Made available in DSpace on 2016-04-28T20:17:26Z (GMT). No. of bitstreams: 1 2006 - Rodrigo Alcantara de Carvalho.pdf: 803403 bytes, checksum: 7a35f34784a3f9cb58b545f52959a6a0 (MD5) Previous issue date: 2006-09-22 / The world wide contamination of foods and byproducts with mycotoxins is a problem of extreme importance. Mycotoxins are secondary metabolites produced by fungi that have adverse effects to human and animal health. The fumonisins and aflatoxins are the mycotoxins of greatest agro-economic importance due to the damage brought to animal production. Important steps in the synthesis of these mycotoxins are described, as such as their metabolism and adverse effects. This work evaluates the determination of the biomarkers sphinganin (Sa) and sphingosin (So) in poultry serum as a diagnose method for fumonisin exposition, and compares the obtained data with ration analysis. Of the evaluated animals only 49% exhibited 2 sphingolipids (Sa and So), 53,04% exhibited only So and 2,04% exhibited only Sa. It was detected at ration the presence of fumonisin B1 at concentration of 374,92μg.kg-1 and aflatoxins B1, B2, and G1 at concentrations of 440,99μg.kg-1, 169,92μg.kg-1 and 494,58μg.kg-1, respectively. It was possible to partially establish the grade of intoxication of the poultry and to verify that the sensibility of the biomarker can be affected by the presence of fumonisin such as aflatoxins. / A contamina??o mundial de alimentos e subprodutos com micotoxinas ? um problema de extrema import?ncia. As micotoxinas s?o metab?litos secund?rios produzidos por fungos, que possuem efeitos adversos ? sa?de humana e animal. As fumonisinas e aflatoxinas s?o as micotoxinas de maior import?ncia agro-econ?mica pelos preju?zos acarretados ? produ??o animal. Passos importantes na s?ntese destas micotoxinas s?o descritos, assim como seu metabolismo e efeitos adversos. Este trabalho avalia a determina??o dos biomarcadores esfinganina (Sa) e esfingosina (So) no soro de frangos de corte como m?todo de diagn?stico para exposi??o ?s fumonisinas e compara os dados obtidos com a an?lise da ra??o. Dos animais avaliados apenas 49% exibiram quantidades detect?veis de esfingolip?deos no soro, dentre estes, apenas 44,9% exibiram os 2 esfingolip?deos (Sa e So), 53,06% exibiram somente So e 2,04% exibiram somente Sa. Foi detectada na ra??o a presen?a de fumonisina B1 na concentra??o de 374,92μg.kg-1 e de aflatoxinas B1, B2 e G1 nas concentra??es de 440,99μg.kg-1, 169,92μg.kg-1 e 494,58μg.kg-1, respectivamente. Foi poss?vel estabelecer parcialmente o grau de intoxica??o dos frangos e verificar que a sensibilidade do biomarcador pode ser afetada tanto pela presen?a de fumonisina quanto de aflatoxina.

Apesar da perda econômica, representada pela contaminação por fungos toxigênicos em castanhas-do-Brasil, um importante produto extrativista da região Norte do Brasil, estudos realizados ainda são incipientes quanto ao controle da contaminação por fungos aflatoxigênicos utilizando métodos como radiação gama (R.G) e, sobretudo, feixe de elétrons (F.E). Esses fatos motivaram a presente pesquisa teve como objetivo avaliar os efeitos da radiação gama e da aplicação de feixe de elétrons em amostras de castanha-do-Brasil inoculadas artificialmente com Aspergillus flavus. Para atingir tal objetivo, foram estudadas 50 amostras de castanha-do-Brasil, previamente, inoculadas com suspensão de esporos de A. flavus e, posteriormente, incubadas a 30 ºC em ambiente com umidade relativa controlada a 93 %. Após 15 dias de incubação, as amostras foram divididas em 5 grupos que receberam as seguintes doses de radiação: controle (0 kGy), 5 e 10 kGy F.E, 5 e 10 kGy R.G. A micobiota foi realizada através de diluição seriada, com semeadura em superfície utilizando ágar batata. Os resultados demonstraram que o tratamento com F.E utilizando a dose de 5 kGy e 10 kGy resultou na redução de crescimento de A. flavus em 74% (37/50) e 94% (47/50) das amostras. Quanto às amostras tratadas com R.G na dose de 5 kGy e 10 kGy não ocorreu crescimento fúngico em 92% (46/50) e 100% (50/50) delas. A pesquisa de aflatoxinas mostrou que doses de F.E de 5 kGy e 10 k Gy reduziram os níveis de AFB1 em 53,32% e 65,66%, respectivamente. Por sua vez, a aplicação de raios gama nas doses de 5 e 10 kGy reduziu os níveis das toxinas em 70,61% e 84,15 % respectivamente. Tal resultado pode ser atribuído a maior penetrabilidade da radiação gama. Análise sensorial demonstrou maior aceitação das amostras irradiadas com F.E e R.G na dose de 10 kGy. Concluímos que, apesar de a análise