M NaCl intake in a two-bottle test. However, the hypertonic NaCl is also known as a mild gastric irritant, which possibly erodes the protective barriers of the gastric mucosa. Therefore, the present work tested the hypothesis that gavage of 2 M NaCl induces mineral intake as a response to gastric irritation. If the gastric irritation is an important factor to induce mineral solution intake, then we can predict that 1) mechanical protectors of the gastric mucosa inhibits the mineral solution intake induced by gavage of NaCl and 2) other irritants such as ethanol and acetic acid, should induce mineral solution intake as the gavage of NaCl did. In previous experiments with adult male Holtzman rats, gavage of 2 M NaCl induced ingestion of appreciable quantities of NaHCO3 and KCl solutions in a five-bottle test (water, 0.01 M KCl, 0.05 mM CaCl2, 0.15 M NaHCO3, 0.15 M NaCl), showing that the effect is not selective for NaCl intake. In subsequent experiments, the animals had access to water and the two preferred mineral solutions (0.01 M KCl and 0.15 M NaHCO3) for ingestion. The ingestion of these solutions was not inhibited by the protective agents of the gastric mucosa, such as aluminum hydroxide, or the NaHCO3, combined with gavage of 2 M NaCl. Non-sodic irritants (50% ethanol or 0.6 N acetic acid) were not able to induce any mineral intake. Furthermore, ethanol - determined by macroscopic inspection as the sole aggressive irritant of the gastric mucosa induced kaolin (hydrated aluminum silicate) intake, a mineral that protects the rat against toxemia-induced malaise. Losartan, an angiotensin II receptor antagonist, injected intracerebroventricularly, inhibited in a dose-response manner the fluid intake induced by gavage of NaCl. The results suggest that NaHCO3 intake induced by gavage of hypertonic NaCl: a) is consistent to cell dehydration rather than to a gastric irritation and b) is mediated by angiotensin II. / A literatura mostra que a gavagem de NaCl 2 M, um método para se induzir desidratação celular, induz ingestão de água e de NaCl 0,15 M em teste de dupla escolha. Entretanto, o NaCl hipertônico é também conhecido como um irritante gástrico, que aparentemente rompe as barreiras de proteção da mucosa gástrica. Portanto, o presente trabalho foi realizado para testar a hipótese de que a gavagem de NaCl 2 M produz ingestão mineral como uma resposta a irritação gástrica. Se a irritação gástrica for um fator importante para induzir ingestão de solução mineral, então podemos prever que: 1) protetor mecânico de mucosa inibe a ingestão de solução mineral induzida pela gavagem de NaCl e 2) outros irritantes, como etanol e o ácido acético, devem induzir ingestão de solução mineral da mesma maneira que o NaCl. Em experimentos preliminares com ratos Holtzman adultos, a gavagem de NaCl 2 M induziu ingestão de quantidades apreciáveis de solução de NaHCO3 e KCl em teste de cinco bebedouros (água, KCl 0,01 M, CaCl2 0,05 mM, NaHCO3 0,15 M, NaCl 0,15 M), mostrando que o efeito não é seletivo para ingestão de NaCl. Nos experimentos subseqüentes, os animais tiveram acesso à água destilada e às duas soluções minerais de preferência (KCl 0,01 M e NaHCO3 0,15 M) para ingestão. A ingestão dessas soluções não foi inibida por agentes protetores da mucosa gástrica, como hidróxido de alumínio, ou o próprio NaHCO3, combinados à gavagem de NaCl 2 M. Irritantes não-sódicos (etanol 50% ou ácido acético 0,6 N) não foram capazes de induzir ingestão de qualquer mineral. Além disso, apenas o etanol - único irritante agressivo da mucosa conforme determinado por inspeção macroscópica - induziu ingestão de caulim (silicato de alumínio hidratado), um mineral que protege o rato do mal-estar induzido por toxemia. Injeção intracerebroventricular de losartan, um antagonista de receptor de angiotensina II, inibiu de forma dose-dependente, a ingestão de líquidos induzida por gavagem de NaCl. Os resultados sugerem que a ingestão de NaHCO3 desencadeada por gavagem de NaCl 2 M: a) ocorre em resposta a desidratação celular, e não por mal-estar gástrico; b) é mediada pela angiotensina II.