Dissertationen zum Thema „Cormels“

[ES] El uso de Colocasia esculenta (L.) Schott y Xanthosoma sagittifolium (L.) Schott como materia prima en forma de almidón o harina es una alternativa sostenible y nutritiva al trigo y otros granos. Esta alternativa permite a los agricultores minimizar las pérdidas después de su cosecha y garantizar la seguridad alimentaria ya que sus propiedades nutricionales, digestivas y saludables son reconocidas por la comunidad científica. Sin embargo, la información existente sobre este rizoma dirigida a su aplicación es bastante limitada. Esta tesis expone la caracterización funcional y tecnológica de los almidones obtenidos a partir de cormos y cormelos de Xanthosoma sagittifolium (L.) Schott. Asimismo, se evaluó tecnológicamente el efecto de la combinación de enzimas, hidrocoloides, almidón de patata, harina pregelatinizada sobre la harina de los cormelos de la Colocasia esculenta (L.) Schott en el desarrollo de un pan sin gluten. Además, se evaluaron las propiedades tecnológicas y digestivas de una fórmula básica para puré desarrollada a partir de harina de cormelos de Xanthosoma spp. y Colocasia spp. Además, la revisión bibliográfica realizada permitió poner en contexto los efectos saludables demostrados clínicamente de esta materia prima y sus componentes. Se demostró que existen diferencias significativas entre el almidón de cormos y cormelos de la misma especie. Se concluyó que la harina de Colocasia esculenta (L.) Schott es una buena opción para incrementar el valor nutricional de los panes sin gluten. Entre las estrategias probadas, el pan elaborado a partir de la mezcla con almidón de patata resultó la menos aconsejable. Además, todas las estrategias aplicadas originaron panes con menor índice glucémico que sus homólogos sin gluten reportados en otros estudios. Por primera vez, este trabajo recomienda el uso de harina de cormelos de Xanthosoma sagittifolium (L.) Schott y Colocasia esculenta (L.) Schott y para desarrollar purés con un valor nutricional agregado. La revisión bibliográfica realizada permitió recopilar los efectos demostrados clínicamente, concretamente antihiperglucémicos, antihepatotóxicos, antihipertensivos, hipoglucemiantes, anticancerosos, hipolipidémicos y prebióticos, entre otros, de los compuestos bioactivos presentes en esta planta.
[CA] L'ús de Colocasia esculenta (L.) Schott i Xanthosoma sagittifolium (L.) Schott com a matèria primera en forma de midó o farina, és una alternativa per a minimitzar les perdues després de la seua collita i garantir la seguretat alimentària ja que les seues propietats nutricionals, digestives i saludables són reconegudes per la comunitat científica. Aquesta tesi exposa la caracterització funcional i tecnològica dels midons obtinguts a partir de corms i cormelos de Xanthosoma sagittifolium (L.) Schott. D'una banda és va avaluar tecnològicament l'efecte de la combinació d'enzims, hidrocol·loides, midó de creïlla, farina pregelatinitzada sobre la farina dels cormelos de la Colocasia esculenta (L.) Schott en el desenvolupament d'un pa sense gluten. D'altra banda, es van avaluar les propietats tecnològiques i digestives d'una fórmula bàsica per a puré desenvolupada a partir de farina de cormelos de Xanthosoma spp. i Colocasia spp. que es pot destinar a poblacions vulnerables amb afeccions gastrointestinals, diabètics, celíacs, entre altres. A més, es va realitzar una revisió on s'exposen els efectes saludables demostrats clínicament, dels seus components. Es va demostrar que existeixen diferències significatives entre el midó de corms i cormelos de la mateixa espècie. Per tant, es va concloure que la farina dels cormelos de la Colocasia esculenta (L.) Schott és una bona opció per a incrementar el valor nutricional dels pans sense glútens i entre les estratègies provades, el pa elaborat a partir de la mescla amb midó de creïlla va resultar la menys aconsellable, però tots van mostrar menor índex glucèmic que els seus homòlegs sense glútens reportats en altres estudis. A més, per primera vegada es va recomanar l'ús de farina cormelos de Xanthosoma sagittifolium (L.) Schott i Colocasia esculenta (L.) Schott per a desenvolupar purés amb un valor nutricional agregat. La revisió bibilogràfica realitza va permetre recopilar el efectes demostrats clínicament, en concret antihiperglucémic, antihepatotóxic, antihipertensiu, hipoglucemiant, anticancerós, hipolipidémic i prebiòtic, entre altres, dels compostos bioactius presents en aquest rizoma.
[EN] The use of Colocasia esculenta (L.) Schott and Xanthosoma sagittifolium (L.) Schott as a raw material in form of starch or flour is a nutritious and sustainable alternative to wheat and other grains. This alternative allows farmers to minimize losses after harvest and guarantee food safety since its nutritional, digestive and healthy properties are recognized by the scientific community. Nevertheless, there is still scarce information about these rhizomes that limit their application. This thesis illuminates the functional and technological characterization of the starches obtained from corms and cormels of Xanthosoma sagittifolium (L.) Schott. Likewise, the effect of the combination of enzymes, hydrocolloids, potato starch, pregelatinized flour on Colocasia esculenta (L.) Schott cormels flour toward the development of a gluten-free bread was technologically evaluated. Furthermore, the technological and digestive properties of a basic formula for pastes developed from cormels flour of Xanthosoma sagittifolium (L.) Schott and Colocasia esculenta (L.) Schott was evaluated. In addition, a review was conducted and the health effects clinically demonstrated are exposed. It was shown that there are significant differences between the starch of corms and cormels of the same species. It was concluded that the flour from Colocasia esculenta (L.) Schott cormels is a good option to increase the nutritional value of gluten-free breads. Among the strategies tested, the bread made from mixtures with potato starch was the least desirable, but all strategies tested gave breads with lower glycemic index than gluten-free counterparts reported in other studies. For the first time, this works recommends the use of cormels flour from Xanthosoma sagittifolium (L.) Schott and Colocasia esculenta (L.) Schott was recommended to develop pastes to nutritional value to gluten-free products. In addition, the bibliographic review carried out allowed the compilation and analysis of the clinically demonstrated effects, especifically, antihyperglycemic, antihepatotoxic, antihypertensive, hypoglycemic, anti-cancer, hypolipidemic and prebiotic effects, among others, of the bioactive compounds present in these plants.
Authors acknowledge the financial support of the Spanish Ministry of Science, Innovation and Universities (RTI2018-095919-B-C21), Spanish Ministry of Economy and Competitiveness (Project AGL2014-52928-C2-1-R), the European Regional Development Fund (FEDER) and Generalitat Valenciana (Project Prometeo 2017/189). Tropical Food Research Institute from Cuba provided the rhizomes.
Calle Domínguez, J. (2021). Properties of Malanga Flours and their Use in Pastes and Gluten Free Breads [Tesis doctoral]. Universitat Politècnica de València. https://doi.org/10.4995/Thesis/10251/168396
TESIS