sensorial ter demonstrado perda de algumas características organolépticas, ambos os processos de radiação foram eficazes na redução da contagem de A. flavus e dos níveis de aflatoxinas. / Despite the economic loss represented by contamination by toxigenic fungi in Brazil nuts, a major product of extractive Northern of Brazil, studies are still preliminary as the control of contamination aflatoxigenic fungal using methods such as gamma radiation (G.R) and mainly, electron beam (E.B). These facts motivated this research, which aimed to evaluate the effects of gamma radiation and application of electron beam in samples of Brazil nut artificially inoculated with Aspergillus flavus. This goal, we were studied 50 samples of the Brazil nut previously inoculated with spores of A. flavus and subsequently incubated at 30 °C in relative humidity controlled at 93%. After incubation, period of 15 days, the average water activity of the samples was 0.80, the samples were divided into 5 groups that received the following doses of radiation: control (0 kGy), 5 and 10 kGy 5 E.B and G.R. The mycobiota was performed by serial dilution, plated on surface using potato dextrose agar. The results demonstrated that treatment with E.B using a dose of 5 kGy and 10 kGy resulted in reduced growth of A. flavus in 74% (37/50) and 94% (47/50) of samples. The samples treated with G.R at the dose of 5 kGy and 10 kGy no fungal growth occurred in 92% (46/50) 100% (50/50) of. The study of aflatoxins showed that doses of E.B of 5 kGy and 10 kGy reduced levels of AFB1 at 53.32% and 65.66% respectively. The application of gamma rays at doses of 5 and 10 kGy reduced levels of toxins in 70.61% and 84.15% respectively. This result may be attributed to higher penetrability of gamma radiation. Sensory analysis showed greater acceptance of the judges for the samples irradiated with E.B and G.R at the dose of 10 kGy. We concluded that although sensory analysis have demonstrated some loss of organoleptic characteristics, both processes of radiation were effective in reducing the count of A. flavus and aflatoxin contamination.

Submitted by Jorge Silva (jorgelmsilva@ufrrj.br) on 2018-03-28T19:08:17Z No. of bitstreams: 1 2017 - Yuri Duarte Porto.pdf: 1910618 bytes, checksum: 969c177c9241195f2d92cb156f231abb (MD5) / Made available in DSpace on 2018-03-28T19:08:17Z (GMT). No. of bitstreams: 1 2017 - Yuri Duarte Porto.pdf: 1910618 bytes, checksum: 969c177c9241195f2d92cb156f231abb (MD5) Previous issue date: 2017-04-06 / CAPES / Corn is one of the most cultivated cereals in Brazil and it is susceptible to contamination by mycotoxins that are toxic secondary metabolites produced by fungi. An adoption of quality management systems during a corn production chain is essential to ensure food safety concerning mycotoxin contamination. However, there is a possibility that these recognized carcinogenic metabolites may already be found in the matrix. Hence the search for alternative solutions capable of reducing contamination to safe levels by the application of emerging technologies has been intense. The methods of decontamination of foods must to obey some premises as: inactivate, destroy or remove the toxins; do not produce others toxic waste; maintain the nutritional value and acceptability of the product. Ozone, which meets almost all of these characteristics, is recognized by the Food and Drug Administration as a GRAS (Generally Recognized as Safe) product for use as a sanitizer in food processing. Since then its use has been investigated in several types of food, including cereals contaminated by mycotoxins. Corn canjiquinha is a cultural food easily accessible by Brazilian consumers at low prices, as well as posing a potential risk of exposure due to contamination with aflatoxins. In this study, aliquots of canjiquinha samples were inoculated with concentrations of 106 CFU/g conidia of Aspergillus spp. and Fusarium spp. previously isolated from this food. Other aliquots were contaminated with aflatoxins B1, B2, G1 and G2 at a concentration of 50 ?g/kg for each. The application of gaseous ozone was tested in different combinations of exposure time, ozone concentration and canjiquinha mass, being these independent variables investigated. After treatment by ozonation, to evaluate the effects on aflatoxin concentrations, the samples were analyzed by high-performance liquid chromatography with fluorescence detection (HPLC-FD system) with pre-column derivatization (C18) by trifluoroacetic acid (TFA). The analytical method used in this study was optimized