Résumé Les hémoglobines tronquées (Hbtrs) sont retrouvées chez plusieurs organismes et leurs fonctions sont encore inconnues pour la plupart. Les Hbtrs du pathogène de plante Agrobacterium tumefaciens C58 ont été inactivées pour vérifier leurs implications dans la production de tumeur chez les plantes. Les expériences ont montré que les souches mutantes pour les Hbtrs AtuHb2 et AtuHb3 étaient capables d'induire la production de tumeur chez la plante Kalanchoe daigremontiana. La protéine recombinante AtuHb2 a été caractérisée par spectroscopie d'absorption et de résonance Raman. L'analyse, par spectroscopie d'absorption, montre que la protéine est hexacoordonnée dans la forme ferrique et ferreuse, qu'il est possible pour AtuHb2 de former des complexes stables avec les ligands CO, CN- et NO, mais pas avec l'O2 et que le complexe formé entre la forme ferreuse et le NO est pentacoordonné. L'analyse par spectroscopie de résonance Raman a montré que le ligand CO est peu stabilisé

Submitted by Automa??o e Estat?stica (sst@bczm.ufrn.br) on 2018-01-15T22:12:26Z No. of bitstreams: 1 EdilaneRodriguesDantasDeAraujo_DISSERT.pdf: 4618681 bytes, checksum: 5ed4bc7a898bcd1ef47ef38661580242 (MD5) / Approved for entry into archive by Arlan Eloi Leite Silva (eloihistoriador@yahoo.com.br) on 2018-01-18T13:59:07Z (GMT) No. of bitstreams: 1 EdilaneRodriguesDantasDeAraujo_DISSERT.pdf: 4618681 bytes, checksum: 5ed4bc7a898bcd1ef47ef38661580242 (MD5) / Made available in DSpace on 2018-01-18T13:59:07Z (GMT). No. of bitstreams: 1 EdilaneRodriguesDantasDeAraujo_DISSERT.pdf: 4618681 bytes, checksum: 5ed4bc7a898bcd1ef47ef38661580242 (MD5) Previous issue date: 2017-06-30 / Coordena??o de Aperfei?oamento de Pessoal de N?vel Superior (CAPES) / Kalanchoe brasiliensis e Kalanchoe pinnata (Crassulaceae), conhecidas como ?sai?o? e ?coirama?, t?m amplo uso popular no tratamento de ?lceras p?pticas e inflama??es cut?neas. Vale destacar que K. pinnata est? presente na Rela??o Nacional de Plantas de Interesse do Sistema ?nico de Sa?de - RENISUS (2009). Dentro deste contexto, o objetivo do presente estudo foi caracterizar os marcadores qu?micos nos sucos das folhas das duas esp?cies e avaliar as atividades gastroprotetora e anti-inflamat?ria t?pica. Foi realizada caracteriza??o fitoqu?mica por Cromatografia em Camada Delgada (CCD) e Cromatografia L?quida de Ultra Efici?ncia Acoplada a Espectr?metro de Massas (CLUE-EM). A atividade gastroprotetora foi avaliada nos modelos de ?lcera ?guda induzida por etanol e indometacina, j? a secre??o g?strica foi avaliada no modelo de ligadura do piloro, em ratos Wistar. Foi realizado o pr?- tratamento com os sucos nas doses de 125, 250 e 500 mg/kg e a ranitidina (50 mg/kg) por via oral. A atividade anti-inflamat?ria t?pica foi avaliada no modelo de edema de pata induzido por carragenina e edema de orelha induzido por ?leo de crot?n em camundongos Swiss, utilizando formula??es na forma de gel contendo os sucos em diferentes concentra??es (1,25%, 2,5% e 5%) e como f?rmaco padr?o a dexametasona (1mg/g), todos administrados por via t?pica imediatamente ap?s a indu??o. A an?lise por CCD revelou a presen?a de manchas caracter?sticas de flavonoides nos sucos das duas esp?cies, ap?s revela??o com o Reagente Natural A, sendo observado que as duas esp?cies t?m perfil flavono?dico diferente. Na an?lise por CLUE-EM K. brasiliensis apresentou flavonoides glicosilados derivados principalmente da patuletina, enquanto que K. pinnata apresentou flavonoides glicosilados derivados principalmente da quercetina. O pr?-tratamento com o suco das folhas de K. brasiliensis nas doses de 125 mg/kg (P<0,01), 250 mg/kg e 500 mg/kg (P<0,001) e K. pinnata nas doses de 125 mg/kg (P<0,01), 250 mg/k e 500 mg/kg (P<0,001) reduziram significativamente as les?es em compara??o ao controle positivo no modelo de indu??o por etanol. No modelo de indu??o por indometacina o suco das folhas de K. brasiliensis apresentou resultado significativo nas doses de 250 (P<0,05) e 500 mg/kg (P<0,01) e K. pinnata nas doses de 250 e 500 mg/kg (P<0,001). A redu??o das les?es foi acompanhada de aumento do conte?do total de glutationa e redu??o dos n?veis de malondialde?do. Al?m disso, houve redu??o dos n?veis de mieloperoxidase, IL-1? e TNF-?. Tamb?m foi observado efeito citoprotetor na avalia??o histol?gica com H&E e manuten??o da produ??o de muco com PAS, al?m da redu??o da express?o de iNOS e NF-?B p65 e aumento da express?o de ZO-1 por imunohistoqu?mica. Os sucos das folhas das duas esp?cies n?o alteraram a acidez, o pH e o volume do suco g?strico. No modelo de edema de orelha, as formula??es contendo as tr?s concentra??es do suco das folhas de K. brasiliensis reduziram significativamente o edema quando comparadas ao grupo placebo (1,25% P<0,05; 2,5% P<0,01 e 5% P<0,01). Entretanto, apenas a