L’ochratoxine A (OTA) est une mycotoxine issue du métabolisme secondaire des champignons filamenteux appartenant aux genres Penicillium et Aspergillus. Cependant, Aspergillus carbonarius est le majeur producteur de l’OTA sur les raisins. L’OTA a été retrouvée dans différents types de denrées alimentaires ainsi que lesurs produits dérivés. Le profil toxicologique de l’OTA due aux effets néfastes qu’elle présente sur la santé humaine et animale (effets hépatotoxiques, immunotoxiques, génotoxiques, tératogènes et cancérogènes) a conféré à cette mycotoxine une attention majeure auprès des instances internationales afin de limiter son occurrence. Ce projet est dédié pour trouver un moyen de lutte biologique, pouvant réduire l’OTA produite par A. carbonarius d’une part, et détoxifier les matrices alimentaires non conformes aux normes d’une autre part. La première stratégie était d’employer des huiles essentielles (cardamome, céleri, cannelle, taramira, origan, feuille de laurier, cumin, fenugrec, mélisse, menthe, sauge, anis, camomille, fenouil, romarin, romarin et thym) ainsi que des composés phénoliques extraits de plantes médicinales (feuille de laurier, cumin, fenugrec, mélisse, menthe, sauge, anis, camomille, fenouil, romarin et thym) afin d’évaluer leur effet sur la production de l’OTA dans le milieu SGM. Cette approche a été complétée par une étude moléculaire dans le but d’évaluer l’expression des gènes de biosynthèse de l’OTA (acpks, acOTApks et acOTAnrps) ainsi que les gènes de régulation (veA et laeA) chez A. carbonarius. Les résultats ont décelé que les huiles essentielles ont une activité fongicide plus élevée que celle des extraits phénoliques. Effectivement, les huiles essentielles du thym, de l’origan, du taramira, et de la cannelle ont bloqué complètement la croissance d’A. carbonarius. Cependant, les huiles essentielles du fenouil, de la cardamome, de l’anise, de la camomille, du céleri et du romarin ont réduit l’OTA sans autant affecter la croissance fongique. Le mode d’action de ces dernières a été mis en évidence en suivant l’expression des gènes acpks, acOTApks, acOTAnrps, veA et laeA, impliqués dans la biosynthèse de l’OTA chez A. carbonarius. Le gène acpks a été réprimée le plus (99.2%) quand A. carbonarius a été mis en culture avec 5 µL/mL du fenouil, entrainant ainsi une réduction de 88.9% de l’OTA. La deuxième stratégie était de développer un moyen de lutte biologique pouvant détoxifier les matrices alimentaires contaminées. Cette méthodologie a été développée suite à l’utilisation de sept souches d’actinobactéries (AT10, AT8, SN7, MS1, ML5, G10 et PT1), en évaluant leur capacité à métaboliser l’OTA, adhérer cette toxine à leur paroi membranaire ainsi que leur effet sur l’expression des gènes impliqués dans la biosynthèse de l’OTA chez A. carbonarius (acpks, acOTApks, acOTAnrps, veA et laeA). Les résultats ont montré que toutes les souches possèdent la capacité d’adhérer l’OTA à leur surface, notamment la souche SN7 qui a réduit 33% de l’OTA après 60 minutes d’incubation dans une solution PBS (Phosphate Buffer Solution) non nutritive. Les souches AT10 et SN7 ont métabolisé 51.94 et 52.68% de l’OTA ajoutée au milieu ISP2 (International Streptomyces Project-2) après 5 jours de culture à 28 °C. Cependant, les souches MS1, ML5 et G10 étaient les seules à avoir un effet sur l’expression des gènes de biosynthèse de l’OTA chez A. carbonarius. Effectivement les gènes acpks, acOTApks et acOTAnrps ont été réprimé respectivement de 37.1, 23.9 et 21% par MS1, de 39, 23 et 11.1% par ML5 et de 39, 18.3 et 11.1% par la souche G10. Ce projet a mis en valeur la capacité des extraits naturels (composés phénoliques et