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The iridoids are a large family of monoterpenoid natural products that possess a wide range of biological activities. A great deal of research has already been done in the field of iridoid total synthesis, but limitations still remain. Specifically, few syntheses of iridoid β-glycosides have been reported. This work describes the 14 step asymmetric total synthesis of the iridoid β-glycoside (+)-geniposide utilizing a phosphine-catalyzed [3+2] cycloaddition as the key step. Other noteworthy steps in the synthesis include a palladium-catalyzed kinetic resolution and a previously unutilized method for iridoid glycosidation. In addition to describing the synthesis of (+)-geniposide, this dissertation will also review 1) phosphine-catalyzed cycloaddition reactions and 2) previous enantioselective total syntheses of iridoid glycosides. / text

Iridoides são metabólitos secundários provenientes de angiospermas eudicotiledôneas, presentes principalmente em espécies das ordens Gentianales e Lamiales. Os iridoides dividem-se em carbocíclicos e seco-iridoides, ocorrendo comumente na forma glicosilada. Estes compostos são marcadores taxonômicos em algumas famílias vegetais e apresentam diversas atividades biológicas tais como cardiovascular, neuroprotetora e anti-Leishmania. Diante da importância dos iridoides, este trabalho teve como finalidade a prospecção química destes metabólitos em espécies nativas do Rio Grande Grande dos Sul, bem como a semissíntese de análogos e a investigação da atividade anti-Leishmania através de ensaios in vitro e modelagem molecular. Os compostos isolados foram identificados através de métodos espectroscópicos e os resultados comparados aos descritos na literatura. A partir de Escallonia bifida e Escallonia megapotamica (Escalloniaceae) foram isolados asperulosídeo, desacetilasperulosídeo, geniposídeo, ácido geniposídico e dafilosídeo, sendo que o asperulosídeo foi convertido em asperulosídeo tetraacetilado por meio de semissíntese. De Angelonia integerrima (Scrophulariaceae) foram obtidos galiridosídeo e antirrídeo. Nos experimentos in vitro para atividade anti-Leishmania, asperulosídeo, galiridosídeo, geniposídeo, ipolamida e teveridosídeo, nas concentrações 5-100 μM, não demonstraram inibição frente às formas promastigotas de Leishmania amazonensis. O estudo de modelagem molecular destes iridoides e daqueles descritos na literatura com atividade anti-Leishmania propôs um modelo farmacofórico que demonstrou que as diferenças estruturais não são responsáveis pela inatividade das moléculas isoladas neste trabalho. A perspectiva é realizar ensaios enzimáticos de tripanotiona redutase, bem com docking molecular e estudos de dinâmica molecular para investigar as interações entre grupamentos farmacofóricos das moléculas isoladas e o sítio de ligação de tripanotiona redutase. / Iridoids are secondary metabolites of eudicotyledonous angiosperms, present mainly in species of the orders Gentianales and Lamiales. The iridoids are divided into carbocyclic and seco-iridoids, occurring commonly in the glycosylated form. These compounds are taxonomic markers in same families of plants and have shown cardiovascular, neuroprotective and anti-Leishmania activities. In view of the importance of iridoids, this work aimed to the chemical prospection of these metabolites of native species of Rio Grande do Sul, as well as semi-synthesis of analogues and to investigate the anti-Leishmania activity through in vitro assays and molecular modeling. The isolated compounds were identified by spectroscopic methods and the results compared to those described in literature. From Escallonia bifida and Escallonia megapotamica (Escalloniaceae) asperuloside, deacetylasperuloside, geniposide, geniposidic acid and daphyloside were isolated, being asperuloside developed in asperuloside tetraacetylated by means of semi- synthesis. From Angelonia integerrima (Scrophulariaceae) galiridoside and antirride were obtained. In the in vitro experiments for anti-Leishmania activity, asperuloside, galiridoside, geniposideo, ipolamiide and theveridoside in concentrations 5-100 μM, did not demonstrate inhibition in promastigote form of Leishmania amazonensis. The molecular modeling study of these iridoids and those described in the literature with anti-Leishmania activity proposed a pharmacophoric model that demonstrated that the structures are not responsible by the inactivity of the molecules isolated in this work. The prospect is to carry out enzymatic assays of trypanothione redutase as well as molecular docking and molecular dynamics studies to investigate the interactions between pharmacophoric grouping of the isolated molecules and the trypanothione reductase binding site.