formula??o contendo o suco das folhas de K. pinnata na concentra??o de 5% apresentou resultado significativo (P<0,01). No modelo de edema de pata, as formula??es contendo o suco das folhas de K. brasiliensis nas concentra??es de 1,25 e 2,5% reduziram significativamente (P<0,05) o edema no tempo 4 h. A formula??o na concentra??o de 5% reduziu significativamente o edema nos tempos 1 h (P<0,001), 2 h, 3 h e 4 h (P<0,01). Em rela??o as formula??es contendo o suco das folhas de K. pinnata, a concentra??o de 1,25% reduziu significativamente o edema no tempo 1h (P<0,01) e 2h (P<0,05), na concentra??o de 5% reduziu significativamente no tempo 1h (P<0,05). A diminui??o do edema foi acompanhada da redu??o de miloperoxidase. Concluiu-se que os sucos das duas esp?cies apresentaram atividade gastroprotetora e anti-inflamat?ria t?pica em modelos in vivo, resultados que justificam a utiliza??o popular das esp?cies. / Kalanchoe brasiliensis and Kalanchoe pinnata (Crassulaceae), known as "sai?o" and "coirama", have wide popular use in the treatment of peptic ulcers and cutaneous inflammations. It is worth mentioning that K. pinnata is present in the National List of Plants of Interest of the Unified Health System - RENISUS (2009). Within this context, the objective of the present study was to characterize the chemical markers in the leaf juices of both species and to evaluate the gastroprotective and topical anti-inflammatory activities. Phytochemical characterization was performed by Thin Layer Chromatography (TLC) and Ultra High Performance Liquid Chromatography coupled to Mass Spectrometer (UHPLC- MS). Gastroprotective activity was evaluated in ethanol and indomethacin induced acute ulcer models, whereas gastric secretion was evaluated in the pylorus ligature model in Wistar rats. Pre-treatment was performed with the juices at the doses of 125, 250 and 500 mg/kg and ranitidine (50 mg/kg) orally. The topical anti-inflammatory activity was evaluated in the carrageenan induced paw edema model and croton oil-induced ear edema in Swiss mice using gel formulations containing the juices at different concentrations (1,25%, 2,5% and 5%) and as the standard drug dexamethasone (1mg/g), all administered topically immediately after induction. The TLC analysis revealed the presence of flavonoid stains in the juices of both species after revelation with the Natural Reagent A, being observed that the two species have different flavonoid profiles. In the analysis by UHPLC-MS the K. brasiliensis leaf juice showed glycosylated flavonoids derived mainly from patuletin, while that of K. pinnata presented glycosylated flavonoids derived mainly from quercetin. The pre-treatment with the K. brasiliensis leaf juice at doses of 125 mg/kg (P<0,01), 250 mg/kg and 500 mg/kg (P<0,001) and K. pinnata at doses of 125 mg/kg (P<0,01), 250 mg/kg and 500 mg/kg (P<0,001) significantly reduced the lesions compared to the positive control in the ethanol induction model. In the indomethacin induction model, the K. brasiliensis leaf juice showed significant results at doses of 250 mg/kg (P<0,05) and 500 mg/kg (P<0,01) and K. pinnata at doses of 250 and 500 mg/kg (P<0,001). Reduction of lesions was accompanied by an increase in total glutathione content and reduction of malondialdehyde levels. In addition, levels of myeloperoxidase, IL-1? and TNF-? were reduced. Cytoprotective effect was also observed in histological evaluation with H&E and maintenance of mucus production with PAS, as well as reduction of iNOS and NF-?B p65 expression and increased expression of ZO-1 by immunohistochemistry. Leaf juices from both species did not change the acidity, pH and volume of the gastric juice. In the ear edema model, the formulations containing the three concentrations of the K. brasiliensis leaf juice significantly reduced the edema when compared to the placebo group (1,25% P<0,05, 2,5% P<0,01 and 5% P<0,01). However, only the formulation containing the juice of the K. pinnata leaf juice at 5% concentration showed a significant result (P<0,01). In the paw edema model, the formulations containing the K. brasiliensis leaf juice at concentrations of 1,25 and 2,5% significantly reduced (P<0,05) the edema in the time 4 h. The formulation at 5% concentration significantly reduced edema at 1 h (P <0,001), 2 h, 3 h and 4 h (P <0,01). Regarding the formulations containing the K. pinnata leaf juice, the concentration of 1,25% significantly reduced the edema in the time 1 h (P<0,01) and 2 h (P<0,05), in the concentration of 5% significantly reduced in time 1 h (P<0,05). The decrease in edema was followed by reduction of miloperoxidase. It was concluded that the juices of both species presented gastroprotective and topical anti-inflammatory activity in vivo models, results that justify the popular use of the species.

