Thesis (Ph. D.)--Columbia University, 1921. / Biographical note. Bibliography: p. 46-47.

Thesis (Ph. D.)--Columbia University, 1927.

Die Fähigkeit Nukleinsäuren in lebenden Organismen nachzuweisen und zu visualisieren, ist entscheidend für das Verständnis zellulärer Prozesse. Die Forschungsgruppe von Prof. Dr. Oliver Seitz hat zu diesem Zweck fluorogene FIT-Hybridisierungssonden entwickelt, die die besondere Eigenschaft der Cyaninfarbstoffe Thiazolorange und Chinolinblau nutzen, stärker zu fluoreszieren, wenn sie in die beengte Umgebung eines Nukleinsäureduplex aus Sonde und spezifischer Zielsequenz eingebracht werden. Obwohl FIT-Sonden eine gute Fluoreszenzverstärkung und Spezifität aufweisen, wäre eine weitere Verbesserung ihrer Helligkeit und des Signal-Hintergrund-Verhältnisses wünschenswert. Um dies zu erreichen, wurde in dieser Arbeit ein Ansatz untersucht, bei dem FIT-Sonden mit zwei Fluorophoren desselben Typs ausgestattet werden (FIT2-Strategie). Dies sollte sowohl die Helligkeit der Sonde erhöhen, als auch die Fluoreszenz im Einzelstrang und bei Hybridisierung mit fehlgepaarter RNA durch eine Mischung aus kontaktvermittelter Fluo-reszenzlöschung und strahlungsfreiem Energietransfer verringern. Verschiedene Sonden-längen, Farbstoffabstände und -positionen wurden untersucht und es konnte bestätigt werden, dass FIT2-Sonden eine höhere Extinktionskoeffizienten, größere Fluoreszenzver-stärkung und eine bessere Selektivität aufweisen als einfach markierte Sonden. Außerdem behalten sie ihre Fähigkeit zur Unterscheidung von Match- und Mismatch-Zielen in visko-sem Zelllysat besser bei. Zudem konnte gezeigt werden, dass das FIT2-Konzept durch Hinzufügen eines hybridisie-rungsunempfindlichen Cyanin 7 Farbstoffs zu den Sonden dahingehend erweitert werden kann, dass eine ratiometrische Detektion der hybridisierten Sonde möglich ist und Hellig-keitsunterschiede aufgrund von lokalen Schwankungen der Sondenkonzentration bei der Bildgebung lebender Zellen korrigiert werden können. Mit diesen qFIT2-Sonden konnten Jurkat und CCRF-CEM T-Zellen in einem Mikroskopie-basierten Experiment unterschieden werden. / The ability to detect and visualize nucleic acids in living organisms is crucial for under-standing cellular processes. For this purpose, the research group of Prof. Dr. Oliver Seitz has introduced fluorogenic forced intercalation (FIT) hybridization probes, which exploit the unique property of the cyanine dyes thiazole orange and quinoline blue to exhibit increased fluorescence when placed in the constrained environment of a nucleic acid du-plex formed between probe and specific target sequence. Although FIT probes demon-strate solid fluorescence enhancement and specificity, further improvement of their abso-lute brightness and signal-to-background ratio would be desirable. To achieve this, the present thesis investigated an approach that equips FIT probes with two identical fluorophores (FIT2 strategy). This should on the one hand increase probe brightness, while simultaneously reducing fluorescence in the single strand and when hy-bridized to mismatched RNA, through a combination of contact-mediated quenching and non-radiant energy transfer. Various probe lengths, dye-dye distances and positions were screened, and it could be confirmed that FIT2 probes have higher extinction coefficients, greater fluorescence enhancement and better selectivity than their mono-dye counter-parts. Moreover, they better retain their ability to discriminate match and mismatch tar-gets in viscous cell lysate. Finally, it was demonstrated that the FIT2 concept can be extended by adding a hybridiza-tion-insensitive Cyanine 7 dye to the probes, allowing ratiometric detection of hybridized probe and correction of brightness differences due to local fluctuations in probe concen-tration during live-cell imaging. Using these qFIT2 probes, Jurkat and CCRF-CEM T-cells could be distinguished in a microscopy-based experiment.