L’objectif de cette recherche est de caractériser les aspects topographiques, fonctionnels et architecturaux de l’abbaye bénédictine de Cormery établie en Touraine, par la communauté de Saint-Martin de Tours en 791 et restée en fonction jusqu’à la Révolution française. Cette approche multi-scalaire du monastère s’appuie sur les sources textuelles et les nombreux vestiges en élévation qui incluent les bâtiments claustraux. La première partie de ce travail s’attache à analyser le contexte de fondation du monastère dans la vallée de l’Indre et tout particulièrement le lien avec la rivière. Dans un deuxième temps, une étude archéologique a été menée sur les vestiges de l’église abbatiale et de la tour-clocher de l’époque romane à partir de relevés réalisés en lasergrammétrie et photogrammétrie. Enfin, l’analyse de l’organisation spatiale de l’établissement monastique et de sa périphérie où s’est formé un bourg conclut ce travail
The purpose of this research is to characterize the topographic, functional and architectural aspects of the benedictin abbey of Cormery, founded in Touraine by the community of St Martin in 791 and which was in use until the French Revolution. This multi-scalar approach of the monastery is based on written sources and standing architectural remains which include the monastic buildings. The first part of this work is dedicated to analyze the foundation process of the monastery in the Indre Valley, especially the link with the river. Secondly, the architectural study of the remains of the abbey church and the still-standing Romanesque western tower was carried out with photogrammetric and lasergrammetric recording. Finally, we conclude this work with the analysis of the spatial organization of the monastic settlement and its periphery where a medieval market town developed

Dissertação para obtenção do Grau de Mestre em Genética Molecular e Biomedicina
There is an increasing need for promoting neuroprotection against cerebral ischemia, which is the main cause of brain damage in adults. Astrocytes are the most abundant cells inboard the central nervous system (CNS), being known as key glial cell for promoting neuronal survival and homeostasis. It is more established in nowdays that astrocytic dysfunction contributes to neurodegenerative processes. Although, carbon monoxide is a well renown as a lethal and toxic gas due to its high affinity to hemoglobin, CO exerts anti-apoptotic, anti-inflammatory and anti-proliferative functions. Recent studies showed likewise that CO induces autophagy, promoting therefore cytoprotective and anti-inflammatory effects. Autophagy is a major catabolic pathway, known as an autodigestive process that degrades cellular organelles and proteins, playing an important role in cellular homeostasis during environmental stress. Due to the great interest on the signaling and cytoprotective actions of CO, novel strategies have been put forward to exploit the potential therapeutic effects of this gaseous molecule. One of these approaches consist on the development of CO-releasing molecules (CO-RMs), compounds that deliver small quantities of CO to tissues and first identified by the group of Motterlini and co-workers. The aim of this Master thesis was to study the action of CORM-A1, a boron-containing compound that spontaneously releases CO, against cell death in primary culture of astrocytes. In particular, we examined the role of CORM-A1 in autophagy, mitophagy and cell metabolism. Here, we demonstrated that CORM-A1 promotes the induction of autophagy in primary culture of astrocytes. Furthermore,autophagy is directly involved in the cytoprotective effect of CORM-A1 against cell death. In some preliminary experiments we have shown that CORM-A1 also induced mitophagy, while autophagy and inhibition of cell death promoted by CORM-A1 seem to occur under hypoxia (5% of oxygen). This master thesis has addressed several important questions on the role of CO in astrocyte function but also opened to many other important questions on the mechanism of action of CO. For instance, future work must be undertaken in order to explore whether CO-mediated induction of reactive oxygen species (ROS), which play an important role in cell signaling, which are the factors directly involved in mitophagy and the cross-talk between apoptosis and modulation of autophagy.
e-COST,(COST Action BM1005)

During spring 1990 and 1991, ichthyoplankton surveys were conducted in lower Chesapeake Bay to estimate seasonal egg production, population biomass and the impact of predation by gelatinous zooplankton on early life stages of black drum, Pogonias cromis. Rearing experiments indicated that at least three species of sciaenid (silver perch, Bairdiella chrysoura; weakfish, Cynoscion regalis and P. cromis) were spawning in the survey area during both years. Specific identification of eggs based on previously published ranges of outside egg diameter (OED) were not reliable due to considerable overlap in diameter distributions. Analysis of weekly OED frequency revealed the presence of three modes which differed in temporal occurrence, suggesting the products of three species. Genetic typing of eggs using RFLP analysis of mtDNA confirmed the presence of three species, but demonstrated that eggs of a certain size class represented two species. Results illustrate that reliance on previously published ranges of egg diameter for specific identification of spring spawning sciaenids may overestimate the spawning biomass of black drum by as much as 50% owing to the misidentification of weakfish eggs as those of black drum. Black drum enter Chesapeake Bay in early spring and spawn throughout the day over a discrete time period from late April through May in a spatially limited area off the city of Cape Charles, Virginia. Seasonal egg production is low compared to other sciaenid stocks and estimates of spawner biomass indicate that the population is comprised of relatively few individuals. During 1990 and 1991, peak egg production of black drum occurred in the lower portion of Chesapeake Bay where potential encounter with gelatinous zooplankton predators was least likely. These observations were consistent between years with vastly different gelatinous zooplankton communities. Analysis of small-scale patterns of spatial coincidence between eggs and predators reveal that chances of physical contact are least in areas of peak production.

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Conselho Nacional de Desenvolvimento Científico e Tecnológico
This work was developed in Gardening at UFMS (Universidade Federal de Mato Grosso do Sul) NCA (Núcleo Experimental de Ciências Agrárias), from September 2004 to February 2006 having the aim of valuating nitrogen, limestone and phosphogypsum over gladiolus development and over some chemical and physical characteristics of a dystroferric red latosol. Plants were valuated by leaf analysis, maximum high, spike and floral stem length, new corm diameter, dry and live mater form shoot area, and produced cormel number. Soil physical characteristics valuated the meaning clay, the meaning clay disperse in water, flocculation degree, aggregation and aggregation stableness. Soil chemical characteristics considered water and CaCl2 pH, macro and micronutrients meaning, exchangeable acidity, effective kation exchange capacity and base saturation. The experiment analysis was done in randomized blocks sub-subdivided in parcels, considering as parcel the nitrogen proportions (150, 300 kgha-1), as sub parcel the limestone proportion (0, 2000, 4000, 8000 kgha-1) and as sub-sub parcel phosphogypsum proportion (0, 500, 1000, 2000 kgha-1), in four repetitions. Nitrogen, limestone and Phosphogypsum proportions used caused no visual deficiencies detected in gladiolus. The smaller flowering time and the bigger live matter plants were gained without using limestone and / or phosphogypsum. Limestone between 6000 and 8000 kg.ha-1, contributed into ideal pH levels
Este trabalho foi desenvolvido na área de Jardinocultura do Núcleo Experimental de Ciências Agrárias da UFMS, durante o período de setembro de 2004 a fevereiro de 2006 tendo por objetivo a avaliação dos efeitos do nitrogênio, calcário e gesso agrícola sobre o desenvolvimento de Gladiolus grandiflorus e sobre alguns atributos físicos e químicos de um Latossolo Vermelho distroférrico. As plantas foram avaliadas por meio de análise foliar e quanto à altura máxima, comprimento da espiga e da haste floral, diâmetro do novo cormo, matéria fresca e seca da parte aérea e número de cormilhos produzidos. Como atributos físicos do solo foram avaliados o teor de argila, teor de argila dispersa em água, grau de floculação, agregação e estabilidade dos agregados. Os atributos químicos considerados foram o pH em água e em CaCl2, teores de macro e micronutrientes, acidez trocáveis, capacidade de troca de cátions efetiva e saturação de bases. O experimento foi analisado como inteiramente casualizado, em parcelas sub-subdivididas, sendo consideradas como parcelas as doses de nitrogênio (150 e 300 kg.ha-1), como subparcelas as doses de calcário (0, 2000, 4000 e 8000 kg.ha-1) e como sub-subparcelas as doses de gesso (0, 500, 1000 e 2000 kg.ha-1), com quatro repetições. As doses de nitrogênio, gesso e calcário utilizadas não induziram Deficiências detectadas visualmente no gladíolo. O menor tempo de florescimento e a maior matéria fresca das plantas de gladíolo foram obtidos sem o uso de calcário e/ou de gesso. As doses de calcário entre 6000 a 8000 kg.ha-1, contribuíram para obtenção de níveis ideais de pH em água para o gladíolo. A dose de nitrogênio de 300 kg.ha-1, promoveu a redução do pH, Ca e Mg e de argila dispersa em água e aumentou o grau de floculação e os teores de Al.