huiles essentielles) et des actinobactéries à prévenir d’une part la production de l’OTA et d’autre part réduire ses taux, sans pourtant affecter l’équilibre naturel ni engendrer l’apparition des débris toxiques dans les aliments traités. / Ochratoxin A (OTA) is a mycotoxin derived from the secondary metabolism of filamentous fungi belonging to the Penicillium and Aspergillus genera. However, Aspergillus carbonarius is the major producer of OTA on grapes. OTA has been detected in different types of foodstuffs as well as several products derived from these commodities. The toxicological profile of OTA and its adverse effects on human and animal health (hepatotoxic, immunotoxic, genotoxic, teratogenic and carcinogenic effects) has given this mycotoxin a major attention of international committees in order to limit its occurrence. The aim of this project was to develop biological techniques that can reduce OTA produced by A. carbonarius and detoxify non-compliant food matrices. The first strategy was achieved by using essential oils (cardamom, celery, cinnamon, taramira, oregano, bay leaf, cumin, fenugreek, melissa, mint, sage, anise, chamomile, fennel, rosemary and thyme) and phenolic compounds extracted from medicinal plants (bay leaf, cumin, fenugreek, melissa, mint, sage, anise, chamomile, fennel, rosemary, and thyme) to evaluate their effect on OTA production in SGM medium. This approach was complemented by a molecular study to evaluate the expression of the OTA biosynthesis genes (acpks, acOTApks and acOTAnrps) as well as the regulatory genes (veA and laeA) in A. carbonarius. The results revealed that essential oils had more significant fungicidal activity than phenolic extracts. Indeed, the essential oils of thyme, oregano, taramira, and cinnamon completely blocked the growth of A. carbonarius. However, essential oils of fennel, cardamom, anise, chamomile, celery and rosemary reduced OTA without affecting fungal growth. The mode of action of these essential oils has been demonstrated by evaluating the expression of acpks, acOTApks, acOTAnrps, veA and laeA genes in A. carbonarius. The expression of acpks was repressed the most (up to 99.2%) when A. carbonarius was cultured with 5 L / mL of fennel essential oil, resulting in a 88.9% of reduction in the OTA produced by this fungus. The second strategy was developed in order to detoxify contaminated food matrices. This methodology was achieved by using seven strains of actinobacteria (AT10, AT8, SN7, MS1, ML5, G10 and PT1), and evaluating their ability to metabolize OTA, adhere this toxin to their membrane walls and their effect on the expression of the genes involved in the biosynthesis of OTA in A. carbonarius (acpks, acOTApks, acOTAnrps, veA and laeA). The results showed that all strains were able to bind OTA to their surfaces, specially the SN7 strain which reduced 33% of OTA after incubation for 60 minutes in PBS (Phosphate Buffer Solution). The strains AT10 and SN7 metabolized 51.94 and 52.68% of the OTA added to the ISP2 medium (International Streptomyces Project-2) after 5 days of culture at 28° C. However, MS1, ML5 and G10 were the only strains to have an effect on the expression of the OTA biosynthesis genes in A. carbonarius. Indeed, acpks, acOTApks and acOTAnrps genes were repressed respectively by 37.1, 23.9 and 21% by MS1, 39, 23 and 11.1% by ML5 and 39, 18.3 and 11.1% by the strain G10. This project highlighted the power of natural extracts (phenolic compounds and essential oils) as well as strains of actinobacteria to prevent OTA production on one hand and to detoxify contaminated commodities on the other hand without altering the natural microbial balance.