Submitted by Fabio Sobreira Campos da Costa (fabio.sobreira@ufpe.br) on 2016-07-28T13:21:21Z No. of bitstreams: 2 license_rdf: 1232 bytes, checksum: 66e71c371cc565284e70f40736c94386 (MD5) TESE_Amanda Aliança_digital.pdf: 2689265 bytes, checksum: 0b25cf703fd666da539c3034a8c40823 (MD5) / Made available in DSpace on 2016-07-28T13:21:21Z (GMT). No. of bitstreams: 2 license_rdf: 1232 bytes, checksum: 66e71c371cc565284e70f40736c94386 (MD5) TESE_Amanda Aliança_digital.pdf: 2689265 bytes, checksum: 0b25cf703fd666da539c3034a8c40823 (MD5) Previous issue date: 2016-02-25 / CAPEs / A leishmaniose é uma doença infecciosa, não contagiosa de caráter zoonótico, causada por vários tipos de protozoários do gênero Leishmania. Em humanos, a doença apresenta três principais formas clínicas, dependendo das espécies envolvidas de Leishmania: a cutânea, mucocutânea e visceral (LV), sendo esta última a forma mais grave que quando não tratada pode evoluir para óbito em mais de 90% dos casos. O agente etiológico da LV compreende Leishmania donovani na Índia e leste da África; Leishmania infantum na China, Ásia central, Europa, África, América do Sul e Central. Calcula-se que a prevalência mundial de leishmaniose esteja em torno de 12 milhões de pessoas, acometendo 80 países e com uma estimativa de 400.000 novos casos da doença por ano. O tratamento atual para a leishmaniose é baseado na utilização de antimoniais pentavalentes como o estibogluconato de sódio (Pentostan®) e o antimoniato de meglumina (Glucantime®), este apresenta várias dificuldades como toxicidade, via de administração intravenosa e tempo prolongado de tratamento, que levam a sua descontinuação. Diante deste cenário, os compostos sintéticos são de grande importância na terapêutica por representarem 85% dos fármacos disponíveis. As ftalimidas e os tiazóis são importantes classes de compostos sintéticos e apresentam amplo espectro de atividades biológicas, como ansiolítica, anti-Parkinson, anti-Alzheimer, analgésica, anti-inflamatória, imunomoduladora e antibacteriana. O presente trabalho teve como objetivo avaliar a atividade in vitro frente a L. infantum e a células de mamíferos de uma série de 15 derivados ftalimido-tiazóis. Os compostos foram avaliados frente a formas promastigotas e amastigotas de L. infantum, quanto à citotoxicidade frente a células vero, macrófagos J774 e macrófagos peritoneais, a produção de óxido nítrico e alterações sobre alvos intracelulares do parasito. Os resultados mostram que os compostos oriundos da hibridação da ftalimida com 1,3 tiazol apresentaram atividade leishmanicida contra formas promastigota de L. infantum, baixa citotoxicidade às células de mamíferos testadas e aumentaram a produção de ON em relação às células controles nos macrófagos não infectados. Os compostos 2j e 2m se mostraram como os mais potentes contra as formas promastigotas da série testada, estes foram testados frente as formas amastigotas, os compostos reduziram a sobrevivência de amastigotas intracelulares e apresentaram baixa citotoxicidade para os macrófagos peritoneais. Formas promastigotas tratadas com esses compostos apresentaram alterações ultraestruturais como: encolhimento do corpo celular, perda da integridade da membrana celular, vacuolização do citoplasma, perfis de membranas circundando organelas e inchaço da mitocôndria. Quando avaliada a marcação pelo iodeto de propídio e pela rodamina 123, formas promastigotas tratadas com os compostos 2j e 2m apresentaram aumento no número de células marcadas com o iodeto de propídio e induziram alterações significativas no potencial de membrana mitocondrial. Dessa maneira, os compostos ftalimido-tiazóis apresentam atividades leishmanicida e devem formar a base para futuros estudos experimentais. / Leishmaniasis is an infectious disease, non-contagious with zoonotic characteristic, caused by several types of protozoa of the genus Leishmania. In humans, the disease has three main clinical forms, depending on the species of Leishmania involved: cutaneous, mucocutaneous and visceral, the latter being the more severe if left untreated can lead to death in more than 90% of cases. The etiologic agent of LV comprises Leishmania donovani in India and East Africa; Leishmania infantum in China, Central Asia, Europe, Africa, South and Central America. It is estimated that the worldwide prevalence of leishmaniasis is around 12 million people in 80 countries and 400,000 new cases per year. The current treatment for leishmaniasis is based on the use of pentavalent antimonials such as