and parameters such as accuracy, precision, linearity, selectivity, limits of detection and limits of quantification of the analytical method were previously evaluated. Additionally, aflatoxin quantification tests were carried out on commercial samples of corn kernels. The method was adequate and presented recovery values within the range of 80-110% with a coefficient of variation of less than 15%, with detection and quantification limits equal to 0.8 and 3.6 ?g/kg, respectively, for each of the aflatoxins. Isolation of fungi was carried out according to the criteria of the Brazilian Ministry of Agriculture. Thus, the efficacy of gaseous ozonation on aflatoxin levels B1, B2, G1 and G2 and on microbial contamination of canjiquinha grains showed reductions of up to 57% in aflatoxin levels. The total fungal count had a reduction of about 3.0 log cycles UFC/g and the total counts of mesophiles were reduced to undetectable levels. These results demonstrated that ozonation is an effective alternative to reduce microbial contamination and the concentration of aflatoxins in corn kernels and, consequently, can improves the safety of this product. / O milho ? um dos cereais mais cultivados no Brasil e est? exposto ? contamina??o por micotoxinas que s?o metab?litos t?xicos secund?rios produzidos por fungos. A ado??o de sistemas de gest?o da qualidade durante a cadeia de produ??o de milho ? essencial para garantir a seguran?a do alimento quanto ? contamina??o por micotoxinas. Contudo, h? possibilidade desses metab?licos reconhecidamente carcinog?nicos j? encontrarem-se na matriz. Da? a busca por solu??es alternativas capazes de reduzir a contamina??o para n?veis seguros pela aplica??o de tecnologias emergentes tem sido intensa. Os m?todos de descontamina??o de alimento devem obedecer algumas premissas como: inativar, destruir, ou remover as toxinas; n?o produzir res?duos t?xicos; manter o valor nutricional e a aceitabilidade do produto. O oz?nio atende quase todas estas caracter?sticas, ? reconhecido pelo United States Food and Drug Administration (FDA) como uma subst?ncia GRAS (Geralmente Reconhecido como Seguro) para uso como sanitizante no processamento de alimentos. Desde ent?o sua utiliza??o tem sido pesquisada em diversos tipos de alimentos, incluindo cereais contaminados por micotoxinas. A canjiquinha de milho ? um alimento cultural de f?cil aquisi??o pelo consumidor brasileiro pelo baixo pre?o, al?m de representar um potencial risco de exposi??o por contamina??o com aflatoxinas. Neste estudo, al?quotas de amostras de canjiquinha foram inoculadas concentra??es de 106 UFC/g de con?dios de Aspergillus spp. e Fusarium spp. previamente isolados do alimento. Outras al?quotas foram contaminadas com aflatoxinas B1, B2, G1 e G2 na concentra??o de 50 ?g/kg para cada. A aplica??o de oz?nio gasoso foi testada em diferentes combina??es de tempo de exposi??o, concentra??o do oz?nio e massa de canjiquinha, sendo essas as vari?veis independentes pesquisadas. Ap?s tratamento por ozoniza??o, para avalia??o dos efeitos sobre as concentra??es de aflatoxinas, as amostras foram analisadas por cromatografia l?quida de alta efici?ncia com detec??o por fluoresc?ncia (sistema CLAE-DF) com derivatiza??o pr?-coluna (C18) por ?cido trifluoroac?tico (TFA). O m?todo anal?tico utilizado neste estudo foi otimizado e par?metros como a exatid?o, precis?o, linearidade da faixa de trabalho, seletividade, limites de detec??o e limites de quantifica??o do m?todo anal?tico foram previamente avaliados. Adicionalmente efetuaram-se ensaios para quantifica??o das aflatoxinas em amostras comerciais de canjiquinha de milho. O m?todo mostrou-se adequado e apresentando valores de recupera??o dentro da faixa de 80-110% com coeficiente de varia??o menor que 15%, sendo os limites de detec??o e quantifica??o iguais a 0,8 e 3,6 ?g/Kg, respectivamente, para cada uma das aflatoxinas. O isolamento de fungos foi realizado de acordo com as normas do Minist?rio da Agricultura. Assim a avalia??o da efic?cia da ozoniza??o gasoso sobre os n?veis de aflatoxinas B1, B2, G1 e G2 e sobre a contamina??o microbiana dos gr?os de canjiquinha apresentou redu??es de at? 57% nos n?veis de aflatoxinas. A contagem de fungos totais teve uma redu??o de cerca de 3,0 ciclos log UFC/g e a contagem total de mes?filos foram reduzidas a n?veis n?o detect?veis. Estes resultados demonstraram que a ozoniza??o ? uma alternativa eficaz para reduzir a contamina??o microbiana e a concentra??o de aflatoxinas em canjiquinha de milho e, consequentemente, melhora a seguran?a desse produto.