Made available in DSpace on 2014-12-17T14:16:30Z (GMT). No. of bitstreams: 1 AnneCOC_DISSERT.pdf: 1977864 bytes, checksum: 512fa415d2a3e86643a5d0929e22bccf (MD5) Previous issue date: 2012-06-05 / Kalanchoe brasiliensis Cambess (Crassulaceae), commonly known as sai?o , coirama branca , folha grossa , is originally from Brazil and commonly found in S?o Paulo to Bahia, mainly in the coastal zone. Regarding of biological activities, most preclinical studies were found in the literature, mainly about the anti-inflammatory activity of extracts obtained from leaves and / or aerial parts of K. brasiliensis. As regards the chemical constitution, it has been reported mainly the presence of flavonoids in the leaves of the species, but until this moment did not knows which are the active compounds. Although it is a species widely used in traditional medicine in Brazil, there is no monograph about the quality parameters of the plant drug. In this context, this study aims to characterize and quantify the chemical markers of hydroethanolic extract (HE) from the leaves of K. brasiliensis, which can be used in quality control of plant drug and derivatives obtained from this species. The methodology was divided into two parts: i. Phytochemical study: to fractionate, isolate and characterizate of the chemical (s) marker (s) of the HE from the leaves of K. brasiliensis; ii. To Developed validate of analytical method by High Performance Liquid Chromatography (HPLC)-diode array detector (DAD) to quantify the chemical (s) marker (s) of the EH. i. The EH 50% was prepared by turbo extraction method. It was then submitted to liquid-liquid partition, obtaining dichloromethane, n-butanol and ethyl acetate (AcOEt) fractions. The AcOEt fraction was selected to continue the fractionation process, because it has a chemical profile rich in flavonoids. The acOEt fraction was submitted to column chromatography using different systems for obtaining the compound Kb1. To identify this compound, it was submitted to UV analysis ii. For quantitative analysis, the EH was analyzed by HPLC, using different methods. After selecting the most appropriate method, which showed satisfactory resolution and symmetrical peaks, it was validated according to parameters in the RE 899/2003. As result, it was obtained from the AcOEt fraction the compound Kb1 (2.7 mg). Until this moment, the basic nucleus was characterized by UV analysis using shift reagents. The partial chemical structure of the compound Kb1 was identified as a flavonol, containing hydroxyls in 3 , 4 position (ring A), 5 and 7 free (ring B) and a replacement of the C3 hydroxyl by a sugar. As the analysis were performed in the HPLC coupled to a DAD, we observed that the UV spectrum of the major peaks of EH from K. brasiliensis shown similar UV spectrum. According to the literature, it has been reported the presence of patuletin glycosydes derivatives in the leaves of this species. Therefore, it is suggested that the compound Kb1 is glycosylated patuletin derivative. Probably the sugar (s) unit(s) are linked in the C3 in the C ring. . Regarding the development of HPLC analytical method, the system used consists of phase A: water: formic acid (99,7:0,3, v / v) and phase B: methanol: formic acid (99,7:0,3, v / v), elution gradient of 40% B - 58% B in 50 minutes, ccolumn (Hichrom ?) C18 (250x4, 0 mm, 5 ?m), flow rate 0.8 mL / min, UV detection at 370 nm, temperature 25 ? C. In the analysis performed with the co-injection of thecompound Kb1 + HE of K. brasiliensis was observed that it is one of the major compounds with a retention time of 12.47 minutes and had a content of 15.3% in EH of leaves from K. brasiliensis. The method proved to be linear, precise, accurate and reproducible. According to these results, it was observed that compound Kb1 can be used as a chemical marker of EH from leaves of K. brasiliensis, to assist in quality control of drug plant and its derivatives / Kalanchoe brasiliensis Cambess (Crassulaceae), conhecida popularmente como sai?o, coirama branca e folha grossa, ? origin?ria do Brasil e comumente encontrada de S?o Paulo at? a Bahia, principalmente na zona litor?nea. Quanto ? atividade microbiol?gica, estudos pr?-cl?nicos foram realizados no sentido de investigar a atividade antiinflamat?ria, para os extratos obtidos a partir folhas e/ou partes ?reas de K. brasiliensis. Em rela??o ? constitui??o qu?mica, h? relatos principalmente da presen?a de flavonoides nas folhas da esp?cie. Embora seja uma esp?cie utilizada amplamente na medicina tradicional no Brasil, n?o h? monografia que especifique os par?metros de qualidade da droga vegetal. Dentro deste contexto, o presente trabalho tem como objetivo caracterizar e a quantificar os marcadores qu?micos do extrato hidroetan?lico (EH) das folhas de K. brasiliensis, que possam ser utilizados no controle de qualidade de derivados obtidos a partir dessa esp?cie. A metodologia do estudo foi dividida em duas partes: i. Estudo fitoqu?mico: fracionamento, isolamento e caracteriza??o de marcador(es) qu?mico(s) do extrato hidroetan?lico das folhas de K. brasiliensis; ii.Desenvolvimento e valida??o de metodologia anal?tica por Cromatografia L?quida de Alta Efici?ncia (CLAE) para quantificar (o)s marcador(es) qu?mico(s) do EH. i. O EH 50% foi obtido pelo m?todo de turboextra??o. Em seguida foi submetido a uma parti??o l?quido-l?quido, obtendo-se as fra??es diclorometano, n-butanol e acetato de etila (FAcOEt). A fra??o acetato de etila foi selecionada para dar continuidade ao fracionamento, e por apresentar um perfil qu?mico rico em flavonoides, uma vez que esses metab?litos parecem estar relacionados ? atividade farmacol?gica. ii. Para a an?lise quantitativa, o EH foi analisado por CLAE, utilizando diferentes m?todos. Ap?s selecionado o m?todo mais adequado, que apresentou picos sim?tricos e resolu??o satisfat?ria, foi realizada a valida??o segundo par?metros da RE No 899/2003. Como resultados, foi obtido a partir da fra??o acetato de etila o composto Kb1 (2,7mg). Neste composto, foi caracterizado o n?cleo b?sico flavono?dico por an?lise por UV utilizando reagentes de deslocamento. O composto Kb1 foi identificado como um