stibogluconate sodium (Pentostan®) and meglumine antimonate (Glucantime®), this presents several problems such as difficulties such as toxicity, intravenous route of administration and prolonged treatment, which leads to its discontinuation of the treatment. The phthalimides and thiazoles are important classes of synthetic compounds and have a broad spectrum of biological activities such as anxiolytic, anti-Parkinsons, anti-Alzheimer, analgesic, antiinflammatory, immunomodulating and antibacterial. This study aimed to evaluate the in vitro activity against L. infantum and mammalian cells in a series of 15-phthalimido thiazoles derivatives. The present work report the in vitro activity of a phthalimido-thiazoles derivatives series. The activities were evaluated against promastigotes and amastigotes of Leishmania infantum, front of cytotoxicity to vero cells, J774 macrophages and peritoneal macrophages, the effect on production of nitric oxide (NO) and changes of intracellular targets on the parasite. The results show that the compounds arising from the hybridization of phthalimido 1, 3 thiazole showed leishmanicidal activity against promastigotes forms of L. infantum, low cytotoxicity to mammalian cells and increased the NO production compared to control cells in uninfected macrophages. The compounds 2j and 2m are shown as the most potent against promastigotes of the test series, these were tested against amastigote forms, the compounds reduced the survival of intracellular amastigotes and showed low cytotoxicity peritoneal macrophages. The promastigotes treated with these compounds exhibited ultrastructural changes such as cell body shrinkage, loss of cell membrane integrity, vacuolization of the cytoplasm membranes surrounding organelles profiles and swelling of mitochondria. The compounds 2j and 2m were the most potent of the series tested and the parasites treated with these compounds showed ultrastructural changes such as cell body shrinkage, loss of cellular membrane integrity, vacuolization of cytoplasm, membrane profiles surrounding organelles and swelling of mitochondria. When assessed by propidium iodide tag and the rhodamine 123 promastigotes treated with the compounds 2j and 2m showed an increase in the number of cells stained with propidium iodide and induced significant changes in mitochondrial membrane potential. Thus, the phthalimido-thiazoles compounds have leishmanicide activities and should form the basis for future experimental studies.

Les statines sont des inhibiteurs sélectifs de la 3-hydroxy-3-méthylglutaryl-coenzyme A réductase, utilisées pour diminuer la biosynthèse du cholestérol. Ces molécules possèdent en plus de leur capacité à réduire le cholestérol des effets pléiotropiques comme les propriétés anti-inflammatoires et anti-oxydantes. Les cucurbitacines sont des triterpènes dérivés des plantes, ayant des propriétés biologiques diverses comme les effets anti-inflammatoires et anticancéreux, associées toutefois à une toxicité élevée. Les dérivés de thiazole sont des molécules contenant un noyau hétérocyclique formé de trois atomes de carbones, un atome de sulfure et un atome d'azote, disposant des effets anti-inflammatoires importantes. Les cyclooxygénases et les hème-oxygénases jouent un rôle dans l'inflammation et le stress oxydatif et sont les cibles des statines et des cucurbitacines in vitro. Les dérivés de thiazole peuvent inhiber plutôt l'activité enzymatique des cyclooxygénases. Le but de ma thèse est d'étudier les effets de ces 3 molécules in vivo et d'analyser si possible les mécanismes impliqués, comme par exemple pour les statines. Pour cela, nous avons eu recours chez les souris C57BL/6 au modèle d'inflammation de la poche à air-stérile injecté par le zymosan.Nous avons d'abord montré que le traitement des souris avec l'atorvastatine pendant 10 jours a réduit la migration des cellules dans l'exsudat de la poche à air ainsi que l'expression de gènes des marqueurs pro-inflammatoires tels que les cytokines, les chimiokines, la cyclooxygénase-2 et la nitric oxide synthase -II. Le taux de la prostaglandine E2 et de l'interleukine-6 a été également réduit. L'inhibition de l'hème-oxygénase-1 par son inhibiteur sélectif, tin protoporphyrine, a partiellement réduit l'effet inhibiteur de l'atorvastatine sur la migration des cellules et sur certaines cytokines suggérant un rôle important de la l'hème-oxygénase-1 dans les propriétés anti-inflammatoires in vivo.En parallèle, nous avons utilisé ce modèle animal tester l'effet de la cucurbitacine E sur