Made available in DSpace on 2017-05-12T14:47:04Z (GMT). No. of bitstreams: 1 Cristiane _Lurdes Paloschi.pdf: 1204212 bytes, checksum: 0429a0e71141d849cf2df342c98d4d01 (MD5) Previous issue date: 2014-07-15 / Fundação Araucária / This work aimed to study the effects of infestation of insects Sitophilus zeamais on the physicochemical and microbiological quality in corn kernels during storage. For the experiment, samples of hybrid maize (Zeamays) were collected from cultivar Dow 2B512Hx, grown in western Paraná, harvested in June / July 2013. For storage, the grains had water contents below 15% bs, and about 300g of them were stored in glass containers closed with tissue type Voil to facilitate gas exchange, with a total of 40 containers stored in a BOD incubator at a temperature of 27±1 °C for 180 days. From all the recipients, 20 were infested with 20 adult insects of Sitophilus zeamais and the other 20 were kept uninfested. Every 45 days, samples were taken and physical and chemical analyzes were carried out (grain weight loss, population variation of insects, classification of corn, mass of 1000 seeds, water content, ash, oil, protein and carbohydrates and dietary fiber total), as well microbiological analyzes (counting of filamentous fungi and identification of aflatoxin-producing fungi) and quantitation of aflatoxin (B1, B2, G1 and G2). Exploratory analysis of the response variables was done by use of the analysis of variance (ANOVA) and mean comparison test was performed by Tukey's test with a significance level of 0.05. The final grain weight in dry material with insects reduced in the course of time, what increased the losses in grain mass and showed statistically significant differences. The population of insects and the classification of maize also showed statistically significant differences. It was observed that when storage time increased, the population of insects also increased and the percentage of whole grains reduced for both storage conditions. The content of water, protein and filamentous fungi incidence increased in grains initially infested during storage, and the 1000 grains weight, the content of carbohydrates and total dietary fiber content in these grains decreased compared to uninfected grains. The incidence of aflatoxin was not detected in maize kernels during the studied storage period. Finally, from these results, it is possible to conclude that the longer the period of contact of insects with the largest grains, larger the damages and losses to producers are. However, S. zeamais did not behave as a mechanical vector of the fungus Aspergillus ssp., with no correlation between the incidence of filamentous fungi and the level of aflatoxins (B1, B2, G1 and G2). / Este trabalho teve como objetivo estudar os efeitos da infestação dos insetos Sitophilus zeamais sobre a qualidade físico-química e microbiológica em grãos de milho durante o armazenamento. Para a condução do experimento, foram coletadas amostras de grãos de milho (Zeamays) híbrido da cultivar Dow 2B512Hx, cultivados na região Oeste do Paraná, colhidas em junho/julho de 2013. Para o armazenamento, os grãos se encontravam com teores de água abaixo de 15% b.s sendo armazenados cerca de 300 g em recipientes de vidros fechados com tecido tipo Voil, a fim de facilitar as trocas gasosas, totalizando 40 recipientes armazenados em câmera incubadora B.O.D à temperatura de 27±1 °C durante 180 dias. Do total de recipientes, 20 foram infestados com 20 insetos adultos de Sitophilus zeamais e os outros 20 foram mantidos sem infestação. A cada 45 dias, foram retiradas as amostras e realizadas análises físico-químicas (perda de massa dos grãos, variação populacional dos insetos, classificação do milho, massa de 1000 grãos, teor de água, cinzas, óleo, proteína e carboidratos e fibra alimentar total), microbiológicas (contagem de fungos filamentosos e identificação dos fungos produtores de aflatoxinas) e de quantificação de aflatoxinas (B1, B2, G1 e G2). A análise exploratória das variáveis respostas foi analisada pelo emprego da análise de variância (ANOVA) e o teste de comparação de médias foi realizado pelo teste de Tukey com nível de significância de 0,05. O peso final dos grãos em matéria seca com insetos reduziu ao longo do tempo, aumentando as perdas na massa dos grãos, apresentando diferenças estatísticas significativas. A população de insetos e a classificação do milho também apresentaram diferenças estatísticas significativas. Observou-se que, conforme aumentava o tempo de armazenamento, aumentava a população de insetos e reduzia a percentagem de grãos inteiros para ambas as condições de armazenamento. O teor de água, proteína e a incidência de fungos filamentosos aumentaram nos grãos inicialmente infestados ao longo do armazenamento e o peso de 1000 grãos e o teor de carboidratos e fibra alimentar total dos grãos diminuiu nestes grãos em relação aos grãos que não infestados. A incidência de aflatoxinas não foi detectada nos grãos de milho durante o período de armazenamento analisado. Pode-se concluir, com base nesses resultados, que quanto maior o período de contato dos insetos com os grãos maiores são os danos causados e os prejuízos para os produtores, porém o S. zeamais não se comportou como um vetor mecânico do fungo Aspergillus ssp., não havendo correlação entre a incidência de fungos filamentosos e o nível de aflatoxinas (B1, B2, G1 e G2).