flavonol, com hidroxilas em 3 , 4 (anel B) e 5 e 7 livres (anel A), al?m de uma hidroxila substitu?da em C3. Como foi utilizado a CLAE acoplada a um detector de arranjo de diodos, foi poss?vel observar que os espectros UV dos picos majorit?rios do EH de K. brasiliensis s?o similares. Em estudos anteriores foi descrita a presen?a de derivados glicosilados da patuletina nas folhas dessa esp?cie. Portanto, sugere-se que Kb1 seja um derivado glicosilado na posi??o C-3 da patuletina. Quanto ao desenvolvimento anal?tico por CLAE, o sistema utilizado foi constitu?do de fase A: ?gua:?cido f?rmico (99,7:0,3, v/v) e fase B: metanol:?cido f?rmico (99,7:0,3, v/v) em elui??o gradiente de 40%B - 58%B em 50 minutos, coluna cromatogr?fica (Hichrom) C18 (250x4,0mm, 5?m), fluxo 0,8 mL/min, detec??o em UV a 370 nm, temperatura de 25oC. Por meio de co-inje??o do padr?o + extrato foi observado que Kb1 trata-se de um dos compostos majorit?rios, com tempo de reten??o de 12,47 minutos e apresentou um teor de 15,3% no EH de K. brasiliensis O m?todo apresentou-se linear, preciso, exato e reprodut?vel. De acordo com os resultados obtidos, foi observado que Kb1 pode ser utilizado como marcador qu?mico do EH das folhas de K. brasiliensis, para auxiliar no controle de qualidade da droga vegetal e seus derivados

Analysis of cuticular wax from Kalanchoe daigremontiana leaves was performed to identify the constituent components within the wax, determine how these changed during leaf ontogenesis, and discover how they were distributed within the cuticle. Analysis of extracted cuticular wax by gas chromatography, mass spectrometry, and comparison with authentic standards led to the identification of triterpenoids including glutinol, friedelin, germanicol, epifriedelanol, glutinol acetate and β-amyrin as well as very long chain fatty acid (VLCFA) derivatives including alkanes, primary alcohols, aldehydes, fatty acids, and alkyl esters. Cuticular wax composition in young K. daigremontiana leaves was dominated by triterpenoids, which made up over 70% of the lipid soluble compounds. During leaf ontogenesis, wax composition changed to include a higher proportion of VLCFA derivatives, which made up approximately 50% of cuticular wax in mature leaves. The most abundant triterpenoids in the wax were glutinol and friedelin, both fairly uncommon pentacyclic triterpenoids with a complex proposed biosynthetic mechanism. Tritriacontane (C33 alkane) was the most abundant compound within the VLCFA derivatives. Cuticular wax accumulation was found to correspond well to leaf growth, with both processes slowing at the same time. Variations in the ratio of friedelin-like compounds to glutinol-like compounds during leaf ontogenesis suggest the presence more than one active triterpenoid synthase enzyme in the leaves of K. daigremontiana. VLCFA compounds were found mainly in the epicuticular wax on both the adaxial and abaxial surfaces, while triterpenoids were relatively more abundant in the intracuticular layer. Two different epicuticular wax crystal forms were observed by scanning electron microscopy (SEM) which can be described as platelets with sinuate margins and twisted ribbons. Based on SEM and chemical data as well as previous reports of crystal composition, it is hypothesized that each crystal type has a unique composition, with the platelets containing one or more triterpenoids and the twisted ribbons containing alkanes and other VLCFA derivatives. Confirmation of this hypothesis will have to await further investigation. This research provides information that will aid in the larger goals of characterizing a glutinol or friedelin synthase and understanding the gradients established within epicuticular and intracuticular wax layers.

Analysis of cuticular wax from Kalanchoe daigremontiana leaves was performed to identify the constituent components within the wax, determine how these changed during leaf ontogenesis, and discover how they were distributed within the cuticle. Analysis of extracted cuticular wax by gas chromatography, mass spectrometry, and comparison with authentic standards led to the identification of triterpenoids including glutinol, friedelin, germanicol, epifriedelanol, glutinol acetate and β-amyrin as well as very long chain fatty acid (VLCFA) derivatives including alkanes, primary alcohols, aldehydes, fatty acids, and alkyl esters. Cuticular wax composition in young K. daigremontiana leaves was dominated by triterpenoids, which made up over 70% of the lipid soluble compounds. During leaf ontogenesis, wax composition changed to include a higher proportion of VLCFA derivatives, which made up approximately 50% of cuticular wax in mature leaves. The most abundant triterpenoids in the wax were glutinol and friedelin, both fairly uncommon pentacyclic triterpenoids with a complex proposed biosynthetic mechanism. Tritriacontane (C33 alkane) was the most abundant compound within the VLCFA derivatives. Cuticular wax accumulation was found to correspond well to leaf growth, with both processes slowing at the same time. Variations in the ratio of friedelin-like compounds to glutinol-like compounds during leaf ontogenesis suggest the presence more than one active triterpenoid synthase enzyme in the leaves of K. daigremontiana. VLCFA compounds were found mainly in the epicuticular wax on both the adaxial and abaxial surfaces, while triterpenoids were relatively more abundant in the intracuticular layer. Two different epicuticular wax crystal forms were observed by scanning electron microscopy (SEM) which can be described as platelets with sinuate margins and twisted ribbons. Based on SEM and chemical data as well as previous reports of crystal composition, it is hypothesized that each crystal type has a unique composition, with the platelets containing one or more triterpenoids and the twisted ribbons containing alkanes and other VLCFA derivatives. Confirmation of this hypothesis will have to await further investigation. This research provides information that will aid in the larger goals of characterizing a glutinol or friedelin synthase and understanding the gradients established within epicuticular and intracuticular wax layers.