Iridoides são metabólitos secundários provenientes de angiospermas eudicotiledôneas, presentes principalmente em espécies das ordens Gentianales e Lamiales. Os iridoides dividem-se em carbocíclicos e seco-iridoides, ocorrendo comumente na forma glicosilada. Estes compostos são marcadores taxonômicos em algumas famílias vegetais e apresentam diversas atividades biológicas tais como cardiovascular, neuroprotetora e anti-Leishmania. Diante da importância dos iridoides, este trabalho teve como finalidade a prospecção química destes metabólitos em espécies nativas do Rio Grande Grande dos Sul, bem como a semissíntese de análogos e a investigação da atividade anti-Leishmania através de ensaios in vitro e modelagem molecular. Os compostos isolados foram identificados através de métodos espectroscópicos e os resultados comparados aos descritos na literatura. A partir de Escallonia bifida e Escallonia megapotamica (Escalloniaceae) foram isolados asperulosídeo, desacetilasperulosídeo, geniposídeo, ácido geniposídico e dafilosídeo, sendo que o asperulosídeo foi convertido em asperulosídeo tetraacetilado por meio de semissíntese. De Angelonia integerrima (Scrophulariaceae) foram obtidos galiridosídeo e antirrídeo. Nos experimentos in vitro para atividade anti-Leishmania, asperulosídeo, galiridosídeo, geniposídeo, ipolamida e teveridosídeo, nas concentrações 5-100 μM, não demonstraram inibição frente às formas promastigotas de Leishmania amazonensis. O estudo de modelagem molecular destes iridoides e daqueles descritos na literatura com atividade anti-Leishmania propôs um modelo farmacofórico que demonstrou que as diferenças estruturais não são responsáveis pela inatividade das moléculas isoladas neste trabalho. A perspectiva é realizar ensaios enzimáticos de tripanotiona redutase, bem com docking molecular e estudos de dinâmica molecular para investigar as interações entre grupamentos farmacofóricos das moléculas isoladas e o sítio de ligação de tripanotiona redutase. / Iridoids are secondary metabolites of eudicotyledonous angiosperms, present mainly in species of the orders Gentianales and Lamiales. The iridoids are divided into carbocyclic and seco-iridoids, occurring commonly in the glycosylated form. These compounds are taxonomic markers in same families of plants and have shown cardiovascular, neuroprotective and anti-Leishmania activities. In view of the importance of iridoids, this work aimed to the chemical prospection of these metabolites of native species of Rio Grande do Sul, as well as semi-synthesis of analogues and to investigate the anti-Leishmania activity through in vitro assays and molecular modeling. The isolated compounds were identified by spectroscopic methods and the results compared to those described in literature. From Escallonia bifida and Escallonia megapotamica (Escalloniaceae) asperuloside, deacetylasperuloside, geniposide, geniposidic acid and daphyloside were isolated, being asperuloside developed in asperuloside tetraacetylated by means of semi- synthesis. From Angelonia integerrima (Scrophulariaceae) galiridoside and antirride were obtained. In the in vitro experiments for anti-Leishmania activity, asperuloside, galiridoside, geniposideo, ipolamiide and theveridoside in concentrations 5-100 μM, did not demonstrate inhibition in promastigote form of Leishmania amazonensis. The molecular modeling study of these iridoids and those described in the literature with anti-Leishmania activity proposed a pharmacophoric model that demonstrated that the structures are not responsible by the inactivity of the molecules isolated in this work. The prospect is to carry out enzymatic assays of trypanothione redutase as well as molecular docking and molecular dynamics studies to investigate the interactions between pharmacophoric grouping of the isolated molecules and the trypanothione reductase binding site.

Iridoides são metabólitos secundários provenientes de angiospermas eudicotiledôneas, presentes principalmente em espécies das ordens Gentianales e Lamiales. Os iridoides dividem-se em carbocíclicos e seco-iridoides, ocorrendo comumente na forma glicosilada. Estes compostos são marcadores taxonômicos em algumas famílias vegetais e apresentam diversas atividades biológicas tais como cardiovascular, neuroprotetora e anti-Leishmania. Diante da importância dos iridoides, este trabalho teve como finalidade a prospecção química destes metabólitos em espécies nativas do Rio Grande Grande dos Sul, bem como a semissíntese de análogos e a investigação da atividade anti-Leishmania através de ensaios in vitro e modelagem molecular. Os compostos isolados foram identificados através de métodos espectroscópicos e os resultados comparados aos descritos na literatura. A partir de Escallonia bifida e Escallonia megapotamica (Escalloniaceae) foram isolados asperulosídeo, desacetilasperulosídeo, geniposídeo, ácido geniposídico e dafilosídeo, sendo que o asperulosídeo foi convertido em asperulosídeo tetraacetilado por meio de semissíntese. De Angelonia integerrima (Scrophulariaceae) foram obtidos galiridosídeo e antirrídeo. Nos experimentos in vitro para atividade anti-Leishmania, asperulosídeo, galiridosídeo, geniposídeo, ipolamida e teveridosídeo, nas concentrações 5-100 μM, não demonstraram inibição frente às formas promastigotas de Leishmania amazonensis. O estudo de modelagem molecular destes iridoides e daqueles descritos na literatura com atividade anti-Leishmania propôs um modelo farmacofórico que demonstrou que as diferenças estruturais não são responsáveis pela inatividade das moléculas isoladas neste trabalho. A perspectiva é realizar ensaios enzimáticos de tripanotiona redutase, bem com docking molecular e estudos de dinâmica molecular para investigar as interações entre grupamentos farmacofóricos das moléculas isoladas e o sítio de ligação de tripanotiona redutase. / Iridoids are secondary metabolites of eudicotyledonous angiosperms, present mainly in species of the orders Gentianales and Lamiales. The iridoids are divided into carbocyclic and seco-iridoids, occurring commonly in the glycosylated form. These compounds are taxonomic markers in same families of plants and have shown cardiovascular, neuroprotective and anti-Leishmania activities. In view of the importance of iridoids, this work aimed to the chemical prospection of these metabolites of native species of Rio Grande do Sul, as well as semi-synthesis of analogues and to investigate the anti-Leishmania activity through in vitro assays and molecular modeling. The isolated compounds were identified by spectroscopic methods and the results compared to those described in literature. From Escallonia bifida and Escallonia megapotamica (Escalloniaceae) asperuloside, deacetylasperuloside, geniposide, geniposidic acid and daphyloside were isolated, being asperuloside developed in asperuloside tetraacetylated by means of semi- synthesis. From Angelonia integerrima (Scrophulariaceae) galiridoside and antirride were obtained. In the in vitro experiments for anti-Leishmania activity, asperuloside, galiridoside, geniposideo, ipolamiide and theveridoside in concentrations 5-100 μM, did not demonstrate inhibition in promastigote form of Leishmania amazonensis. The molecular modeling study of these iridoids and those described in the literature with anti-Leishmania activity proposed a pharmacophoric model that demonstrated that the structures are not responsible by the inactivity of the molecules isolated in this work. The prospect is to carry out enzymatic assays of trypanothione redutase as well as molecular docking and molecular dynamics studies to investigate the interactions between pharmacophoric grouping of the isolated molecules and the trypanothione reductase binding site.