Os objetivos foram: identificar a microbiota fúngica e a ocorrência de aflatoxinas em cultivar de amendoim, identificar a microbiota fúngica do solo e correlacionar os resultados obtidos com os níveis de atividade de água encontrados. As amostras (solo e amendoim) foram provenientes de: Jaboticabal, Rosália, Tupã e Cafelândia (SP). A microbiota fúngica do solo revelou que o fungo do gênero Penicillium foi o mais frequente (52,1 %) nas quatro regiões durante as duas coletas (após a emergência das plantas e duas semanas antes do arranquio). Dentro do gênero Aspergillus, A. flavus foi a espécie mais frequente (13,5 %). A microbiota fúngica dos grãos e das cascas, das quatro regiões, nas duas coletas (duas semanas antes do arranquio e após o arranquio), demonstrou maior frequência de isolamento do fungo do gênero Fusarium (70,2 %). Do gênero Aspergillus, a espécie A. flavus foi a mais frequente (9,8 %). A análise de aflatoxinas revelou a presença de aflatoxinas em 5 % das amostras de grãos analisadas. Nas cascas, 13,75 % das amostras apresentaram contaminação por aflatoxinas. / The objectives were: to identify the mycoflora and the occurrence of aflatoxins in a peanut variety, identify the soil mycoflora and compare the acquired results with the levels of water activity found. The samples (peanut and soil) were collected from: Jaboticabal, Rosália, Tupã and Cafelândia (SP/Brazil). The mycoflora of the soil revealed that the genus Penicillium was the most frequent (52.1 %) in the four regions during the two trials (after the emergence of the plants and two weeks before the uprooting). Within the genus Aspergillus, A. flavus was the most frequent specie (13.5 %). The mycoflora of the kernels and peanut hulls, from the four regions in the two trials (two weeks before the uprooting and after the uprooting), demonstrated greater frequency of isolation of the genus Fusarium (70.2 %). Within the genus Aspergillus, A. flavus was the most frequent specie (9.8 %). The analysis of aflatoxins revealed the presence of aflatoxins in 5 % of the kernels samples analyzed. In peanut hulls, 13.75 % of the samples presented aflatoxin contamination.

Graduation date: 1992

Since the discovery of mycotoxins in food, the thrust of biochemical and toxicological research has been carried out on animals which has proven to be uncoordinated and not easily extrapolated to humans. Over the last decade, there have been increasing pressures to review and reduce the use of animals in experimental toxicological studies. Consequently in this study aflatoxin B1 (AFB1), zearalenone (Zea) and their epoxide derivatives have been evaluated using in vitro assays. The HepG2, A549 and Hela cell lines were used for assessing the cytotoxicity, effects on cellular metabolism and sites of action of AFB1, Zea and their derivatives. The cytotoxicity of these mycotoxins was evaluated using the methylthiazol tetrazolium (MTT) reduction assay. Cells, treated with mycotoxins were prepared for transmission electron mlcroscopy (TEM), immunocytochemistry (ICC), scanning electron microscopy (SEM), confocal and light microscopy. From the cytotoxicity assay it was found that the epoxide derivatives were more toxic than the parent toxin when exposed to HepG2 cells with no significant differences in toxicity levels in A549 and Hela treated cells. Both epoxide derivatives displayed a regression of hepatoma cell proliferation at high doses (25ug/ml) while lower concentrations (<12.5ug/ml) enhanced cell growth. Microscopy analyses showed distinct cellular alterations. When exposed to AFB1 (12.5ug/ml) hepatoma cells showed prominent ultrastructural alterations such as areas of cytoplasmic lysis and increased numbers of secondary lysosomes while cells exposed to Zea (l2.5ug/ml) displayed numerous ovoid mitochondria and proliferation of rough endoplasmic reticulum which is indicative of enhanced protein synthesis. The presence of label in toxin treated cells is suggestive of the effects of these mycotoxins. Such cellular changes may lead to altered metabolism and cell function. / Thesis (M.Med.)-University of Natal, Durban, 1996.