Conselho Nacional de Desenvolvimento CientÃfico e TecnolÃgico / Taninos sÃo substÃncias polifenÃlicas que fazem parte dos metabÃlitos secundÃrios de vegetais. Tais compostos tÃm sido muito utilizados no curtimento de couro, como substitutos de fenol na produÃÃo de adesivos para madeira e em aplicaÃÃes mÃdicas. As principais fontes de taninos sÃo cascas de Ãrvores, e de acordo com alguns estudos, o caule de cajueiro possui grande potencial para ser utilizado como fonte de taninos. Nesse contexto, o objetivo deste trabalho foi otimizar a extraÃÃo de taninos da casca de cajueiro. Inicialmente, conduziram-se experimentos para avaliar o efeito de diferentes solventes sobre a extraÃÃo de taninos condensados (TC) e polifenÃis totais (PT). A tÃcnica de extraÃÃo utilizada foi a maceraÃÃo dinÃmica. TambÃm foi realizado um planejamento experimental para estudar a influÃncia de algumas variÃveis crÃticas (tempo, temperatura e razÃo lÃquido/sÃlido) sobre a extraÃÃo aquosa de PT e TC, alÃm do efeito sobre o teor de sÃlidos totais (TST). Testou-se o efeito da reutilizaÃÃo da casca e do solvente na extraÃÃo de TC e por fim foi avaliado o efeito da secagem por atomizaÃÃo e por liofilizaÃÃo sobre os extratos. De acordo com os resultados, os extratos obtidos com etanol 50% em Ãgua apresentaram maiores quantidades de PT e TC em relaÃÃo aos outros solventes. A variÃvel razÃo lÃquido/sÃlido foi o fator mais importante para todas as respostas, apresentando efeito positivo para PT e TC e efeito negativo para sÃlidos totais. TambÃm foi observado que hà uma relaÃÃo diretamente proporcional entre a temperatura e os teores de PT e TC. E o fator tempo sà foi significativo para TC, onde foi observado que o tempo de 100 minutos favoreceu a extraÃÃo. Avaliando a relaÃÃo das variÃveis, a melhor condiÃÃo encontrada para obter maiores teores de taninos foi: temperatura de 100ÂC, por 100 minutos e razÃo L/S 30:1. Nessa condiÃÃo, o teor de TC extraÃdos foi de 7,63 mg equivalente de catequina/ 100 mg casca. A utilizaÃÃo do mesmo solvente em atà duas extraÃÃes se mostrou viÃvel para a extraÃÃo de TC, ao contrÃrio da casca que, ao ser extraÃda uma segunda vez, proporcionou ganhos mÃnimos. A secagem por atomizaÃÃo se mostrou mais adequada para aplicaÃÃo industrial em comparaÃÃo com a liofilizaÃÃo. / Tannins are secondary plant metabolites with polyphenolic structure. These compounds have been used in leather tanning, also as replacement for phenol in adhesives manufacture for wood and medical applications. The main sources of these compounds are tree barks, and according to some studies, the stem cashew has great potential to be a source of tannins. In this context, the aim of this study was to optimize the tannins extraction from the cashew bark. Initially, experiments were performed to evaluate the effect of different solvents on the condensed tannins (CT) and total polyphenols (TP) extraction. Dynamic maceration was the technique used for extraction. An experimental design was conducted to understand the influence of some critical variables (time, temperature and liquid/solid ratio) on aqueous extraction of TP and CT, as well as the effect on total solids content (TSC). We tested the effect of bark and solvent reuse in the CT extraction and finally it was evaluated the effect of extracts freeze drying and spray drying. According to results, the extracts obtained with 50% ethanol in water showed higher amounts of TP and CT when compared to other solvents. The variable liquid/solid ratio, the most important factor for all responses, showed positive effect for TP and CT and negative effect on total solids. There is a directly proportional relationship between temperature and levels of TP and CT. Time factor was significant only for CT, where it was observed that 100 minutes favored the extraction. Evaluating the variables relationship, the best condition found for higher levels of tannins was: temperature of 100ÂC for 100 minutes and liquid/solid ratio 30:1. In this condition, the amount of CT extracted was 7.63 mg catechin equivalent / 100 mg bark. In extraction process, solvent used twice is feasible to extract CT. Unlike the bark, that obtained minimal gains in a second extraction. Spray drying was more suitable for industrial application compared to freeze drying.