In South Africa part of the cultural and religious beliefs of the African people is the use of traditional remedies to treat diseases. These remedies are obtained from medicinal plants (Steenkamp, 2003). One of the most frequently traded plants in the Eastern Cape is Hypoxis, commonly known as Afrika patat, or African potato. South African traditional healers instruct patients to brew the fresh Hypoxis corm as a tea and then ingest it (Steenkamp, 2006a). This prompted an investigation into the digestive stability of a traditionally prepared Hypoxis extract. The H. sobolifera extracts were digested using a simulated gastric/small intestinal digestion and their biological activity determined. The hot water H. sobolifera extract before digestion only showed cytotoxic activity against cancer cell lines at very high concentrations which are not likely to be achieved under normal ingestion circumstances. In Chang liver cells on the other hand, chronic exposure to the hot water H. sobolifera extract increased glucose uptake in amounts similar to that of metformin. On the negative side, the glucose utilization stimulation was lost due to the simulated digestion process. The significant inhibition of AGEs by hot water H. sobolifera extract (IC50 of 6.3 Ig/ml) is a very encouraging result as treatment in the management of diabetes. This activity was only slightly reduced by the in vitro digestion process. Also observed was enzyme inhibition activity by traditionally prepared H. sobolifera, with ∝-amylase being inhibited (IC50 of approximately 250 Ig/ml) and therefore preventing or limiting starch breakdown. From the DPPH results it was clear that H. sobolifera, even when digested, is a potent anti-oxidant (IC50 of 134.4 Ig/ml when undigested compared to 162.9 when digested with β-glucosidase added to stomach digestive step). HPLC and TLC experiments revealed that rooperol which has previously been thought to be the compound responsible for the anti-oxidant activity in Hypoxis extracts, was absent from the traditional extract of H. sobolifera and therefore cannot be the sole compound exhibiting anti-oxidant activity; other compounds such as phenolics may be contributing. The phenolic and flavonoid content results revealed very highconcentrations of these compounds in the traditionally prepared H. sobolifera extract. These compounds may therefore play major roles in all of the biological activities observed from treatment with Hypoxis spp. The ROS results yielded interesting and promising results. Using standard or traditionally prepared H. sobolifera extracts, activation of differentiated U937 cells with PMA was greatly enhanced by cotreatment with the extracts, while extracts on their own did not cause significant activation. Future studies should investigate this property of the extracts as a promising immune boosterThe HPLC results showed that hypoxoside was undetectable in the hot water traditional extract and the TLC anti-oxidant experiment proved that rooperol is not present in the hot water traditional extract after treatment with β-glucosidase. This indicates that neither one of the Hypoxis compounds previously believed to be responsible for the biological activities observed are present in the extract when prepared the traditional way. Therefore, the biological activities observed in this study can be attributed to other phytochemical compounds.

Antibiotic-resistant pathogenic bacteria pose a major, growing public health risk. Antibiotics affect specific targets so that bacteria may develop cognate resistance to the target site. To cope with antibiotic resistance, combinatorial therapies involving two (or more) antimicrobial agents may be useful for minimizing antibiotic resistance phenomena. It is, therefore, necessary to investigate other antimicrobial agents. Carbon monoxide-releasing molecules (CORMs) have been shown to exert antimicrobial actions on several bacterial species both in the laboratory and in animal models. The aims of this thesis were to investigate the antibacterial effects of CORMs on the bacterial growth and viability for both strains of E. coli wild type (MG1655) and multidrug-resistant uropathogenic E. coli (EC958) and more importantly to test their interactions with antibiotics. The results revealed that CORMs have antimicrobial effects against both strains of E. coli. Importantly, a combination of sub-inhibitory concentrations of CORMs and antibiotics showed a significant potency on the action of conventional antibiotics with reduction of bacterial growth and viability. CO gas showed only slight antibacterial effects and there was no effect of CO gas on antibiotic activities when combined. Furthermore, conjunction of CORM-2 or CORM-3 with doxycycline, minocycline, gentamicin or chloramphenicol revealed that minimal inhibitory concentration (MIC) values of the antibiotics decreased between 16 - 640 fold with CORM-2 and 13 - 60 fold with CORM-3; the minimal bactericidal concentration (MBC) values were also reduced 2 - 6 fold with CORM-2 and 1 - 150 fold with CORM-3. Moreover, the drug interactions between CORMs and antibiotics were assessed using checkerboard microdilution methods, time-killing curves and Etests; the results revealed that CORMs have synergistic interactions with several antibiotics such as trimethoprim, novobiocin and doxycycline with fractional inhibitory concentration indexes (FICi) between 0.35 and 0.50. Real-time PCR data showed that CORM-2 significantly upregulates the transcription of recA and spy, which encode proteins for DNA recombination and repair and an envelope-stress-induced periplasmic protein respectively, However, there were no significant changes in the transcriptional level of sodA and katG, which encode for oxidative stress enzymes. Furthermore, ruthenium-based CORMs caused DNA degradation and also membrane leakage of nucleic acids in a concentration-dependent manner. Finally, CORM-2 produces a significant amount of ROS when assayed using a DCF-DC probe. However, using the HFP probe that selectively measures the generation of hydroxyl radical (OH^), we showed that CORM-2 does not produce OH^. Furthermore, the Amplex Red assay revealed that CORM-2 at lower concentrations was not able to produce H2O2 while 4 μM H2O2 was produced at 150 μM CORM-2. In conclusion, these findings reveal that the CO released from CORMs, especially CORM-2 and CORM-3 has bactericidal activity and, more importantly, these CORMs showed synergistic interactions with various antibiotics against both strains of E. coli.

Carbon monoxide (CO) is now well established as one of the signaling molecules in higher organisms, including humans. Due to its physiological roles, CO is now accepted as a potential therapeutic agent. The use of CO gas has been studied in multiple clinical trials. Vasodilation, anti-inflammatory, anti-apoptotic, anti-proliferative and cytoprotective effects are just a few of the pharmacological actions attributed to CO gas in various models of diseases. Use of inhaled CO gas as a therapeutic has many limitations which necessitate the development of a new approach for CO delivery. In order to handle CO safely, compounds that release CO (CO-releasing molecules, CORMs) have been developed. CORMs that release CO only when triggered, and with the ability to target certain tissue sites, are of particular interest. Our lab is developing molecules that release CO only when illuminated with visible light (photoCORMs). These photoCORMs are based on a motif found in naturally-occurring flavonols, which are chemical compounds found in wide variety of foods including fruits, vegetables, tea and dark chocolate. The research presented in this dissertation outlines the results of studies on extended flavonols as CO release agents. The specific studies described herein focus on understanding visible light-induced CO-releasing flavonols in terms of their: a) structure/reactivity relationships, especially in biological environments; b) interactions with metal ions and proteins; c) reaction pathway of CO release; and d) their properties when combined with a CO-sensing motif.