l'inflammation et évaluer le rôle d'encapsulation in vivo dans des liposomes à base de phosphatidylcholine. Nous avons démontré que la cucurbitacine E libre (12.5 μg/poche ou 25 μg/poche) a tendance à diminuer l'interleukine-6, l'hème oxygénase-1 et la cyclooxygénase-2 alors que la cucurbitacine E encapsulée (12.5 μg/poche) a significativement réduit la prostaglandine E2, l'interleukine-6 et le taux de nitrite sans affecter le niveau d'ARNm de la cyclooxygénase-2 et l'hème oxygénase-1. Nos résultats suggèrent que l'incorporation de la cucurbitacine E dans des liposomes améliore son effet anti-inflammatoire et réduit sa cytotoxicité.Finalement, deux dérivés de thiazole qui diffèrent dans leur structure par la présence d'un groupement butyle- ou benzyle- sur leur chaîne latérale, ont été explorés dans ce modèle. Nous avons montré que le dérivé de thiazole contenant le groupe benzyle est sélectif de la cyclooxygénase-2 dans les macrophages et le modèle in vivo chez la souris.En résumé, mon travail de thèse met en évidence in vivo les effets anti-inflammatoires des statines et le rôle de l'hème oxygénase-1, et permet en plus la caractérisation des effets anti-inflammatoires des cucurbitacines et des dérivés de thiazole. L'ensemble de ces résultats renforcent les effets anti-inflammatoires de ces substances et démontre in vivo leur effet et les suggèrent comme molécules anti-inflammatoires. / Statins are selective competitive inhibitors of 3-hydroxy-3-methylglutaryl coenzyme A reductase used to lower cholesterol biosynthesis and have multiple pleiotropic effects beyond lowering cholesterol such as anti-inflammatory and anti-oxidant properties. Cucurbitacins are triterpenoid derived from plants and exhibit potential anti-inflammatory and anticancer properties though they have a high cytotoxicity. Thiazole derivatives are molecules containing heterocyclic ring with three carbons, one sulfur and one nitrogen atom, with important anti-inflammatory activities. Cyclooxygenases and heme-oxygenases play a role in inflammation and oxidative stress and are targets for statins or cucurbitacins in vitro. Thiazole derivatives are potential inhibitors of cyclooxygenases. The aim of my thesis is to investigate the anti-inflammatory effect of these three compounds in vivo and attempt to analyze the mechanisms involved, mainly for statins. Thus we set up an animal model of inflammation in C57BL/6 corresponding to the zymosan -injected dorsal sterile air pouch.First we have shown that treatment of mice with atorvastatin for 10 days reduced cell migration in the exudate of the air pouch as well as the expression of inflammatory markers such as cytokines, chemokines, cyclooxygenase-2 and nitric oxide synthase -II. The synthesis of prostaglandin E2 and interleukin-6 was also reduced. Inhibition of heme-oxygenase-1 by the selective inhibitor tin protoporphyrin partially diminished the inhibitory effect of statins on cell migration and some cytokines suggesting a significant role of HO-1 in the anti-inflammatory properties for atorvastatin in vivo.For cucurbitacins, we used the same animal model to investigate the effect of this substance in vivo and further assess the beneficial effect of encapsulating cucurbitacin in phosphatidylcholine-liposomes. Free cucurbitacin E (12.5 μg/pouch or 25 μg/pouch) tended to increase interleukin-6, decrease heme oxygenase-1 and cyclooxygenase-2 whereas cucurbitacin E loaded liposomes (12.5 μg/pouch) significantly reduced prostaglandin E2, interleukin-6 and nitrite without affecting cyclooxygenase-2 and heme oxygenase-1 mRNA levels. We demonstrated that the incorporation of Cuc E into liposomes enhances its anti-inflammatory effect and reduces its cytotoxicity.Finally, we used the dorsal air pouch model to investigate the anti-inflammatory effect of two thiazole derivatives that differ in their side chain by the presence of butyl or benzyl group. In addition to analyzing their effect on cyclooxygenase activity in human blood platelets, on recombinant COX-1 and in culture macrophage cell lines, we demonstrated their capacity to block cyclooxygenase-dependent prostaglandin synthesis in the lumen of the air pouch.In summary, my thesis presents in vivo evidence for the anti-inflammatory effects of atorvastatin dependent on HO-1 activity. My studies characterized the anti-inflammatory effects of cucurbitacins and thiazole derivatives. All these results support the anti-inflammatory effects of these substances and suggested them as potential anti-inflammatory substances.