Submitted by Automa??o e Estat?stica (sst@bczm.ufrn.br) on 2016-06-23T20:55:01Z No. of bitstreams: 1 CinthyaSaraivaDeAssis_DISSERT.pdf: 1209351 bytes, checksum: 5bb50037084ea9c40b94cff6abdaf523 (MD5) / Approved for entry into archive by Arlan Eloi Leite Silva (eloihistoriador@yahoo.com.br) on 2016-06-28T17:58:44Z (GMT) No. of bitstreams: 1 CinthyaSaraivaDeAssis_DISSERT.pdf: 1209351 bytes, checksum: 5bb50037084ea9c40b94cff6abdaf523 (MD5) / Made available in DSpace on 2016-06-28T17:58:44Z (GMT). No. of bitstreams: 1 CinthyaSaraivaDeAssis_DISSERT.pdf: 1209351 bytes, checksum: 5bb50037084ea9c40b94cff6abdaf523 (MD5) Previous issue date: 2015-10-27 / O uso terap?utico de plantas medicinais tem contribu?do desde a antiguidade de forma ben?fica para a sa?de. No entanto, muitas esp?cies carecem de evid?ncias cient?ficas que forne?am embasamento para a sua utiliza??o na pr?tica terap?utica. Neste contexto, encontra-se a esp?cie Genipa americana L. (Rubiaceae), conhecida popularmente como jenipapo e utilizada como antissifil?tica, antiulcerosa e anti-hemorr?gica, contra hematomas, como t?nico, afrodis?aco, etc. Visto que, essa esp?cie carece de estudos toxicol?gicos, o objetivo deste trabalho foi avaliar a toxicidade in vivo (toxicidade aguda e subcr?nica) e in vitro (citotoxicidade) do extrato dos frutos da G. americana. O extrato hidroetan?lico dos frutos de G. americana foi preparado por macera??o. Foi realizada uma an?lise fitoqu?mica preliminar para avaliar a presen?a de metab?litos secund?rios no extrato. No estudo da citotoxicidade do extrato (0,1; 1,0; 10; 100 e 1000 ?g/100?L) foram utilizadas linhagens de c?lulas normais (3T3) e tumorais (786-0, HepG2 e B16), analisados pelo ensaio do MTT. Para a avalia??o da toxicidade aguda (dose ?nica de 2000 mg/Kg) e subcr?nica (100, 500 e 1000 mg/Kg por 30 dias) foram utilizados camundongos Swiss de ambos os sexos. Ao final dos experimentos, amostras de sangue e ?rg?os foram coletados para an?lise. Os dados entre os grupos foram comparados pelo test T ou por ANOVA com p?s-teste de Dunnett com n?vel de signific?ncia de 5%. O estudo fitoqu?mico do extrato indicou principalmente a presen?a de irid?ides. Os ensaios de citotoxicidade apresentaram resultados de at? 70% de inibi??o celular frente ? linhagem de B16, na dose de 1000 ?g/100?L, e at? 29% frente ? linhagem 786-0, na dose de 10 ?g/100?L. O extrato n?o promoveu morte nas c?lulas 3T3 e HepG2. Durante as avalia??es in vivo, n?o houve mortes de animais. A an?lise das amostras de sangue revelou que os animais submetidos ? avalia??o da toxicidade aguda apresentaram leves altera??es hep?ticas, e que os animais submetidos ? avalia??o da toxicidade subcr?nica apresentaram altera??es no peso relativo do rim esquerdo e na concentra??o plasm?tica de ur?ia. N?o foram observadas diferen?as entre grupos testes e controles na avalia??o histopatol?gica dos ?rg?os coletados. Apesar das altera??es encontradas nos ensaios de toxicidade in vivo, segundo os crit?rios descritos pelos Guias OECD, sugere-se que o extrato hidroetan?lico dos frutos da G. americana seja classificado como de baixa toxicidade. A citotoxicidade do extrato sugere que omesmo possui potencial contra as linhagens de melanoma (B16) e carcinoma renal (786-0). / The therapeutic use of medicinal plants has contributed since antiquity in a beneficial way for health. However, many species lacks of scientific evidence which provide basis for their use in therapeutic practice. In this context is the Genipa americana L. species (Rubiaceae), popularly known as jenipapo and used to treat syfilis, ulcer and hemorrhagic disturbs. It's also used against bruising, as tonic and as aphrodisiac. Due this species lacks toxicological studies, the aim of this study was to evaluate the toxicity in vivo (acute and sub-chronic toxicity) and in vitro (cytotoxicity) of the hydroethanolic extract from G. americana fruits. The hydroethanolic extract of G. americana fruits was prepared by maceration. A preliminary phytochemical analysis was performed to assess the presence of secondary metabolites in the extract. The cytotoxicity study of the extract (0.1, 1.0, 10, 100 and 1000 mg / 100 ul) were performed against normal cells (3T3) and tumor (786-0, HepG2 and B16), analyzed by the MTT assay. To evaluate the acute (single dose of 2000 mg / Kg) and subchronic (100, 500 and 1000 mg / kg for 30 days) toxicity Swiss mice of both sexes were used. At the end of the experiment, blood samples and organs were collected for analysis. Data between groups were compared by t test or ANOVA with Dunnett's post-test with 5% significance level. The phytochemical study of the extracts mainly indicated the presence of iridoids. Results for cytotoxicity tests showed up to 70% inhibition of B16 cell line at a dose of 1000 mg / 100 ul, and up to 29% inhibition of 786-0 at a dose of 10 ug / 100 ul. The extract did not cause death in 3T3 and HepG2 cells. During the in vivo assays, there were no animal deaths. Analysis of blood samples revealed that the animals submitted to the evaluation of acute toxicity had changes in AST and ALT, and that the animals evaluated for subchronic toxicity showed changes in the relative wet weight of the kidney and plasma urea concentration. No differences were observed between groups on histopathological evaluation of the collected organs. Despite the changes found in the in vivo toxicity tests, using the criteria described by the OECD Guidelines, it is suggested that the hydroethanolic extract of the fruits of the G. americana is classified as low toxicity. The cytotoxicity of the extract suggests that they have potential against melanoma cell lines (B16).