Dissertação para obtenção do Grau de Mestre em Biotecnologia
Outrora considerado um produto tóxico, reconhecem-se, hoje, ao monóxido de carbono (CO) efeitos biológicos e terapêuticos (nomeadamente anti-inflamatórios); neste sentido, desenvolveram-se metodologias para a sua aplicação destacando-se as Moléculas Libertadoras de Monóxido de Carbono (CORMs). Perspectivando a passagem a fármacos, é indispensável elucidar os mecanismos de acção e de transporte no organismo que dependem, significativamente, das interacções estabelecidas com proteínas sanguíneas (hemoglobina, albumina e transferrina). Assim, no presente trabalho, empregaram-se técnicas de Cristalografia de Raios-X e de Espectroscopia (Ultra-Violeta/Visível e Infra-vermelhos) para caracterizar interacções entre as referidas proteínas (juntamente com lisozima) e possíveis CORMs desenvolvidos pela empresa Alfama. Os estudos com a lisozima indicam a formação de aductos com o ALF 850 confirmando-se, por Cristalografia, a ligação do composto à Histidina 15 com a libertação de 2 moléculas de CO e comprovando-se o respectivo efeito terapêutico. Analogamente, conseguiram-se dados indicativos da formação de aductos entre a hemoglobina e o ALF 850, embora seja necessário prosseguir a investigação no sentido de caracterizá-los estruturalmente. Optimizou-se ainda a sobre-expressão e a purificação da transferrina em Pichia pastoris. Efectuaram-se vários ensaios de screening para determinar condições de cristalização da albumina e da transferrina, não se obtendo, contudo, cristais de proteína para o seguimento dos estudos envolvendo os CORMs.

碩士
國立臺灣大學
園藝學系
81
Corm and cormel production of gladioli was studied in southern part of Taiwan. The best time to harvest corms and cormels was two to four weeks after flowers harvesting when using No.3 medium-size corms to produce cut flowers and corms of gladiolus during winter in southern part of Taiwan.It was suggested to leave four leaves on stem at harvesting flowers to get both favorable-length stalks and large quantity corm production. ''Spic and Spic''was proved to be a heat-tolerance cultivar for summer crops. Cormels were planted after October when the temperature became cool. With the increasing of days after cormels plan- ting, mother cormels and absorbing roots would shrink and weaken. Weight of shoot was increasing at beginning, and then leaves turn senescent after all leaves were emerged. Changes of weight in contractile roots showed the same trend as that in shoot, and then contractile roots would contract and turn fiberous. Basal part of shoot began to swell up af- ter planting and formed into daughter corm. When two leaves emerging, buds of stolon could be observed on sheath leaves nodes between daughter corm and mother corm.Stolon tip would swell and then cormels were formed. With the growth and development of corm, starch content was increasing, and it could accumulate up to 40% dry weight of corm. Dormancy of cormels could be broken totally after 5 mon- ths of 5Ｃ storage. According to results of three corm production trials, the potential for corm production in Taiwan was discussed. We could get No.3 and No.2 corms after 4 to 5 months of cormels planting.It''s suggested to cut spikes competing nutrition in corm production. Smaller sized cormels (dia. below 5mm) were of potential to produce corms.

碩士
國立中興大學
園藝學系所
94
The study is about the effect of Ethrel on sprouting and development in the different cormel size of Gladiolus ’ Princess Margaret Rose’ cormlets from anther culture in vivo and in vitro. in order to accelerate the speed of breaking the physiological dormancy of the cormel, recover the growth, shorten the time of culture. The situation of dormancy of Gladiolus ’ Princess Margaret Rose’ cormlets from anther culture had been changed by the different cormlet size, the dormancy were strong in the small cormlte size, and were weak in the large one. The study result could be shown that the sprouting of the cormlets of Gladiolus ’ Princess Margaret Rose’ were 32% in diameter 1~15 cm were higher than when the sprouting were 18% in diameter 0.6-0.9 cm, and the sprout were 10% in diameter <0.6 cm. To affect the corm in vitro with the various concentration of Ethrel in 0, 2.5, 5,and 10 mM, at the fifth week in 5 mM Ethrel, the sprouting were 80% (the medium corm) and 78%(the large corm), both of them had the higher sprouting than others, and the fresh weight were 792 mg(the medium cormlets) and 1453 mg(the large cormlets), also higher than others. The study result could be shown that when Ethrel in 5 mM could break the physiological dormancy of the cormlets of Gladiolus ’ Princess Margaret Rose’ cormlets from anther culture in vitro, recover the growth, shorten the time of culture efficiently. The sprouting did not raise in medium and large cormlets when we used the 0, 5, 10 mM concentration of Ethrel in Gladiolus ’ Princess Margaret Rose’ cormlets from anther culture in vivo. The sprouting were lower than 25% in the medium cormlets, and 45% in the large cormlets. And the Ethrel did not work on the germinated cormlets, and relative growth ratio were getting down when the culture time were increased, and it were easily to be observed the appearance of blight on the top of the leaf. The blight could be more serious when in the higher concentration of Ethrel. The effect of the low temperature storage in the sprouting of the cormlets of Gladiolus ’ Princess Margaret Rose’ cormlets from anther culture in vitro, we could see that at the lower 5℃temperature for one month and took it out for 3 weeks, the medium and large cormlets’ sprout rate were 23% and 28%, but in the 25℃, the medium and large cormsel spouting had no obvious different, and the spout rate became lower. It had shown that the low temperature could not break the physiological dormancy of the cormlets of Gladiolus ’ Princess Margaret Rose’ cormlets from anther culture in vitro efficiently. The result of the experiment had showed that, when Ethrel in 5 mM could break the physiological dormancy of the cormlets of Gladiolus ’ Princess Margaret Rose’ cormlets from anther culture in vitro, recover the growth efficiently, and it were more effective to break the physiological dormancy of the cormlets of Gladiolus ’ Princess Margaret Rose’ cormlets from anther culture in vitro more that in the lower temperature storage, Ethrel in 5 mM it helped to increase the sprouting and shorten the time of culture.

碩士
國立清華大學
分子與細胞生物研究所
104
Nymphaea capensis is an aquatic plant which produces recalcitrant seeds. Its’ seeds were desiccation intolerant and metabolically active. Cryopreservation is suggested because the conventional methods could not provide a stable storage environment for the seeds. Vitrification pretreatment was used to avoid the formation of ice crystals in cryopreservation. This study showed that cryopreserved seeds which progressed 12 h and 18 h of plant vitrification solution 3 (PVS3) treatments brought the highest germination rate (66.7%) among various pretreatment periods. The glass transition temperature (Tg) of 75% w/v sucrose + 25% w/v propylene glycol (PG) solution (-63.6℃) had the largest improvement among the alternative vitrification solutions compared to PVS3 (-90.7℃). Isoetes taiwanensis is a semiaquatic plant which is listed as a critically endangered species based on the criteria of IUCN. In contrast to the seeds of N. capensis, the plants of I. taiwanensis could tolerate air-dried for ca. one year in room temperature. Air-dried corms were retained 10% survival rate after 11 months storage under 4℃ and able to regrow new leaves and roots after rehydration. However, corms completely lost viability in cryopreservation even if various pretreatments were used.