Made available in DSpace on 2014-12-17T15:01:19Z (GMT). No. of bitstreams: 1 BrunoAAS.pdf: 4258750 bytes, checksum: 23e94d243172bf35909690337987df9d (MD5) Previous issue date: 2009-04-30 / Conselho Nacional de Desenvolvimento Cient?fico e Tecnol?gico / Solid substrate cultivation (SSC) has become an efficient alternative towards rational use of agro industrial wastes and production of value-added products, mainly in developing countries. This work presents the production and functional application results of phenolic extracts obtained by solid substrate cultivation of pineapple (Ananas comosus L.) and guava (Psidium guajava L.) residues associated to soy flour and bioprocessed by Rhizopus oligosporus fungus. Two experimental groups were tested: (1) 9g of fruit residue and 1g of soy flour (A9 or G9); (2) 5g of fruit residue and 5g of soy flour (A5 or G5). After SSC, 100ml of distilled water was added to each Erlenmeyer flask containing 10g of bioprocessed material in order to obtain the phenolic extracts. Samples were taken every two days for total phenolic concentration (TPC) and antioxidant capacity evaluation by DPPH test during 12-day cultivation. The 2-day and 10-d ay extracts were selected and concentrated by ebullition until 1/10 of original volume was reached. After that, both non-concentrated and concentrated extracts were evaluated for their antimicrobial activity against Staphylococcus aureus and Salmonella enterica and a-amylase inhibitory capacity. It was observed an inverse relationship between total phenolic concentration (TPC) and antioxidant capacity during the cultivation. Besides that, the concentrated pineapple samples after two days were able to inhibit both pathogens tested, especially S. aureus. Guava concentrated extracts after 2 days showed expressive inhibition against S. enterica, but negative results against S. aureus growth. When it comes to a-amylase inhibition, A9 extracts after 2 days, both concentrated or not, completely inhibited enzyme activity. Similar behavior was observed for G9 samples, but only for concentrated samples. It was shown that concentration by ebullition positively affected the enzymatic inhibition of G9 and A9 samples, but on the other side, decreased antiamylase activity of A5 and G5 samples / O cultivo semi-s?lido tem se tornado nos ?ltimos anos uma alternativa eficiente no aproveitamento de res?duos agroindustriais para a produ??o de compostos de alto valor agregado, sobretudo em pa?ses em desenvolvimento. Este trabalho apresenta resultados da produ??o e funcionalidade de extratos fen?licos obtidos atrav?s de cultivo semi-s?lido de res?duos de abacaxi (Ananas comosus L.) e goiaba (Psidium guajava L.) associados ao farelo de soja por meio do fungo Rhizopus oligosporus. Dois grupos experimentais foram estudados: (1) 9g de res?duo e 1g de farinha de soja (A9 ou G9); (2) 5g de res?duo e 5g de farinha de soja (A5 ou G5). Ap?s o cultivo semi-s?lido, 100 mL de ?gua destilada foram adicionados a cada frasco Erlenmeyer contendo 10g de material bioprocessado para obten??o dos extratos fen?licos n?o concentrados. Amostras foram tomadas ao longo do cultivo e avaliadas quanto ? concentra??o de fen?licos totais e capacidade antioxidante pelo teste DPPH ao longo de 12 dias de cultivo. Os extratos aquosos obtidos ap?s dois e dez dias de cultivo foram selecionados e concentrados at? redu??o de 1/10 do volume original. Em seguida, tanto os extratos concentrados quanto os n?o-concentrados desses pontos foram submetidos ? avalia??o da atividade antimicrobiana contra Staphylococcus aureus e Salmonella enterica e poder inibit?rio da enzima a-amilase. Durante o cultivo foi observada rela??o inversa entre a produ??o de fen?licos livres e capacidade antioxidante dos extratos. As amostras fen?licas concentradas dos res?duos de abacaxi ap?s dois dias de cultivo apresentaram capacidade de inibir o crescimento dos pat?genos testados, especialmente o S. aureus. Para o res?duo de goiaba, foi observado que os extratos fen?licos concentrados de ambos os grupos experimentais ap?s 2 dias de cultivo demonstraram expressiva inibi??o da Salmonella enterica, por?m, n?o apresentaram resultados positivos contra S. aureus. Quanto ? atividade anti-amilase, os extratos fen?licos do grupo experimental A9 ap?s dois dias de cultivo apresentaram capacidade de inibir completamente a a??o da enzima α-amilase. Comportamento similar foi detectado para as amostras do grupo experimental G9, por?m, apenas para as amostras concentradas. A concentra??o por ebuli??o dos extratos fen?licos nos grupos A9 e G9 favoreceu o aumento da inibi??o enzim?tica. Comportamento diferente foi observado nas amostras dos grupos A5 e G5 que apresentaram baixa atividade anti-amilase, possivelmente influenciada por modifica??es decorrentes da concentra??o por ebuli??o