Submitted by Automa??o e Estat?stica (sst@bczm.ufrn.br) on 2016-03-02T23:14:27Z No. of bitstreams: 1 AldilaneGoncalvesDaFonseca_DISSERT.pdf: 2225811 bytes, checksum: 5e7b981fdc77d287792a011dffc52374 (MD5) / Approved for entry into archive by Arlan Eloi Leite Silva (eloihistoriador@yahoo.com.br) on 2016-03-04T23:20:31Z (GMT) No. of bitstreams: 1 AldilaneGoncalvesDaFonseca_DISSERT.pdf: 2225811 bytes, checksum: 5e7b981fdc77d287792a011dffc52374 (MD5) / Made available in DSpace on 2016-03-04T23:20:31Z (GMT). No. of bitstreams: 1 AldilaneGoncalvesDaFonseca_DISSERT.pdf: 2225811 bytes, checksum: 5e7b981fdc77d287792a011dffc52374 (MD5) Previous issue date: 2014-09-25 / As esp?cies do g?nero Kalanchoe (Crassulaceae) s?o bastante utitlizadas em todo o Brasil, como medicamento para tratar tosse, fur?nculos, gastrite e outras enfermidades. Nesse cen?rio destaca-se K. brasiliensis, conhecida popularmente como coirama ou sai?o. Este trabalho tem como proposta a exposi??o por via oral de camundongos a um extrato hidroalco?lico de folhas de Kalanchoe brasiliensis. Os animais (total de 100) foram divididos em grupos de 12 (6 machos e 6 f?meas) para avalia??o aguda; e grupos de 10 para avalia??o subcr?nica.Os grupos teste foram tratados com doses de 250 mg/Kg, 500 mg/Kg, 1000 mg/Kg e 2000 mg/Kg, e o grupo controle recebeu solu??o salina 0,9 %. Os animais foram observados por 14 dias para avalia??o aguda e 30 dias para avalia??o subcr?nica, e nesse per?odo analisou-se o aparecimento de sinais cl?nicos, altera??es no peso e no consumo de ?gua e ra??o. Ap?s o per?odo de observa??o, foram realizadas a an?lise histopatol?gica dos ?rg?os, as concentra??es s?ricas bioqu?micas e par?metros hematol?gicos para os grupos da avalia??o subcr?nica. N?o foram observadas diferen?as e altera??o do peso corp?reo entre os grupos, nem de consumo de ?gua e ra??o, tamb?m n?o houve mortes entre os grupos, nos dois tipos de ensaios. As an?lises histopatol?gicas demonstraram algumas altera??es compat?veis com baixa toxicidade aguda hep?tica. Os resultados de glicemia, triglicer?deos, ALT, ureia e creatinina mostraram diferen?as entre o grupo controle e os testes nas concentra??es estudadas (p<0,05), por?m estas diferen?as n?o demonstram mudan?a relevante no quadro cl?nico dos animais. Os resultados mostraram que o extrato de K. brasiliensis apresenta baixa toxicidade aguda nas doses utilizadas e n?o apresenta toxicidade quando administrado durante 30 dias. Isto destaca a import?ncia desta esp?cie como futuro e promissor candidato a fitoter?pico, assim que tenha seus ensaios farmacol?gicos conclu?dos. / The species of the genus Kalanchoe (Crassulaceae) are widely used in Brazil as a medicinal product to treat cough, boils, gastritis and other diseases. In this scenario stands out K. brasiliensis, popularly known as coirama or sai?o. This paper aims at the oral exposure of mice to a hydroalcoholic extract of leaves of Kalanchoe brasiliensis. The animals (total of 100) were divided into 12 groups (6 males and 6 females) for the acute assessment; and groups of 10 for subchronic evaluation. Test groups were treated with doses of 250 mg / kg, 500 mg / kg, 1000 mg / kg and 2000 mg / kg and the control group received 0.9% saline. The animals were observed for 14 days for acute evaluation and 30 days for subchronic evaluation, and in that period analyzed the appearance of clinical signs, changes in weight and consumption of water and food. After the observation period, histopathological analysis of the organs, biochemical serum and haematological parameters for the assessment of the subchronic groups were processed. Differences and changes in body weight were not observed among the groups, nor consumption of water and food, there were no deaths among the groups in the two types of assays. Histopathological analysis showed some alterations compatible with low acute hepatic toxicity. The results of blood glucose, triglycerides, ALT, urea and creatinine showed differences between the control group and the test concentrations studied (p <0.05), but these differences do not show relevant change in the clinical picture of animals. The results showed that the extract K. brasiliensis has low acute toxicity at the doses used and no toxicity when administered for 30 days. This highlights the importance of this species as future and promising candidate for phytotherapics, so has its pharmacological trials completed.