Kohlenstoffmonoxid ist ein wichtiges kleines Signalmolekül das im menschlichen Körper durch die enzymatische Wirkung von Häm-Oxygenase (HO) auf Häm produziert wird. Für eine thera-peutische Anwendung werden Metallcarbonyl-Komplexe als CO-releasing molecules (CORMs) untersucht, die eine kontrollierte Freisetzung in biologischen Zielstrukturen erlauben. Dafür wird entweder die Ligandenperipherie ("drug sphere") modifiziert oder die CORMs an bio-molekulare Trägersysteme konjugiert. Im Rahmen dieser Arbeit stand dabei die lichtinduzierte Freisetzung von Kohlenstoffmonoxid aus Mangan(I)tricarbonyl-Komplexen im Vordergrund. Die oktaedrische Koordinationssphäre des Metallzentrums wurde dabei durch verschiedene faciale tridentate Liganden komplettiert, welche außerdem eine einfache und modulare Verknüpfung mit biologischen Träger-molekülen ermöglichen sollten. Als Chelatoren wurden Derivate von N,N-Bis(pyridin-2-ylmethyl)amin (bpa) ausgewählt, in denen das zentrale Stickstoffatom mit Alkylaminen unterschiedlicher Kettenlänge funktionalisiert ist, welche über Amid-Bindungen mit Carboxylat-modifizierten Trägermolekülen verknüpft werden können. Diesen bpa-Liganden sollte ein neuartiges Ligandensystem auf der Basis von N-(Phenanthridin-6-ylmethyl)-N-(chinolin-2-ylmethyl)ethan-1,2-diamin (pqen) gegenübergestellt werden, in denen die Phenanthridin-Gruppe interessante photophysikalische und photochemische Eigenschaften erwarten lässt. Die CO-releasing molecules sollten zudem mit den isostrukturellen Rhenium(I)tricarbonyl-Komplexen verglichen werden, die als Marker für die Fluoreszenz-mikroskopie dienen
In many organisms, carbon monoxide is generated in a controlled fashion by the degradation of heme by heme oxygenase (HO) enzymes. This small signaling molecule is involved in the control of blood pressure and possess anti-inflammatory, anti-apoptotic, and cytoprotective properties. However, a key issue is the tissue-specific delivery of carbon monoxide without concomitant formation of elevated toxic levels of CO in blood. In that context, metal carbonyl complexes show great potential for a safe CO delivery in a spatially and temporally well-controlled manner. Such CO-releasing molecules (CORMs) are composed of an inner "CORM sphere", which determines the CO release kinetics, and an outer "drug sphere", which controls bioavailability and tissue-specific uptake. In the context of this work, a series of photoactivatable CO-releasing molecules based on manganese(I) tricarbonyl groups was synthesized. In these systems, the octahedral coordination sphere of the metal center is completed by a variety of facial tridentate N^N^N ligands. Derivatives of bis(2-pyridylmethyl)amine (bpa) were selected as the chelator, in which the central tertiary nitrogen atom is functionalized with alkylamines of different chain lengths that can be linked to carboxylate-modified biological carrier molecules via amide bonds. The series of bpa ligands was contrasted with a novel ligand system based on N-(phenanthridin-6-ylmethyl)-N-(quinolin-2-ylmethyl)ethane-1,2-diamine (pqen), in which the phenanthridine group possesses interesting photophysical and photochemical properties. The series of CO-releasing molecules was complemented with the isostructural rhenium(I) tricarbonyl complexes, which might serve as markers for fluorescence microscopy

碩士
南臺科技大學
生物科技系
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Paraquat (PQ) is a widely used herbicide in the world but highly toxic to both humans and animals. After ingestion, PQ is rapidly absorbed into the bloodstream and concentrates primarily in pulmonary tissue, and then develops into irreversible pulmonary fibrosis, which is the primary cause of death. Pulmonary fibrosis is characterized by fibroblast proliferation, myofibroblast differentiation, and deposition of extra-cellular matrix in the lung parenchyma. Carbon monoxide (CO) is toxic to humans and animals, recent studies suggest low dose, 10 to 500 ppm, CO has anti-inflammatory effects, and protection against hypoxic as well as ischemic lung injury. Studies revealed Carbon monoxide releasing molecules (CORMs) can improve viability of paraquat-treated cardiomyocytes and decrease pulmonary fibrosis induced by bleomycin. These findings suggest CORMs may have therapeutic effect on paraquat-induced pulmonary fibrosis. This study showed first in vitro CORMs (CORM-3 or CORM-A1) has dose related anti-oxidant effects (free radicals, ABTS+•, DPPH•, scavengers). In vitro cell study, 20 μM CORM-3 can improve cell injury of RAW264.7/A549/MRC-5 cell after paraquat/H2O2-treatment 24 hours (increase survival rate of cells).Pretreatment with 20-100 μM CORMs (CORM-3 or CORM-A1) can decrease high level of CTGF and ANGII which released by paraquat-treated A549/MCR-5. In vivo animal study showed intraperitoneal injection of CORMs improve the survival for 14 days of mice fed with paraquat, and decrease overexpression of collagen fiber (PAS and Masson stain) and TGF-β (IHC stain). By western blot analysis, 10 mg/kg CORMs (CORM-3 or CORM-A1) decrease protein expression of COX-2, α-SMA and TNF- increased by paraquat. We also found prescription with 5 mg/kg CORMs (CORM-3 or CORM-A1) decrease serum level of Hydroxyproline of mice fed with paraquat. Overall, this study indicates that CORMs would be an attractive therapeutic approach to attenuate the progression of pulmonary fibrosis induced by paraquat.