Orientador: Raquel Marques Braga / Tese (doutorado) - Universidade Estadual de Campinas, Instituto de Quimica / Made available in DSpace on 2018-08-04T22:47:27Z (GMT). No. of bitstreams: 1 Lima_ValeriaBittencourtde_D.pdf: 9598037 bytes, checksum: 83f061fb32519074a309e1975981ace5 (MD5) Previous issue date: 2005 / Resumo: Nosso trabalho tem por objetivo o isolamento e a elucidação estrutural dos metabólitos secundários de Himatanthus obovatus (família Apocynaceae, sub-família Rauvolfioideae). Apenas cinco espécies de Himatanthus já foram estudadas do ponto de vista químico. O material de H. obovatus utilizado nesse trabalho foi coletado na Chapada dos Guimarães (MT) e em Casa Branca (SP). Utilizando diferentes metodologias de extração e tratamento dos extratos etanólicos brutos foram isoladas 5 lignanas: pinoresinol, isolariciresinol, hidroxipinoresinol, lariciresinol e olivil; 3 nor-isoprenóides: blumenol C, blumenol A e um nor-isoprenóide inédito; o iridóide plumieride, misturas dos terpenos: acetato de lupeol + acetato de a-amirina + acetato de b-amirina + germanicol e stigmasterol + b-sitosterol + campesteroI a, após a acetilação do extrato etanólico bruto, o glicitol inositol. Os extratos Diclorometânico (CDCb) e Etanólico (CECb) da casca de H. obovatus (Casa Branca (SP) foram submetidos aos testes com Artemia salina Leach. e de atividade antiproliferativa frente à 4 linhagens celulares derivadas de tumores humanos: leucemia (K562), pulmão (NCI460), melanoma (UACC62) e mama (MCF7). Os resultados dos dois testes foram bastante coerentes, já que ambos mostraram resultados promissores para o extrato CDCb. Os testes de bioautografia foram realizados com os extratos da casca de H. obovatus (Casa Branca): CHCb (heptano), CDCb (diclorometano) e CECb (etanol) e com as substâncias isoladas: pinoresinol, isolariciresinol, blumenol C, blumenol A, nor-isoprenóide inédito, hidroxipinoresinol, lariciresinol, plumieride e inositol, frente aos fungos: Alternaria alternata, Aspergillus fumigatus, A. niger, Candida albicans, Cladosporium cladosporioides, Fusarium oxysporium, Penicillium oxalicum, P. funicullosum e Rhizopus orizae. e frente às bactérias: Bacillus subtilis, Escherichia coli, Micrococcus luteus, Salmonella typhimurim, Staphilococcus aureus e Streptococcus mutans. Foram observadas atividades bactericida para as lignanas: isolariciresinol frente à bactéria S. mutans e lariciresinol frente à bactéria S. aureus. Os compostos isolados de H. obovatus permitiram comparar filogeneticamente este gênero aos gêneros Tabernaemontana e Rauvolfia (pertencentes às mesmas família e sub-família), estudados anteriormente em nosso grupo de pesquisas e ricos em alcalóides indólicos. / Abstract: Our objective is the isolation and identification of the compounds from Himatanthus obovatus (family Apocynaceae and sub-family Rauvolfioideae). Five species from genus Himatanthus have been chemically studied. H. obovatus was collected in Chapada dos Guimarães (MT state, Brazil) and in Casa Branca (SP State, Brazil). We used different methodologies for extraction and purification of the extracts, yielding 5 lignans: pinoresinol, isolariciresinol, hydroxypinoresinol, lariciresinol and olivil; 3 nor-isoprenoids: blumenol C, blumenol A and one unknown nor-isoprenoid; the iridoid plumieride, a mixture of terpenes: lupeol acetate + a-amirin acetate + b-amirin acetate + germanicol and stigmasterol + b-sitosterol + campesterol and, after acetylation of the crude ethanolic extract, the glycitol inositol. The diclorometanic (CDCb) and ethanolic (CECb) extracts trom the bark of H. obovatus (Casa Branca - SP) have been tested with Artemia salina Leach. and for antiproliferative activity against 4 carcinoma cell lines derived from human cancer: leukemia (K562), lung (NCI1460), melanoma (UACC62) and breast (MCF7). The good results with the CDCb extract in both tests suggest that this extract is a development candidate. The Bioautography tests were made with the heptanic (CHCb), diclorometanic (CDCb) and ethanolic (CECb) extracts from the bark of H. obovatus (Casa Branca - SP) and with the isolated substances: pinoresinol, isolariciresinel, blumenol C, blumenol A, the unknown nor-isoprenoid, hydroxypinoresinol, lariciresinol, plumieride and inositol against the fungi: Alternaria alternata, Aspergillus fumigatus, A. niger, Candida albicans, Cladosporium cladosporioides, Fusarium oxysporium, Penicillium oxalicum P. funicullosum and Rhizopus orizae and against the bacteria: Bacillus subtilis, Escherichia coli, Micrococcus luteus, Salmonella typhimurim, Staphilococcus aureus and Streptococcus mutans. We observed bactericide activity at the lignans: isolariciresinol against the bacteria S. mutans and lariciresinol against the bacteria S. aureus. The coumpounds isolated from H. obovatus allowed us to phylogenetically compare this genus to the genera Tabernaemontana and Rauvolfia (belonging to the same family and sub-family), previously studied in our group and rich in indolic alkaloids. / Doutorado / Quimica Organica / Doutor em Ciências