Made available in DSpace on 2014-06-12T16:31:58Z (GMT). No. of bitstreams: 2 arquivo6256_1.pdf: 1729667 bytes, checksum: d62ce5ef03ff18914736fc6bce682329 (MD5) license.txt: 1748 bytes, checksum: 8a4605be74aa9ea9d79846c1fba20a33 (MD5) Previous issue date: 2007 / Kalanchoe brasiliensis Cambess é uma Crassulácea conhecida popularmente como saião, coirama-branca, folha-grossa ou folha-suculenta. Na medicina popular é utilizada contra lesões teciduais, bronquites, inflamações e úlceras em muitas partes do mundo. Contra essas afecções, a folha é a parte mais utilizada. Este estudo reporta uma avaliação do extrato hidroetanólico das folhas de K. brasiliensis quanto ao seu potencial toxicológico in vivo e in vitro e sua propriedade antitumoral. Além disso, também foi avaliada a atividade antimicrobiana do extrato hidroetanólico e do óleo essencial das folhas, bem como a alcoolatura do caule de Kalanchoe brasiliensis. A citotoxicidade in vitro da planta foi realizada em larvas de Artemia salina. A CL50 determinada em 704,8 &#956;g/mL tornou o extrato tóxico ao microcrustáceo. Foram realizados ensaios de toxicidade aguda com camundongos fêmeas Albinos Swiss (Mus muscullus), por via intraperitoneal, para observação de parâmetros como alterações comportamentais e efeitos sobre os sistemas nervoso central e autônomo. Nas doses de 1000 mg/kg a 3000 mg/kg de peso corpóreo administradas, foram observadas ações estimulantes seguidas de efeitos depressores ao sistema nervoso central e alterações na locomoção. A DL50 determinada em 1925 mg/kg de peso corpóreo tornou o extrato nocivo por via intraperitoneal. Na análise macroscópica o fígado e os rins apresentaram-se empaledecidos nas doses 250 mg/kg e 125 mg/kg de peso corpóreo. Na dose 62,5 mg/kg foram observadas apenas alterações microscópicas. Na avaliação antitumoral foram testadas as linhagens de Carcinoma de Ehrlich e Sarcoma-180 nas doses 250 mg/kg e 62,5 mg/kg de peso corpóreo do extrato hidroetanólico de K. brasiliensis. Nessas doses a inibição do Sarcoma-180 foi superior a 50%, sendo ambas estatisticamente significantes em comparação ao controle. Para Carcinoma de Ehrlich a inibição foi de 66,59% na maior dose, e apenas esta se mostrou significativa estatisticamente. Na avaliação mascroscópica a massa tumoral apresentou-se delimitada em todas as doses. Dentre os órgãos avaliados nos camundongos com neoplasias, o baço mostrou-se hipotrofiado e o fígado com coloração escurecida no grupo tratado com 250 mg/kg do extrato, no controle e no grupo padrão, para os dois tumores testados. Na menor dose não houve nenhuma alteração nos órgãos dos animais com o tumor de Ehrlich, porém ocorreu esplenomegalia nos animais com Sarcoma-180. A análise fitoquímica foi realizada através de cromatografia em camada delgada analítica utilizando diversas fases móveis e reveladores específicos, indicando os flavonóides como os fitoconstituintes predominantes. No ensaio antimicrobiano foram utilizadas amostras bacterianas gram-positivas e gram-negativas, dentre elas algumas cepas multiresistentes, além de fungos leveduriformes. Entre o óleo essencial e extrato hidroalcoólico das folhas e alcoolatura do caule, apenas o óleo mostrou-se efetivo nas concentrações de 0,25% a 8%, com halos de inibição de até 17 mm. Na concentração de 8%, foi obtido um efeito bacteriostático bem pronunciado, após a sexta hora de exposição da amostra ao óleo essencial. O efeito antimicrobiano foi observado em amostras multiresistentes de Staphylococcus aureus

Submitted by Celso Magalhaes (celsomagalhaes@ufrrj.br) on 2017-06-21T11:22:42Z No. of bitstreams: 1 2016 - Raquel Teixeira Ferreira.pdf: 4123117 bytes, checksum: d2271c63e8c851f1e25afd9a0bd8cf91 (MD5) / Made available in DSpace on 2017-06-21T11:22:42Z (GMT). No. of bitstreams: 1 2016 - Raquel Teixeira Ferreira.pdf: 4123117 bytes, checksum: d2271c63e8c851f1e25afd9a0bd8cf91 (MD5) Previous issue date: 2016-03-30 / Coordena??o de Aperfei?oamento de Pessoal de N?vel Superior - CAPES / Kalanchoe pinnata (KP) is popularly used for treating inflammatory diseases. This study investigated the antinociceptive and anti-inflammatory potential of the KP flower aqueous extract (KPFE), the ethyl acetate (EtOAcF), butanol (BuOHF) and aqueous (AqF) fractions and the main flavonoid [quercetin 3-O-?-L-arabinopyranosyl (1??) ?-L-rhamnopyranoside] (KPFV) of KP, as well as possible mechanisms of action. Swiss albino mice, male (25-35g) were pretreated with KPFE (30-300 mg/kg, s.c.) producing dose-related inhibition of acetic acid-induced writhing (ID50 164.8 mg/kg), and the subcutaneous administration of KPFE (300 mg/kg), EtOAcF (12 mg/kg), BuOHF (15 mg/kg) or AqF (210 mg/kg) reduced leukocyte migration on carrageenan-induced pleurisy in mice at 56.1%, 47.3%, 39.6% and 43.1%, respectively. At doses of up to 240 times smaller than KPFE, KPFV (0.3-3.0 mg/kg, s.c.), also inhibited leukocyte migration (ID50 1.78 mg/kg). In croton oil-induced ear edema in mice, KPFE (3-30 mg/kg, s.c.) and KPFV (0.3-3.0 mg/kg, s.c.) presented dose-related antiedematogenic activity (KPFE - ID50 4.3 mg/kg, KPFV - ID50 0.76 mg/kg). Treatment with KPFE reduced the MPO and TNF-? concentration in the pleural exudates on carrageenaninduced pleurisy test, while KPFV inhibited both isoforms of cyclooxygenase (COX), with IC50 3.8 x 10-5 M (22.1 ?g/mL) for COX-1 and a maximum COX-2 inhibition of 43.5% (IC50 >8.4 x10-5M or 50 ?g/mL). The selectivity index (COX-1 IC50/COX-2 IC50) was <0.44. These results indicate that KPFE and KPFV produced anti-inflammatory and antinociceptive activity through the COX inhibition and TNF-? reduction. This study reveals that the main flavonoid in KP flowers and leaves plays an important role in the ethnomedicinal use of this species for healing inflammatory processes / Kalanchoe pinnata (KP) ? popularmente utilizada para tratamento de doen?as inflamat?rias. Este estudo investigou o potencial antinociceptivo e anti-inflamat?rio do extrato aquoso das infloresc?ncias de KP (EAFL), a fra??o acetato de etila (FAcOEt), a fra??o butan?lica (FBuOH), a fra??o aquosa (FAq) e o flavonoide majorit?rio [quercetina 3-O-?