RESUMO: Doenças neurológicas refere-se a qualquer tipo de patologia que afecta o cérebro, espinal medula e sistema nervoso periférico. Entre outras, podemos identificar doenças como o acidente vascular cerebral (AVC), epilepsia, doenças psiquiátricas, demência, infecções neurológicas e tumores. A Organização Mundial da Saúde (OMS) estima que, em 2030, quase 8 mil milhões de pessoas irão morrer devido a este tipo de doenças. A fisiopatologia e progressão dos distúrbios do Sistema Nervoso Central (SNC) são, na maior parte das vezes, multifatoriais e mal compreendidos, tornando um tratamento eficiente particularmente complicado. Porém, existem vários mecanismos característicos de disfunção e dano. Neuroinflamação descontrolada é uma das causas principais de propagação de dano e desequilíbrios homeostáticos. Desregulação de neurogénese adulta e morte de populações neuronais especificas são outra característica patológica. Descobertas recentes têm ajudado a esclarecer mecanismos moleculares de doença, mas muitas questões permanecem sem resposta. Desta forma, existe um esforço contínuo para melhor entender estas patologias e encontrar novas alternativas terapêuticas. O monóxido de carbono (CO) é uma molécula gasosa endógena, que é protetora em vários tecidos, incluindo no SNC. O principal objectivo desta tese de Doutoramento foi entender de melhor forma os mecanismos de neuroprotecção do CO. O efeito modulatório do CO foi estudado em dois processos biológicos diferentes: neurogénese e neuroinflamação. Para isto, vários modelos experimentais foram implementados. O Capítulo I introduz conceitos teóricos relativos ao cérebro, ao neurónio e às células da glia. É também feita uma descrição do estado-da-arte relativo às vias moleculares de diferenciação, função da microglia na imunidade e homeostasia, comunicação microglia neurónio, fagocitose e morte celular. Existe um foco nas propriedades biológicas do CO. No Capítulo II, é reportado que o CO reforça a produção de neurónios num modelo de diferenciação neuronal usando a linha celular de neuroblastoma SH-SY5Y. O efeito estimulatório do CO na diferenciação neuronal havia sido relacionado com modulação de morte celular e melhoramento do metabolismo mitocondrial em outros modelos. Aqui, o CO melhorou a neurogénese na linha celular SH-SY5Y, ao actuar sobre o enzima Glucose-6-fosfato desidrogenase (G6PD) e aumentar o fluxo da via das pentoses fosfato. O efeito aqui reportado foi independente da síntese de novo de glutatião. Knockdown da expressão do enzima limitante da via, G6PD, bloqueou a estimulação da diferenciação neuronal induzido pelo CO. Sumariamente, o CO promove uma reprogramação metabólica que tem um papel importante no destino da célula e em mecanismos de diferenciação neuronal O Capítulo III foca-se no CO como mediador da comunicação remota neurónio-microglia. Utilizando um protocolo de meio condicionado, o CO protegeu os neurónios incubados com meio pro-inflamatório de microglia. O CO inibiu a elevada secreção de TNF-α e NO em microglia reactiva. Além disso, o envolvimento do CO na comunicação neurónio-microglia foi também avaliado na ausência de um estímulo inflamatório. O meio condicionado de microglia tratada com CO protegeu parcialmente os neurónios contra morte celular induzida via stress oxidativo. Adicionalmente, o sobrenadante de microglia tratada com CO promoveu um aumento na complexidade morfológica neuronal. Este efeito neurotrófico gerado pela microglia tratada com CO parece ser dependente de um aumento na secreção microglial de IL-10. Este capítulo indica que o CO tem um papel anti-inflamatório celular não-autónomo, promovendo neuroprotecção através da regulação da função microglial e comunicação unidirecional com os neurónios. Aqui, foi demonstrado pela primeira vez que o CO estimula a função neurotrófica da microglia. Um sistema de cultura de células em microfluídicas é usado no Capítulo IV para explorar o interactoma directo neurónio-microglia no contexto neuroinflamatório, focando também na regulação da fagocitose. Administração de CO inibiu a secreção de mediadores inflamatórios por parte da microglia e limitou apoptose neuronal e atrofia morfológica. Sob condições inflamatórias, a remoção de conteúdo sináptico por parte da microglia decresceu. Porém, o CO reverteu este efeito, indicando que esta molécula melhora a fagocitose de sinapses sob condições de stress. Em conjunto com a poda sináptica, o CO também regulou a fagocitose microglial de neurónios durante a inflamação. Na verdade, o CO teve um efeito de controlo de qualidade na fagocitose de neurónios: bloqueou a remoção disfuncional de células viáveis, mas promoveu a fagocitose de resíduos neuronais e material apoptótico. Este mecanismo aparentemente homeostático, promovido pelo CO, não havia sido descrito e experiências estão a decorrer de modo a entender as vias moleculares que o regulam. Por último, a discussão final é apresentada no Capítulo V, integrando a informação de todos os capítulos. Aqui, a inovação e impacto geral dos resultados são contextualizados tendo em conta a literatura existente. Melhorias experimentais e próximos passos são discutidos. Em suma, esta tese de Doutoramento oferece uma forte contribuição para o entendido de mecanismos chave no funcionamento do sistema nervoso. Adicionalmente, contribui para a melhor compreensão do CO como molécula citoprotectora e homeostática no SNC.
ABSTRACT: Neurological disorders refer to any pathology that affects the brain, spinal cord, and peripheral nerves. These include stroke, epilepsy, psychiatric disorders, dementia, neuroinfections and tumours. The World Health Organization (WHO) estimates that, in 2030, almost 8 billion people will die because of neurological complications. Moreover, it reveals that formal care costs related to neuropathology morbidities will continue skyrocketing in the future. The pathophysiology and progression of central nervous system (CNS) disorders are often multifactorial and poorly understood, which makes efficient treatment particularly difficult. There are several hallmark mechanisms of dysfunction: Uncontrolled neuroinflammation is a key driving force for damage propagation and homeostatic imbalances. Dysregulation of adult neurogenesis and loss of specific neuronal population are also neurological hallmarks of ageing and disease. Current day findings have helped to illuminate the underlying mechanisms of disease, but several questions remain unanswered. Therefore, there is an ongoing struggle to further disclose neurological disease aetiology and find new therapeutic alternatives in brain pathologies. Carbon monoxide (CO) is an endogenous gaseous molecule, which confers protection in different tissues, including the CNS. The main aim of this PhD thesis was to better understand the mechanisms of COinduced neuroprotection. The modulatory role of CO was addressed on two distinct processes: neurogenesis and neuroinflammation. Different models were employed. CO-releasing molecules (CORMs) were used to deliver CO in vitro. Chapter I introduces concepts regarding the brain, neuron, and glial cells. Additionally, state-of-the art knowledge about molecular pathways of neurodifferentiation, microglia function in immunity and homeostasis, as well as microglia-neuron communication, phagocytosis, and cell death. There is a great focus on the biological role of CO. In Chapter II, it is reported that CO enhances neuronal production yield in an SH-SY5Y neuroblastoma cell line differentiation model. The CO’s stimulatory effect on neuronal differentiation had previously been linked to modulation of cell death and improvement of mitochondrial metabolism in other models. In here, CO improved SH-SY5Y neuroblastoma cell line neurogenesis by acting on the Glucose-6-phosphate dehydrogenase (G6PD) and enhancing Pentose Phosphate Pathway (PPP) flux. The reported effect was independent of de novo Glutathione (GSH) synthesis. Knocking down the expression of the rate-limiting enzyme G6PD blocked the CO-induced improvement of SH-SY5Y differentiation. In summary, CO-driven metabolic reprogramming has a central role in cell fate and neuronal differentiation processes. Chapter III focuses on CO as a mediator of neuron-microglia remote communication. Neuronmicroglia conditioned media protocol revealed that CO provides cytoprotection to neurons exposed to microglia pro-inflammatory media. In fact, CO suppressed exacerbated microglia reactivity by limiting Tumour necrosis factor α (TNF-α) and nitric oxide (NO) secretion. Furthermore, the effect of CO on neuronmicroglia communication was also assessed in the absence of an inflammatory context. CO-treated microglia conditioned media partially protected neurons against cell death induced by oxidative stress. Moreover, supernatant from CO-treated microglia improved neuronal morphological complexity. This neurotrophic effect generated by CO-treated microglia might be dependent on microglia increased secretion of Interleukin-10 (IL-10). This chapter demonstrates that CO has an anti-inflammatory non-cell autonomous role, providing neuroprotection via regulation of microglia function and unidirectional communication with neurons. It was demonstrated for the first time that CO enhances microglial neurotrophic functions. After exploring CO’s mediation of remote communication (secretome) between neurons and microglia, a microfluidic cell system is used in Chapter IV, to explore neuron-microglia direct interactome during inflammation, while also focusing on the phagocytic process. CO’s administration inhibited microglia secretion of inflammatory mediators and limited neuronal apoptosis and morphological atrophy. Focusing on the direct cell contact, microglial clearance of synaptic material is decreased under inflammatory conditions, while CO reverted phagocytosis to control levels, indicating that CO improves microglial removal of synaptic content under stress conditions. Along with synaptic pruning, CO also regulates microglial phagocytosis in an inflammatory context. In fact, CO had a quality control effect on microglia engulfment of neuronal cells under inflammation by blocking removal of viable neurons but enhancing the phagocytosis of neuronal debris and apoptotic cells. This apparent homeostatic role of CO had never been described, and the underlying molecular pathways are under study. Lastly, a final discussion is presented in Chapter V, which integrates information from all chapters. The novelty and overall impact of the thesis generated data are contextualized in the scope of the existing literature. Experimental shortcomings and future steps are discussed. Altogether, this PhD thesis provides a strong contribution to comprehending key CNS mechanisms. Additionally, it contributes for the better understanding of CO as a potentially therapeutic cytoprotective and homeostatic molecule.