Bacillus amyloliquefaciens FZB42 ist ein grampositives Bodenbakterium. Es kann in der Rhizosphäre das Wachstum von Pflanzen fördern und durch die Produktion von Sekundärmetaboliten phytopathogene Organismen hemmen. Aus der Genomanalyse und den dazugehörigen Arbeiten war bekannt, dass Bacillus amyloliquefaciens FZB42 nicht-ribosomal je drei antimikrobielle Polyketide und Lipopeptide herstellt, sowie zwei Siderophore und das Dipeptid Bacilysin. Für Bacillus typische Lantibiotika oder große Bacteriocine wurden nicht gefunden. In dieser Arbeit wird erstmalig gezeigt, dass Bacillus amyloliquefaciens FZB42 auf ribosomale Weise antibakterielle Peptide herstellt. Zwei bisher unbekannte Bacteriocine, Amylocyclicin A und Plantazolicin, und deren dazugehörigen Gencluster konnten identifiziert und charakterisiert werden. Amylocyclicin A ist ein unmodifiziertes Peptid, dessen N- und C-Terminus kovalent verbunden sind. Es wurde der Gruppe I der zirkulären Bacteriocine zugeordnet, dessen Mitglieder sich durch schwache Homologie untereinander, aber durch wahrscheinlich ähnliche 3D-Strukturen auszeichnen. Die Masse beträgt 6381 Da und die Substanz ist stark aktiv gegen grampositive Bakterien. Das Biosynthesecluster umfasst sechs Gene für die Synthese, den Export, die Zyklisierung und die Immunität. Plantazolicin ist ein hydrophobes, stark modifiziertes Peptid aus der TOMM-Gruppe, einer Gruppe aus Microcin B17-ähnlichen Peptiden, die nach neueren Erkenntnissen verbreiteter ist, als bisher bekannt. Plantazolicin ist schwach aktiv gegen grampositive Bakterien und besitzt die Masse 1335 Da. Das Biosynthesecluster umfasst zwölf Gene, mit allen nötigen Genen für Synthese, Modifikation, Regulation, Immunität und Export. / Bacillus amyloliquefaciens FZB42 is a Gram-positive, plant-associated bacterium, which stimulates plant growth and produces secondary metabolites that suppress soil-borne plant pathogens. Five gene clusters direct the non-ribosomal synthesis of the cyclic lipopeptides surfactin, bacillomycin, fengycin, an unknown peptide and the iron-siderophore bacillibactin. Three gene clusters direct the non-ribosomal synthesis of the antibacterial acting polyketides macrolactin, bacillaene and difficidin; in addition to the non-ribosomal synthesis of the antibacterial dipeptide bacilysin. Genes involved in ribosome-dependent synthesis of lantibiotics and other peptides are scarce. Only two incomplete gene clusters directing immunity against mersacidin and subtilin were found. In this work two ribosomally synthesized antibacterial peptides, amylocyclicin A and plantazolicin, and their corresponding gene clusters were identified. Amylocyclicin A is a circular peptide with a mass of 6381 Da and strong activity against Gram-positive bacteria. Six genes are responsible for the synthesis, maturation, export and immunity of this peptide belonging to group I of circular bacteriocins. Plantazolicin is a strongly modified hydrophobic peptide bearing a molecular mass of 1,335 Da and displaying antibacterial activity toward closely related Gram-positive bacteria. Essential modification contains the incorporation of azole heterocycles, which derive from Cys, Ser, and Thr residues of the precursor peptide and addition of two methyl groups. Twelve genes are responsible for synthesis, modification, export and immunity of this peptide belonging to the TOMM group of thiazol/oxazol modified microcins.