-Larabinopiranosil (1??) ?-L-ramnopiranos?deo] (KPFV) de KP, assim como poss?veis mecanismos de a??o. Camundongos albinos, machos (25-35g) foram pr?-tratados com EAFL (30-300 mg/kg, s.c.), produzindo inibi??o dose dependente das contor??es induzidas por ?cido ac?tico (DI50 164,8 mg/kg), e a administra??o subcut?nea de EAFL (300 mg/kg), FAcOEt (12 mg/kg) ou FBuOH (15 mg/kg) reduziu a migra??o leucocit?ria na pleurisia induzida por carragenina em camundongos em 56,1%, 47,3%, 39,6% e 43,1%, respectivamente. Em doses at? 240 vezes menores que o EAFL, KPFV (0,3-3,0 mg/kg, s.c.), tamb?m foi capaz de inibir a migra??o leucocit?ria (DI50 1,78 mg/kg). No teste do edema de orelha induzido por ?leo de cr?ton em camundongos, EAFL (3-30 mg/kg, s.c.) e KPFV (0,3-3,0 mg/kg, s.c.) apresentaram atividade antiedematog?nica dose dependente (EAFL - DI50 4,3 mg/kg, KPFV - DI50 0,76 mg/kg). O tratamento com EAFL reduziu a concentra??o de MPO e de TNF-? no exsudato pleural obtido do teste da pleurisia induzida por carragenina, enquanto o KPFV inibiu ambas as isoformas de cicloxigenase (COX), com CI50 3,8 x 10-5M (22,1 ?g/mL) para COX-1 e um m?ximo de inibi??o da COX-2 de 43,5% (CI50 >8,4 x10-5M ou 50 ?g/mL). O ?ndice de seletividade (COX-1 CI50/COX-2 CI50) foi < 0,44. Estes resultados indicam que EAFL e KPFV produziram atividade anti-inflamat?ria e antinociceptiva envolvendo a inibi??o da COX e a redu??o de TNF-?. Este estudo revela que o principal flavonoide presente nas infloresc?ncias e folhas de KP possui papel importante no uso etnomedicinal desta esp?cie nos processos de natureza inflamat?ria

Analysis of cuticular wax from Kalanchoe daigremontiana leaves was performed to identify the constituent components within the wax, determine how these changed during leaf ontogenesis, and discover how they were distributed within the cuticle. Analysis of extracted cuticular wax by gas chromatography, mass spectrometry, and comparison with authentic standards led to the identification of triterpenoids including glutinol, friedelin, germanicol, epifriedelanol, glutinol acetate and β-amyrin as well as very long chain fatty acid (VLCFA) derivatives including alkanes, primary alcohols, aldehydes, fatty acids, and alkyl esters. Cuticular wax composition in young K. daigremontiana leaves was dominated by triterpenoids, which made up over 70% of the lipid soluble compounds. During leaf ontogenesis, wax composition changed to include a higher proportion of VLCFA derivatives, which made up approximately 50% of cuticular wax in mature leaves. The most abundant triterpenoids in the wax were glutinol and friedelin, both fairly uncommon pentacyclic triterpenoids with a complex proposed biosynthetic mechanism. Tritriacontane (C33 alkane) was the most abundant compound within the VLCFA derivatives. Cuticular wax accumulation was found to correspond well to leaf growth, with both processes slowing at the same time. Variations in the ratio of friedelin-like compounds to glutinol-like compounds during leaf ontogenesis suggest the presence more than one active triterpenoid synthase enzyme in the leaves of K. daigremontiana. VLCFA compounds were found mainly in the epicuticular wax on both the adaxial and abaxial surfaces, while triterpenoids were relatively more abundant in the intracuticular layer. Two different epicuticular wax crystal forms were observed by scanning electron microscopy (SEM) which can be described as platelets with sinuate margins and twisted ribbons. Based on SEM and chemical data as well as previous reports of crystal composition, it is hypothesized that each crystal type has a unique composition, with the platelets containing one or more triterpenoids and the twisted ribbons containing alkanes and other VLCFA derivatives. Confirmation of this hypothesis will have to await further investigation. This research provides information that will aid in the larger goals of characterizing a glutinol or friedelin synthase and understanding the gradients established within epicuticular and intracuticular wax layers. / Science, Faculty of / Chemistry, Department of / Graduate

O desenvolvimento de cepas bacterianas multirresistentes tornou-se uma preocupação global. As plantas medicinais têm sido exploradas como uma fonte potencial terapêutica para o tratamento de várias doenças e para o desenvolvimento de medicamentos mais eficazes contra infecções bacterianas. Assim, o objetivo deste estudo foi avaliar a atividade antibacteriana do extrato de cascas e folhas de Anacardium humile (St.) Hil, extrato de cascas de Davilla nítida (Vahl.) Kubitski, e extrato de folhas de Kalanchoe pinnata (Lam.) Pers. contra bactérias resistentes a múltiplos fármacos isoladas de infecções do pé diabético, e também contra cepas de referência da American Type Culture Collection (ATCC). A atividade antibacteriana foi avaliada pelos métodos de Ágar Disco-Difusão (DD), Microdiluição em Caldo (MC), Ensaio “Checkerboard” e Ensaio “Time-Kill”. O extrato das cascas de Davilla nitida (Vahl.) Kubitski mostrou atividade antibacteriana expressiva contra todos os grupos de bactérias testadas. O método de microdiluição em caldo foi mais sensível que o método disco-difusão para determinar a atividade antibacteriana presente no extrato das cascas. O extrato de cascas inibiu o crescimento de bactérias com altos níveis de resistência aos antibióticos, como Pseudomonas spp. (100,00%), Enterobacter spp. (88,89%), Staphylococcus aureus (54,55%), Streptococcus pneumoniae (75,00%), Staphylococcus saprophyticus (92,86%). A combinação do extrato das cascas com antibióticos resultou em efeito aditivo contra a maioria das cepas testadas. O perfil cinético de tempo de morte do extrato de cascas mostrou propriedades bactericidas tempo-dependentes. Nossos resultados sugerem a presença de compostos