Substituted tetrazines have been found to undergo facile inversed electron demand Diels-Alder reactions with “tunable” reaction rates. By varying the substituents on tetrazine, cycloaddition rate variations of over 200 fold have been achieved with the same dienophile. Coupled with the availability of different dienophiles, such as norbornene, the reaction rate difference can be over 14,000 folds. These substituted tetrazines can be very useful for selective labeling under different conditions. This finding paves the way to utilize tetrazine conjugation reactions for not only DNA but also stage labeling work. Carbon monoxide (CO) belongs to the gasotransmitter family of signalling molecules in the mammalian systems with importance on par with that of NO and H2S. Studies have shown that endogenous production of CO has anti-inflammatory, anti-proliferative, and anti-apoptotic effects in mammalian system. Besides of the conventional metal-based carbon monoxide releasing molecules (CORMs) to deliver CO for therapeutic purposes, organic CO prodrugs represent a new direction. Here we report the “click and release” approached to release CO. Unlike the metal-based CORMs, our system does not contain transition metal and liberates CO with controllable manner and possesses potential tunable releasing rate property under physiological conditions.

Romulea is a genus with numerous attractive and endangered species with horticultural potential. This genus in the Iridaceae has its centre of diversity in the winter-rainfall zone of South Africa. This thesis uses ecophysiological and biotechnological techniques to investigate the physiology behind the propagation of some species in this genus. The ecophysiological techniques of soil sampling and analysis and germination physiology were used to determine the natural and ex vitro growth and development requirements of these plants, while biotechnological techniques are used to determine the in vitro growth and development requirements of these plants and to increase the rate of multiplication and development. Soil sampling and analysis revealed that R. monadelpha and R. sabulosa, two of the most attractive species in the genus, grow in nutrient poor 1:1 mixture of clay and sandy loam soil with an N:P:K ratio of 1.000:0.017:0.189 with abundant calcium. To investigate the physical properties of the seeds, imbibition rate, moisture content and viability of seeds were determined. The seed coat and micropylar regions were examined using scanning electron microscopy. To test for suitable stimuli for germination, the effect of temperature and light, cold and warm stratification, acid and sand paper scarification, plant growth promoting substances, deficiency of nitrogen, phosphorous and potassium, and different light spectra (phytochromes) on germination were examined. An initial germination experiment showed germination above 65% for R. diversiformis, R. leipoldtii, R. minutiflora and R. flava seeds placed at 15°C; while seeds of other species placed at 15°C all had germination percentages lower than 30%. More extensive germination experiments revealed that R. diversiformis and R. rosea seed germinate best at 10°C, R. flava seed germinates best when cold stratified (5°C) for 21 days and R. monadelpha germinates best at 15°C in the dark. Seeds of R. diversiformis, R. flava, R. leipoldtii, R. minutiflora, R. monadelpha and R. sabulosa seem to all exhibit non-deep endogenous morphophysiological dormancy while seeds of R. camerooniana and R. rosea appear to have deep endogenous morphophysiological dormancy. The suitability of various explant types and media supplementations for culture initiation was examined for various species of Romulea. Both embryos and seedling hypocotyls can be used for R. flava, R. leipoldtii and R. minutiflora in vitro shoot culture initiation. R. sabulosa shoot cultures can only be initiated by using embryos as explants, because of the lack of seed germination in this species. Shoot cultures of R. diversiformis, R. camerooniana and R. rosea could not be initiated due to the lack of an in vitro explant shooting response. Shoot cultures can be initiated on media supplemented with 2.3 to 23.2 M kinetin for all species that showed an in vitro response. The most suitable concentration depended on the species used. Some cultures appeared embryogenic, but this was shown not to be the case. A medium supplemented with 2.5 M mTR is most suitable for R. sabulosa shoot multiplication. BA caused vitrification of shoots in all the experiments in which it was included and is not a suitable cytokinin for the micropropagation of these species. The effect of various physical and chemical parameters on in vitro corm formation and ex vitro acclimatization and growth was examined. Low temperature significantly increased corm formation in R. minutiflora and R. sabulosa. A two step corm formation protocol involving placing corms at either 10 or 20°C for a few months and then transferring these cultures to 15°C should be used for R. sabulosa. When paclobutrazol and ABA were added to the medium on which R. minutiflora shoots were placed, the shoots developed corms at 25°C. This temperature totally inhibits corm formation when these growth retardants are not present. BA inhibited corm formation in R. leipoldtii. Corms can be commercialized as propagation units for winter-rainfall areas with minimum temperatures below 5°C during winter. Although an incident of in vitro flowering was observed during these experiments, these results could not be repeated. Although none of the corms or plantlets planted ex vitro in the greenhouse survived, a small viability and an ex vitro acclimatization experiment shows that the corms produced in vitro are viable. One embryo of the attractive R. sabulosa, produces 2.1 ± 0.7 SE shoots after 2 months; subsequently placing these shoots on a medium supplemented with 2.5 μM mTR for a further 2 months multiplies this value by 5.5 ± 1.3 SE. Each of these shoots can then be induced to produce a corm after 6 months. This means that 1 embryo can produce about 12 corms after 10 months or about 65 corms after 12 months (if shoots are subcultured to medium supplemented with 2.5 μM mTR for another 2 months). Embryo rescue can enable wider crosses within this genus. These results can be used for further horticultural development of species in this genus and their hybrids and variants.
Thesis (Ph.D.)-University of KwaZulu-Natal, Pietermaritzburg, 2012.