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No. of bitstreams: 1 Dissertação - Aimêe Almeida de Oliveira.pdf: 2799798 bytes, checksum: 18ae1473c9870ce5fe4e2cba3093128f (MD5) Previous issue date: 2013-07-19 / FAPEAM « Fundação de Amparo à Pesquisa do Estado do Amazonas / The family Apocynaceae presents a wide variety of secondary metabolites with potential biological and pharmaceutical activities, which has attracted great interest in studies of their genres. The genus Himatanthus has thirteen species that are widely distributed in northern Brazil. The species of the Amazon region, Himatanthus sucuuba is a latex containing tree, whose trunk is erect and rough skin, popularly known as sucuba, sucuuba or janaguba. The latex, barks and leaves are used in folk medicine as antitumor, antifungal, anti-anemic, anthelmintic and for treatment of gastritis and arthritis. In the present study the roots of H. sucuuba were used to obtain hexane, dichloromethane and methanol extracts. From the hexane extract was isolated the mixture of lupeol cinnamates, α and β-amyrin. The dichloromethane extract was subjected to fractionation providing the iridoids plumericine, isoplumericine, plumieridine and the steroid sitosterol. From the methanol extract were isolated after successive fractionations, the iridoid plumieride and the terpenes lupeol acetate and the mixture of ursolic and oleanolic acids. The antioxidant activity was evaluated by sequestration capacity of the stable radical 2,2 - diphenyl-1-picrylhydrazyl (DPPH) using quercetin as standard. The methanol extract of the dried roots / the dichloromethane extract was better with antioxidant activity with the concentration of CS50 in 244.56 mg/mL. The toxicity test on Artemia salina hexane extract obtained from the fresh root, showed activity more pronounced at 111.06 mg/mL. Quantification of iridoids in different species of the genus Himatanthus proved the presence of plumieride as the major constituent in all analyzed species. The analysis of different samples analyzed by ultra efficient liquid chromatography coupled with mass spectrometry (UPLC-MS) led to the identification of iridoid 15-desmethylplumieride and plumieride, present only in the green fruits of the species H. sucuuba. / A família Apocynaceae apresenta uma grande diversidade de metabólitos secundários, com potencial biológico e farmacêutico, o que tem despertado grande interesse nos estudos de seus gêneros. O gênero Himatanthus possui treze espécies que estão amplamente distribuídas na região norte do Brasil. A espécie da região amazônica, Himatanthus sucuuba, é latescente, arbórea, cujo tronco é ereto e a casca rugosa, popularmente conhecida como sucuba, sucuuba ou janaguba. O látex, cascas e folhas são utilizados na medicina popular como antitumoral, antifúngico, antianêmico, vermífugo e no tratamento de gastrites e artrites. No presente estudo as raízes de H. sucuuba foram utilizadas para a obtenção dos extratos hexano, diclorometano e metanol. Do extrato hexano foi isolada a mistura dos cinamatos de lupeol, α e β-amirina. O extrato diclorometano foi submetido a fracionamento cromatográfico fornecendo os iridóides plumericina, isoplumericina, plumieridina e o esteróide sitosterol. Do extrato metanol foram isoladas, após sucessivos fracionamentos, o iridóide plumierídeo, e os terpenos acetato de lupeol e a mistura dos ácidos ursólico e oleanólico. A atividade antioxidante foi avaliada por meio da capacidade de sequestro do radical estável 2,2- difenil-1-picrilhidrazila (DPPH), utilizando quercetina como padrão. O extrato metanol das raízes secas/parte diclorometano foi o extrato com melhor atividade antioxidante com a concentração de CS50 em 244, 56 μg/mL. No teste de toxicidade frente à Artemia salina o extrato hexano obtido da raiz fresca, apresentou a atividade mais pronunciada 111,06 μg/mL. A quantificação de iridóides em diferentes espécies do gênero Himatanthus comprovou a presença do plumierídeo constituinte majoritário em todas as espécies analisadas. As análises de diferentes amostras realizadas pela cromatografia líquida de ultra eficiência acoplada ao espectrômetro de massas (CLUE-EM) conduziu à identificação dos iridóides 15-desmetilplumierideo e plumierídeo, presentes apenas nos frutos verdes da espécie H. sucuuba.