[pt] As doenças negligenciadas são causadas por agentes infecciosos ou parasitas, gerando altas taxas de morbidade e mortalidade em todo o mundo. Os investimentos dos países desenvolvidos em pesquisa, produção de medicamentos e controle dessas enfermidades são muito reduzidos, pois elas afetam, majoritariamente, as populações com baixo Índice de Desenvolvimento Humano. Uma dessas enfermidades é a Doença de Chagas, causada pelo Trypanosoma cruzi. Apresenta-se na forma de uma infecção parasitária crônica e altamente debilitante, acometendo mais de 6 milhões de pessoas na América Latina e este número pode ter um incremento, de aproximadamente 25 por cento nos próximos anos. O fármaco referência para o tratamento da Doença de Chagas disponível no Brasil é o benznidazol, extremamente eficaz na fase aguda da doença, mas sua eficácia na fase crônica é limitada, o que torna essencial a busca por novos compostos que atuem também nesta fase. No presente trabalho, foram descritas rotas sintéticas para uma série de 12 derivados de tiazol, destes 8 são inéditos. Posteriormente, 10 deles foram avaliados quanto a sua atividade tripanocida e toxicidade em modelos in vitro. Os compostos possuindo o anel tiazólico foram estruturalmente planejados, a partir do levantamento bibliográfico, onde verificou-se que certos compostos com um centro tiazólico apresentam propriedade tripanocida. Nas sínteses, a 4-bromoacetofenona (55) foi utilizada como material de partida para a obtenção dos intermediários tiazólicos 4-(4-bromofenil)-2- metiltiazol (56) e 4-(4-bromofenil)-2-aminotiazol (57). A partir desses, diversas modificações foram feitas para se chegar às moléculas alvo. A elucidação estrutural dos compostos foi realizada por RMN de 1H e RMN de 13C. A avaliação preliminar da atividade tripanocida foi realizada na forma amastigota intracelular da Cepa Tulahuen e a toxicidade aguda (LC50) foi feita in vitro em células hospedeiras de mamíferos (linhagem celular L929). Dentre os compostos sintetizados, a maioria apresentou baixa toxicidade, com valores de LC50 maiores que 400 (micro)mol.L-1. O composto tiazólico piridil-substituído 59c apresentou os melhores resultados em termos de atividade tripanocida, tendo reduzido 76 por cento da infecção. em células hospedeiras em concentração de 20 (micro)mol.L-1 Esse composto servirá como base para otimizações estruturais visando a melhoria da atividade tripanocida. / [en] Neglected diseases are mainly caused by infectious agents or parasites, generating high morbidity and mortality rates worldwide. The investments from developed countries in research, production of drugs/medicines and control of these diseases are small, since they mainly affect populations with low Human Development Index. One of these illnesses is Chagas disease, caused by Trypanosoma cruzi. It presents as a chronic and highly debilitating parasitic infection, affecting more than 6 million people only in Latin America with a perspective of increasing approximately 25 percent in few years. The reference drug for the treatment of Chagas disease in Brazil is benznidazole, which is extremely effective in the acute phase of the disease, but its efficacy in the chronic phase is limited, which makes essential the search for new compounds that also act in this phase. In the present work, synthetic routes were developed for a series of 12 thiazole derivatives, from which eight have been described for the first time. These molecules had their preliminary trypanocidal activity and toxicity evaluated in in vitro models. The compounds with the thiazole ring were structurally planned, based on the bibliographic review, where it was found that certain compounds with this heterocycle display trypanocidal properties. In the synthesis, 1-(4- bromophenyl)ethenone (54) was used as a starting material to obtain the thiazole intermediates 4-(4-bromophenyl)-2-methylthiazole (56) and 4-(4-bromophenyl)thiazol-2- amine (57). Several modifications were performed on these intermediates to produce the target molecules. The structural elucidation of the compounds was performed by 1H NMR and 13C NMR. Preliminary evaluation of trypanocidal activity was performed in the amastigotes intracellular form of the Tulahuen strain and the acute toxicity (LC50) was tested in vitro in mammalian host cells (cell line L929). Among the synthesized compounds, most of them presented low toxicity, with LC50 values greater than 400 (micro)mol/mol.L-1. The thiazole compound pyridil-substituted 59c showed the best results in terms of trypanocidal activity, reducing 76 percent of the infection in host cells in a concentration of 20 (micro)mol/mol.L-1. This thia\ole derivative will be used as a lead compound for structural optimizations aiming at improving trypanocidal activity.