The role of light and temperature on flowering of South African Watsonia species were evaluated to assess the potential for this genus as a commercial flower crop. Species were selected that represent different climatic regions of South Africa, with the aim of understanding how ecologically distinct species perform under cultivation. The four selected species were W. borbonica and W. tabularis (winter-rainfall area), W. angusta (shared rainfall) and W. pillansii (summer-rainfall area). In order to establish the optimum temperature required for flowering, plants were exposed for 12 weeks to three temperature regimes (12/7 °C, 21/15 °C and 29/21 °C) after attaining their first and/or second leaves. A temperature shift of 12/7 °C was used to assess if the plants had a vemalisation requirement. Controls were maintained under 25 % shade under natural conditions, with an average temperature of 24/7 °C. An elevated temperature of 29/21 °C was detrimental to plant growth. Moderate temperatures of 21/15 °C significantly (P<0.001) increased the height and the number of leaves produced per plant relative to the 12/7 °C treatment. These temperatures significantly (P<0.001) increased the total number of flowers produced per plant compared to low temperatures. However, flowering percentage and quality of flowers were reduced. A low temperature regime of 12/7 °C was efficient in satisfying vernalisation requirements and inducing flowering in four selected species. However, the total number of leaves produced per plant was signifcantly reduced. The summer-rainfall species, W pillansii, displayed a qualitative response to vernalisation, as no flowering was observed in non-vernalised plants. Two winter-rainfall species, W borbonica and W. tabularis, demonstrated a quantitative response to vernalisation. These species flowered at non-vernalising temperatures. W angusta behaved like the winter-rainfall species in terms of flowering. Overall, a vernalisaton treatment marginally reduced days to flower while flowering percentage was increased compared to other temperature regimes. However, there was no increase in the total number of flowers produced per plant. Low temperatures were not only effective for flower induction, but also for releasing corm dormancy, thus synchronising growth. Storing corms at either 4 or 10 QC resulted in 100 % sprouting within 4-6 weeks. The role of daylength in flowering of Watsonia plants was established by subjecting plants to long days (LO) of 16 h light and 8 h dark and to short days (SO) of 8 h light and 16 h dark. The number of leaves and flowering were significantly (P<0.01) promoted under the LO regime. However, there was strong temperature and daylength interaction in terms of flowering potential, as at low temperatures flowering was induced irrespective of daylength. In W. pillansii, flowering was obtained under both regimes (LO and SO) applied at the second leaf stage. Flowering in W. borbonica and W. tabularis was only observed under the LO regime at the second leaf stage. In both species, flowering was also obtained in SD-treated plants, provided treatment occurred after the formation of the third leaf. However, the total number and quality of flowers were reduced. To examine the effect of light intensity on flowering, plants at different developmental stages (first and/or second or beyond the third leaf stage) were exposed to photosynthetically active radiation (PAR) of 150 jJmol m-2s-1 or 39.5 jJmol m-2s-1 for 7 weeks. Exposure to low light intensity at either developmental stage compromised leaf quality. No flowering was observed following low light intensity treatment during the first to third leaf stages, even though plants were exposed to low temperature and LO regimes, both of which promoted flowering. Observation of the shoot apical meristem revealed that the second leaf stage was critical as the anatomical transition to flowering occurred at this level. When beyond the third leaf stage, low light intensity did not prevent flowering. However, the number of flowers produced per plant was reduced compared to plants maintained at 150 jJmol m-2s-1. Thus, light intensity played a role in both plant morphogenesis and flowering. LDs were effective in promoting vegetative growth whereas high light intensity and low temperature regimes played pivotal roles in flower induction. This makes them useful horticulture tools to produce desirable Watsonia plants for commercialisation.
Thesis (M.Sc.)-University of KwaZulu-Natal, Pietermaritzburg, 2006.

A artrite reumatoide (AR) é uma patologia crónica e incapacitante com grande impacto na qualidade de vida dos doentes. Caracteriza-se pela existência de inflamação nas articulações podendo mesmo levar à destruição do tecido articular. Afeta cerca de 0.5 a 1% da população mundial, verificando-se que tem maior incidência em mulheres do que em homens. Não é conhecida uma cura para a doença, pelo que é fundamental uma terapia mais eficaz para que se consiga controlar os sintomas e se possível retardar o avanço da doença. O monóxido de carbono (CO) em tempos visto apenas como um agente tóxico, atualmente é explorado pelas suas capacidades anti-inflamatórias, tendo-se desenvolvido moléculas libertadoras de CO, os CORMs, de modo a tirar partido dos seus efeitos terapêuticos. Nos últimos anos, a nanotecnologia suscitou um grande interesse na comunidade científica, uma vez que, permite criar sistemas de libertação controlada de fármacos tornando possível ultrapassar alguns dos problemas que se encontram nos fármacos ditos convencionais. Aliando o facto de a AR ser uma patologia inflamatória, do CO ter propriedades anti-inflamatórias e de ser possível criar sistemas de libertação controlada de fármacos, surge o presente trabalho. Pretende-se assim, desenvolver nanoagentes que permitam a libertação controlada de CO, para serem aplicados no tratamento da AR permitindo atenuar os seus sintomas e reduzir os efeitos colaterais, relativamente aos fármacos já existentes para esta patologia. Os objetivos englobam: o desenvolvimento de uma nova abordagem terapêutica para a AR, baseada no design e otimização de nanopartículas lipídicas incorporando CORMs, que possuam um índice terapêutico mais elevado que os tratamentos convencionais e com efeitos colaterais escassos.

Vários complexos metálicos têm sido explorados visando uma melhoria de atuais tratamentos das mais variadas doenças. Os compostos de vanádio apresentam uma capacidade de mimetizar a ação da insulina, insurgindo-se assim como uma aposta no tratamento da doença Diabetes mellitus. O monóxido de carbono (CO), outrora visto como apenas um agente tóxico, atualmente é explorado pelas suas capacidades anti-inflamatórias, tendo-se desenvolvido moléculas libertadoras de CO, os CORMs, de modo a tirar partido dos seus efeitos biológicos. Com vista a determinar a formação de aductos entre os complexos organometálicos de vanádio e diferentes proteínas e caracterizar o tipo de interação estabelecida recorreu-se à Cristalografia de raios-X. A estrutura cristalográfica do aducto VOIV(picolinato)2–tripsina bovina foi obtida a 1,09 Å de resolução, mostrando o complexo VOIV(pic)2 no centro ativo da enzima, coordenado à cadeia lateral da serina catalítica (Ser195), numa geometria octaédrica. Ensaios cinéticos preliminares realizados com o complexo VO(pic)2 foram efetuados para determinar potenciais efeitos inibitórios na atividade das enzimas lisozima da clara do ovo (HEWL) e tripsina. O potencial terapêutico dos CORMs associado à Nanotecnologia foi explorado para o [Ru(CO)3Cl(glicinato)] (CORM-3), visando desenvolver nanopartículas de ouro (AuNPs) funcionalizadas com o composto e diferentes proteínas. A toxicidade celular direta do CORM-3, bem como alguns potenciais efeitos biológicos de CORM-3–albumina de soro humano (BSA) e CORM-3–HEWL foram também avaliados, recorrendo a ensaios de viabilidade celular , riscados celulares, e RT-qPCR em várias linhas celulares tumorais e linha primária de fibroblastos humanos. Foi possível iniciar o desenvolvimento de nanosistemas de AuNPs@PEG@BSA e AuNPs@PEG@HEWL, seguindo-se futuramente a conjugação com o CORM-3. Não foi possível determinar o efeito anti-inflamatório do CORM-3 e seus conjugados através da análise dos genes de resposta por RT-qPCR em tempo real, no entanto determinou-se que o composto CORM-3 não possui toxicidade em linhas celulares tumorais, e que aparenta fomentar a regeneração celular em fibroblastos e células tumorais NINA, sendo este efeito potenciado aquando conjugação com BSA.