bioativos nesta planta que podem ser utilizados para o desenvolvimento de novos agentes antibacterianos. O extrato de folhas de Kalanchoe pinnata (Lam.) Pers. mostrou atividade antibacteriana contra Citrobacter spp. (100,00%), Citrobacter youngae (ATCC-29935) (100,00%), Enterobacter spp. (33,33%), Pseumodonas spp. (66,67%), Pseudomonas aeruginosa (ATCC-27853) (100,00%). O extrato de folhas quando associado com antibiótico amoxicilina/ácido clavulânico ou cefoxitina não evidenciou qualquer potencialização da atividade antibacteriana contra as bactérias Gram-positivas e Gram-negativas, sugerindo que extrato e antibióticos possuem mecanismos independentes relacionados à inibição do crescimento bacteriano. Nossos resultados mostraram que o extrato etanólico das folhas de Kalanchoe pinnata (Lam.) Pers. teve uma boa atividade antimicrobiana contra algumas bactérias resistentes a múltiplos fármacos. Entretanto, estudos adicionais e mais específicos são recomendados para determinar a eficácia deste extrato no tratamento de infecções bacterianas. O extrato das cascas e folhas de Anacardium humile (St.) Hil, tanto no método Disco-difusão quanto em microdiluição em caldo, inibiu significantemente o crescimento das bactérias Gram-positivas e Gram-negativas, como também o crescimento das bactérias comumente encontradas em lesões de pés diabéticos. Pelo método “Checkerboard”, a combinação do extrato das cascas (35,00%) e folhas (40,00%) sobre as bactérias Gram-negativas apresentaram efeitos sinérgicos semelhantes, enquanto que para as bactérias Gram-positivas, os extratos das folhas (45,00%) apresentaram um efeito sinérgico mais efetivo do que os extratos das cascas (17,50%). O perfil cinético “Time-Kill” apresentou atividade bactericida com propriedades dose e tempo dependente, sugerindo que os extratos das cascas e folhas podem potencializar os efeitos dos antibióticos, o que sugere que os extratos de Anacardium humile (St.) Hil podem ser usados com uma fonte alternativa de pesquisa para agentes antibacterianos com ação em bactérias Gram-positivas e Gram-negativas resistentes a multidrogas. No entanto, o isolamento dos compostos bioativos e estudos adicionais devem ser realizados para entender os mecanismos de ação bactericida, para definir a real eficácia e os efeitos tóxicos. / Development of multidrug resistant bacterial strains has become a global concern. Medicinal plants have been explored as a source of molecules with therapeutic potential for the treatment of various diseases and to the development of better drug against bacterial infections. Thus, the aim of this study was to evaluate the antibacterial activity of the bark and leaves extracts of Anacardium humile (St.) Hil, bark extract of Davilla nitida (Vahl.) kubitski, of leaf extract of Kalanchoe pinnata (Lam.) Pers. and against multidrug resistant bacteria isolated from diabetic foot infections, and also of reference strains from the American Type Culture Collection (ATCC). Antibacterial activity of the bark extract was evaluated by agar Disk-Diffusion (DD), Broth Dilution (BD), “Checkerboard” and “Time-Kill” methods. The bark extract of Davilla nitida (Vahl.) kubistki showed a significant antibacterial activity against all groups of bacteria tested. Broth dilution was more sensitive for determining the antibacterial activity of the bark extract than the Disk-Difusion method. The bark extract inhibited the growth of bacteria with high-levels of antibiotic-resistance, such as Pseudomonas spp. (100,00%), Enterobacer spp. (88,89%), Staphylococcus aureus (54,55%), Streptococcus pneumoniae (75,00%), Staphylococcus saprophyticus (92,86%). The combination of extract with antibiotics resulted in an additive effect against most of the strains tested. “Time-Kill” kinetics profiles of bark extract showed bactericidal and time-dependent properties. Our results suggest the presence of bioactive compounds in this plant that could be useful for the development of new antibacterial agents. The leaf extract of Kalanchoe pinnata (Lam.) Pers. showed antibacterial activity against Citrobacter spp. (100,00%), Citrobacter youngae (ATCC-29935) (100,00%), Enterobacer spp. (33.33%), Pseumodonas spp. (66.67%), Pseudomonas aeruginosa (ATCC-27853) (100.00%). Extract of leaf associated with amoxicillin/clavulanic acid or cefoxitin antibiotics did not show any potentiation of antibacterial activity against both Gram-positive and Gram-negative bacteria, suggesting that extract and antibiotics have independent mechanisms related to inhibition of bacterial growth. Our results showed that the ethanolic leaf extract of Kalanchoe pinnata (Lam.) Pers. had a good antimicrobial activity against some multidrug resistant bacteria. However, further and more specific studies, are recommended to determine the efficacy of this extract in the treatment of bacterial infections. Barks and leaves extracts of Anacardium humile (St.) Hil, both in the Disk-Difusion and Broth Dilution methods, inhibited the growth of Gram-positive and Gram-negative bacteria, as well as the growth of bacteria commonly found in diabetic foot lesions. By the “Checkerboard” method, the combination of bark extract (35,00%) and leaves (40,00%) on Gram-negative bacteria had similar synergistic effects, while for Gram-positive bacteria, leaf extracts (45,00%) presented a synergistic effect more effective than the bark extracts (17,50%). The “Time-Kill” kinetic profile showed bactericidal activity with dose and time-dependent properties, suggesting that bark and leaf extracts may potentiate the effects of antibiotics, suggesting that Anacardium humile (St.) Hil extracts can be used with an alternative source of research for antibacterial agents acting on multidrug resistant Gram- positive and Gram- negative bacteria. However, the isolation of bioactive compounds and additional studies should be performed to understand the mechanisms of bactericidal action, to define real efficacy and toxic effects.