Made available in DSpace on 2014-12-17T15:01:25Z (GMT). No. of bitstreams: 1 KatiaCB_DISSERT.pdf: 2815552 bytes, checksum: 5538a2db54454e17355a1ea7d2b7b8e5 (MD5) Previous issue date: 2011-02-15 / Coordena??o de Aperfei?oamento de Pessoal de N?vel Superior / Fruits are rich sources of bioactive compounds, including phenolic compounds. Tropical fruit cultivation is an important productive segment in Brazilian Northeast. Its industrialization generates solid wastes as co-products, with potential environmental impact. Considering the recognized bioactive content of fruit and its derivatives, this research has the objective of investigating acerola (Malpighia glabra L.), caj?-umbu (Spondia ssp), jambolan (Syzygium cumini) and pitanga (Eugenia uniflora) dried wastes obtained by spouted bed drier. It was analyzed the physical-chemical composition, solubility and microphotographic aspect of these dried wastes. Besides this, it was also evaluated the bioactive content, antioxidant activity and inhibitory activity against aamylase and a-glycosidase enzymes of water and ethanol (70%, 80% e 100% v/v) extracts prepared from fruit dried wastes, as well as their possible correlations. The dried fruit wastes showed high phenolic (606.04 to 3074.6 mg GAE eq/100 g sample), anthocyanin (478.7 mg/100 g for jambolan) and ascorbic acid (2748.03 mg/100 g for acerola) contents, as well as high antioxidant DPPH activity (14.27 a 36.30 mg Trolox eq/g sample). The extracts exhibited moderate to high a-amylase inhibition (23.97% a 76.58%) and high α-glycosidase inhibition, which 99.32% peak was reached for ethanol 70% pitanga extracts. It was also observed great positive correlation between phenolic content and DPPH activity (0.97 for acerola), anthocyanin (0.95 for jambolan) and α- glycosidase inhibition (0.98 for acerola). The α-glycosidase inhibition also correlated well with the antioxidant activity for all fruit extracts. The results show that these dried fruit wastes are valuable material for further applications as functional ingredients / As frutas possuem importantes compostos bioativos, dentre eles os compostos fen?licos. O cultivo de frutas tropicais ? um importante segmento produtivo do Nordeste Brasileiro, mas sua industrializa??o gera res?duos s?lidos como c?-produtos da atividade, pass?veis de gerar impacto ambiental. Tendo em vista o reconhecido conte?do bioativo de frutas e seus c?-produtos, objetivou-se estudar o baga?o desidratado da acerola (Malpighia glabra L.), caj?-umbu (Spondia ssp), jambol?o (Syzygium cumini) e pitanga (Eugenia uniflora) obtidos em secador de leito de jorro. A partir disso, foi analisada a composi??o f?sico-qu?mica, solubilidade e aspecto microfotogr?fico dos baga?os secos. Al?m disso, foi avaliada a concentra??o de compostos bioativos, atividade antioxidante e atividade inibit?ria contra as enzimas α-amilase e α-glicosidase do extrato aquoso e extratos etan?licos (70%, 80% e 100% v/v) preparados a partir dos baga?os desidratados, bem como suas correla??es. Os p?s de fruta apresentaram elevado teor de compostos fen?licos (606,04 a 3074,6 mg GAE eq/100 g amostra), antocianinas (478,7 mg/100 g para jambol?o) e ?cido asc?rbico (2748,03 mg/100 g), bem como expressiva atividade antioxidante medida pelo m?todo DPPH (14,27 a 36,30 μg Trolox eq/g amostra). Os extratos foram capazes de expressar inibi??o da α-amilase de moderada a alta (23,97% a 76,58%) e expressiva inibi??o da α-glicosidase, cujo m?ximo de 99,32% foi alcan?ado pelos extratos etan?licos 70% da pitanga. Foi observada correla??o positiva entre o teor fen?lico e atividade antioxidante (0,97 para acerola), antocianinas (0,95 para jambol?o), inibi??o da α-amilase (0,99 para jambol?o) e inibi??o da α-glicosidase (0,93 para pitanga) e dessas duas ?ltimas com a atividade antioxidante para todos os frutos estudados. Os dados apresentados demonstram que os res?duos desidratados constituem valioso material de estudo para aplica??es como ingredientes funcionais