Orientador: Adriana Zerlotti Mercadante / Tese (doutorado) - Universidade Estadual de Campinas, Faculdade de Engenharia de Alimentos / Made available in DSpace on 2018-08-25T12:01:22Z (GMT). No. of bitstreams: 1 Bentes_AdriadeSousa_D.pdf: 1114101 bytes, checksum: 01c7b622801719d19ba492f29a102dc1 (MD5) Previous issue date: 2014 / Resumo: A ingestão de alimentos ricos em compostos bioativos, tais como iridoides, compostos fenólicos e carotenoides, está relacionada à redução do risco de desenvolvimento de doenças crônico-degenerativas. Para identificar as melhores fontes destes compostos é necessário o desenvolvimento de métodos adequados de extração que preservem a composição original da amostra. Os perfis de iridoides e de ácidos clorogênicos do endocarpo e do mesocarpo de jenipapo (Genipa americana L.) foram determinados nos estádios de maturação verde e maduro. Foram identificados ao todo, 17 compostos por HPLC-DAD-MSn (14 iridoides e 3 compostos fenólicos). O geniposídeo foi o composto majoritário encontrado no jenipapo verde, tanto no endocarpo quanto no mesocarpo. No jenipapo maduro, a genipina-gentiobiosídeo foi o composto majoritário encontrado no endocarpo. O teor de iridoides encontrado no jenipapo maduro reduziu mais de 90% em relação ao jenipapo verde. Esta grande redução nos teores de geniposídeo que ocorre com a maturação, pode explicar a não formação de pigmento azul no jenipapo maduro. Não foram detectados compostos fenólicos no jenipapo maduro. Uma nova metodologia de extração assistida por micro-ondas (MAE) para compostos fenólicos foi desenvolvida utilizando laranja cv. Pera [Citrus sinensis (L.) Osbeck], com diferentes teores de umidade: congelada e seca. O método MAE otimizado consistiu nos seguintes parâmetros: massa de 2,0 g para amostra congelada e de 0,2 g para amostra liofilizada, razão metanol:água de 8:2, volume de solvente de 25 mL, rampa de 30 s para atingir a temperatura de 50 °C, mantida por 10 min, utilizando como potência máxima 400 W e agitação de 200 rpm, seguido por um período de resfriamento de 10 minutos. Não houve diferença (P > 0,05) entre os teores de compostos fenólicos totais obtidos para ambas as amostras pelo novo método. Além disso, a MAE não provocou alterações no perfil de compostos fenólicos da amostra. O método de MAE desenvolvido apresentou ótima repetibilidade (RSD < 5%) e boa recuperação (> 92 %); foi 2,5 vezes mais rápido, consumiu 75 % menos solvente e utilizou metade da massa de amostra em relação à SLE, além de permitir a extração de várias amostras simultaneamente. Uma nova metodologia para extração de carotenoides foi desenvolvida. Para isso, três formas de extração foram avaliadas: MAE, agitação magnética e extração tradicional por maceração, utilizando manga cv. Haden (Mangifera indica L.) liofilizada como matriz. Além disso, também foi avaliada a eficiência de extração dos solventes acetona e da mistura acetato de etila:metanol (1:1 v/v). Os teores de carotenoides totais obtidos com acetato de etila:metanol foram superiores (P < 0,05) aos obtidos com acetona em todos os métodos avaliados. A comparação dos resultados obtidos pelos diferentes métodos mostrou que as micro-ondas não exerceram influência na extração, e também não provocaram alterações no perfil de carotenoides da amostra. Assim, o método utilizando apenas agitação magnética foi escolhido para ser validado utilizando os seguintes parâmetros: 0,2 g de amostra, 20 mL de acetato de etila:metanol, 200 rpm por 5 min a temperatura ambiente. O novo método apresentou elevada precisão (RSD < 3%) e exatidão (recuperação entre 101 % e 109 %). Além disso, o método foi cerca de 20 vezes mais rápido, consumiu 80 % menos amostra e 67 % menos solvente, quando comparado à extração por maceração / Abstract: The intake of foods rich in bioactive compounds, such as iridoids, phenolic compounds and carotenoids, is related to the decreased risk of chronic degenerative diseases. To identify the best sources of these compounds is necessary to develop efficient quantitative extraction methods which are able to preserve the original composition of the sample. The composition of iridoids and chlorogenic acids from endocarp and mesocarp of unripe and ripe genipap (Genipa americana L.) were determined by HPLC-DAD-MSn and a total of 17 compounds were identified (14 iridoids and 3 phenolic compounds). Geniposide was the major compound in both the endocarp and the mesocarp of unripe genipap, while genipin-gentiobioside was the major one in the endocarp of ripe genipap. A reduction above 90 % of the iridoid content was observed in ripe genipap in comparison with the unripe fruit. The large reduction in the levels of geniposide that occurred during ripening could explain the lack of formation of blue pigment in the ripe genipap. No phenolic compounds were detected in ripe genipap. A new microwave-assisted extraction (MAE) method for the extraction of phenolic compounds was developed and validated using orange cv. Pera [Citrus sinensis (L.) Osbeck] in physical states with different moisture contents: frozen and dried. The optimized MAE method consisted of the following parameters: 2.0 g of frozen sample or 0.2 g of lyophilized sample; methanol:water ratio of 8:2; solvent volume of 25 mL; microwave program: 30 s ramp to reach 50 °C, this temperature was maintained for 10 min using the maximal power of 400 W; and 200 rpm of agitation, followed by a cooling period of 10 min. No differences were found (P > 0.05) between the total phenolic contents of both liophilized and fresh samples using the new MAE method. In addition, MAE did not cause any alterations in the phenolic compound profile. The MAE method developed showed excellent repeatability (RSD < 5%) and good recovery (> 92 %). It was 2.5 faster than SLE; consumed 75 % less solvent and used half the mass of sample when compared with SLE. In addition, MAE allows the simultaneous extraction of several samples. A new methodology for the extraction of carotenoids was developed. For this purpose, three methods of extraction were evaluated using lyophilized mango cv. Haden (Mangifera indica L.) as food matrix: MAE, magnetic stirring and conventional extraction by maceration. In addition, the extraction efficiency of the solvents acetone and a mixture of ethyl acetate:methanol (1:1 v/v) was also evaluated. The contents of total carotenoids obtained with ethyl acetate:methanol were higher (P < 0.05) than those obtained with acetone in all the methods evaluated. The comparison of the results obtained from the different methods showed that the microwaves did not influence the carotenoid extraction and did not significantly change the profile of carotenoids of the sample. Therefore, the method using only magnetic stirring was chosen to be validated using the following parameters: 0.2 g of sample, 20 mL of ethyl acetate:methanol, 200 rpm for 5 min at room temperature. The new method for carotenoid extraction showed high precision (RSD < 3%) and accuracy (recoveries between 101 % and 109%). Moreover, the new method was almost 20 times faster, consumed 80% less sample and 67% less solvent than to the conventional extraction by maceration / Doutorado / Ciência de Alimentos / Doutora em Ciência de Alimentos

Thesis (MSc Food Sc )--Stellenbosch University, 2003. / ENGLISH ABSTRACT: The effect of drying conditions on the retention of harpagoside, a glucoside iridoid found in the indigenous Southern African plant Harpagophytum procumbens (Devil's Claw), was investigated. Additionally, the feasibility of using near infrared spectroscopy (NIRS) as a rapid and nondestructive analytical tool for the prediction of the iridoids harpagoside (HS), 8-p-coumaroyl harpagide (8pCHG) and moisture contents, as well as the HS:8pCHG ratio in Devil's Claw was also studied. Due to the purported medicinal activity ofthe tuberous secondary roots, Devil's Claw is of commercial interest with a well established export market in Germany. The stability of HS during processing has, however, never been investigated and the current study evaluated the effect of three drying methods (sun, tunnel and freeze-drying), as well as three tunnel-drying temperatures (40°C, 50°C and 60°C, dry bulb temperature) on HS content. Methanolic extracts (70% methanol-water) were analysed by high-performance liquid chromatography (HPLC) and a multiple extraction test (n = 12) indicated a 91.70% recovery for HS. The comparative drying results showed significant (P < 0.05) differences between tunnel and freeze drying on the one hand and sun-drying on the other, with the latter being the most detrimental technique. In comparing tunnel-drying temperatures, at a constant relative humidity (RH) of30%, drying at 40°C showed the lowest retention ofHS, differing significantly (P < 0.05) from that at 50°C. Drying at 60°C did not differ significantly (P > 0.05) from either of the other two conditions, but also resulted in lower retentions ofHS content than drying at 50°C. Overall, freeze-drying showed excellent results for HS retention (1.565% ± 0.394%; mean ± standard deviation), but economic constraints limit the commercial application of this drying method. Although further study is required to optimise tunnel-drying conditions, this method indicated acceptable and practical results (1.526% ± 0.396%), especially with conditions set to 50°C and 30% RH (1.750% ± 0.438%). Future research should concentrate on elucidating the degradation mechanisms of HS during processing, transport and storage of dried Devil's Claw root. The analysis of dried, ground Devil's Claw root by NIRS was referenced to HPLC analyses of the same methanolic extracts used in the drying studies. The spectra were generated on two different NIRS instruments by measuring diffuse reflectance of the powder whilst calibrations were performed by partial least squares regression. Both independent data set validation and full cross-validation were used to evaluate the performance and predictive abilities of the various models. Excellent moisture content (MC) calibrations were developed with a standard error of prediction (SEP) ofO.24% (range: 2.44% to 10.43%) and correlation coefficient (r) of 0.99. For the given MC range, the model performed very well against the standard error of laboratory (SEL) ofO.14%. Considering the sensitivity of the HPLC reference method (SEL = 0.035%), the HS content calibration (range: 0.693% to 2.244%) performed admirably with an SEP ofO.134%. The available range was, however, relatively small and future NIRS studies should incorporate a larger range if possible. The 8pCHG model (range: 0.069% to 0.290%) performed similarly, with a standard error of cross-validation of 0.028% and SEL of 0.007%. By inclusion of a second species, H. zeyheri, both the 8pCHG content and especially HS:8pCHG ratio (range: 1.84 to 34.48) calibrations were evaluated as possible methods of distinguishing between species. With 8pCHG values of ca. 0.9%, good separation was achieved for the first model but, contrary to previous studies, the ratio was found to be less successful. Principle component analysis of the spectra, however, showed great potential as a qualitative tool for this purpose. It is believed that NIRS can become an invaluable tool for the division of Devil's Claw into export classes, depending on HS content. This content is, however, dependant on the drying conditions used and drying parameters should be optimised to ensure the best possible quality and retention ofHS. / AFRIKAANSE OPSOMMING: Die invloed van drogingskondisies op die behoud van harpagosied, 'n iridoïed glikosied wat in die inheemse Suider-Afrikaanse plant, Harpagophytum procumbens (Duiwelsklou) voorkom, is ondersoek. Die uitvoerbaarheid van naby infra-rooi spektroskopie (NIRS) as 'n vinnige en niedestruktiewe analitiese metode vir die voorspelling van die inhoud van iridoïede, harpagosied (HS), 8-p-kumaroïel-harpagied (8pCHG) en vog, sowel as die HS:8pCHG verhouding in Duiwelsklou is ook ondersoek. Die beweerde medisinale aktiwiteit van die knolagtige sekondêre wortels van Duiwelsklou het gelei tot 'n gevestigde uitvoermark in Duitsland, en is dus van kommersiële belang. Die stabiliteit van HS gedurende prosessering is nog nooit ondersoek nie. Die huidige studie het die invloed van drie drogingsmetodes (son-, tonnel- en vriesdroging) en drie tonneldrogingstemperature (40°C, 50°C en 60°C, droëbol temperatuur) op die HS inhoud evalueer. Metanolies ekstrakte (70% metanol-water) is met hoë-druk vloeistof chromatografie ("HPLC") geanaliseer en 'n veelvoudige ekstraksie toets (n =12) het 'n 91.70% herwinning van HS getoon. Betekenisvolle (P < 0.05) verskille is gevind tussen tonnel- en vriesdroging enersyds en sondroging andersyds, met laasgenoemde die nadeligste metode. Die vergelyking van tonneldrogingstemperature, by 'n konstante relatiewe humiditeit (RH) van 30%, het getoon dat droging by 40°C die minste HS behou het en betekenisvol (P < 0.05) verskil het van droging by 50°C. Alhoewel die droging by 60°C nie betekenisvol verskil het van die ander twee kondisies nie, het dit laer HS waardes as droging by 50°C getoon. Vriesdroging het uitstekende resultate ten opsigte van die behoud van HS gelewer (1.565% ± 0.394%; gemiddelde ± standaard afwyking), maar ekonomiese beperkings plaas noodgedwonge limiete op die toepaslikheid van hierdie drogingsmetode. Verdere studie om tonneldrogingskondisies te optimiseer word is nodig (1.526% ± 0.396%), maar veral droging by 50°C en 30% RH (1.750% ± 0.438%) het prakties aanvaarbare resultate gelewer. Toekomstige navorsing moet daarop gemik word om die spesifieke degradasie meganismes van HS gedurende die droging, vervoer en opberging van Duiwelsklouwortel te verklaar. Die NIRS ontleding van gedroogde, gemaalde Duiwelsklou wortel is vergelyk met HPLC analises van dieselfde metanolise ekstrakte wat tydens die drogingstudies gebruik is. Twee verskillende NIRS instrumente is gebruik om spektra deur diffuse weerkaatsing in die poeier te meet en kalibrasies is daaropvolgend met gedeeltelike minste vierkante ("PLS") regressie gedoen. Beide onafhanklike data stel validasie en volle kruisvalidasie is gebruik om die prestasie en voorspellingsvermoë van die onderskeie modelle te evalueer. Uitstekende voginhoud (VI) kalibrasies met 'n standaard voorspellings fout (SVF) van 0.24% (klasinterval: 2.44% tot 10.43%) en 'n korrelasie van 0.99 is gevind. Die model het goed vergelyk met die standaard laboratorium fout (SLF) van 0.14%, oor die gegewe klasinterval. Met inagneming van die sensitiwiteit van HDVC as verwysingsmetode (SLF = 0.035%), het die HS inhoud kalibrasie (klasinterval: 0.693% tot 2.244%) aanvaarbaar gepresteer met 'n SVF van 0.134%. Die beskikbare klasinterval was egter relatief klein en toekomstige NIRS studies moet gebruik maak van 'n wyer klasinterval indien moontlik. Die 8pCHG model (klasinterval: 0.069% tot 0.290%) het soortgelyk gepresteer, met 'n standaard kruisvalidasie fout van 0.028% en SLF van 0.007%. Met die insluiting van monsters van 'n tweede spesie, H. zeyheri, is beide die 8pCHG inhoud en HS:8pCHG verhouding (klasinterval: 1.84 tot 34.48) kalibrasies as moontlike metodes vir onderskeiding tussen die spesies geëvalueer. Met'n 8pCHG inhoud van ca. 0.9% het die eerste model 'n goeie skeidingsvermoë getoon. In teenstelling met vorige studies egter, het die verhoudingsmodel minder sukses getoon. Hoofkomponent analise ("PCA") van die spektra was egter baie belowend as 'n kwalitatiewe onderskeidingsmetode. Volgens hierdie studie kan NIRS 'n waardevolle tegniek wees vir die verdeling van Duiwelsklou in uitvoer klasse op grond van HS inhoud. Hierdie inhoud is egter grotendeels afhanklik van die drogingskondisies en drogingsparameters wat verder verfyn behoort te word om 'n produk van die hoogste kwaliteit met behoud van HS te verseker.