Aflatoxin B1 (AFB1) and Fumonisin B1 (FB1), potentially cytotoxic and carcinogenic mycotoxins are common contaminants of agricultural commodities in South Africa and thus could be detrimental to the human immune system. Many of the cytotoxic effects of AFB1 require its bioactivation to an epoxide, which will bind covalently to macromolecules to form protein and DNA adducts. Fumonisin B1 is a competitive inhibitor of sphingosine and sphinganine N aceyltransferase, which are key components in the pathways for sphingolipid biosynthesis. Accumulation of free sphingoid bases, which are both cytotoxic and mitogenic, could provide a plausible explanation for the toxicity and carcinogenicity of FB1. The cytotoxic effects of AFB1 and FB1 on normal human lymphocytes, individually and in combination were assessed using the methylthiazol tetrazolium (MTT) bioassay. Two different methods of treatment were used, the treatment of isolated normal human lymphocytes for 12, 24, 48, 72 and 96 hours and whole blood treated for 12 hours. Flow cytometry and fluorescent microscopy were used to determine whether AFB1 and FB1 (5uM and 50uM), individually or in combination, were capable of inducing apoptosis, necrosis or nuclear fragmentation in isolated lymphocytes and whole blood treated for 12 hours. DNA damage was evaluated using the comet assay. The results showed that AFB1routinely induced higher levels of cytotoxicity in isolated lymphocytes than FB1. In the combination treatment, the mitogenic properties of FB1 appeared to partially counteract the cytotoxic effect exerted by AFB1. When whole blood was treated with the same concentration and ratio of toxin, FB1 was shown to be more cytotoxic than AFB1. The combination treatment of whole blood was shown to be cytotoxic in a dose dependent manner. The toxins appeared to exert a greater cytotoxic effect, when treated in combination than individually at higher concentrations. Aflatoxin B1 induced increased levels of apoptosis and necrosis in isolated lymphocytes while treatment with the FB1 resulted in increased levels of apoptosis at both concentrations. Treatment with the combination also resulted in increased levels of apoptosis. The levels of apoptosis were reduced in whole blood lymphocytes when compared to isolated lymphocytes. However, treatment with AFB1 and FB1 resulted in increased levels of apoptosis. Both AFB1 and FB1 are capable of inducing nuclear fragmentation. Treatment with FB1 (5uM and 50uM) resulted in greater degree of fragmentation than AFB1. The most nuclear fragmentation was induced by the 5uM combination treatment. The 50uM combination treatment of isolated lymphocytes induced the most DNA damage. As both toxins are common contaminants and have been known to coexist, this could be a potential area of concern for public health. / Thesis (M.Med.)-University of KwaZulu-Natal, 2004.

Thesis (D.Tech.: Biotechnology)-Dept. of Biotechnology, Durban Institute of Technology, 2005 xx, 175, [14] leaves : ill. ; 30 cm / Previous studies indicate that a mutation in the non-oncogenic p53 gene is epidemiologically linked to human HCC (Ozturk, 1991; Chan et al., 2003). Hsu et al. (1991) found this link in Chinese, South African and Asian patients and Hollstein et al. (1993) found the same gene mutation in Taiwanese patients. The incidence of these aberrations is reported to be about 20- 50% in HCC’s (Kishimoto et al., 1997). There is sufficient evidence to indicate that carotenoids in addition to their well known antioxidant properties (Paiva and Russel, 1999), also affect intercellular communication, immune responses, neoplastic transformations and growth control, and cellular levels of enzymes that detoxify carcinogens (Zhang et al., 1991; Brockman et al., 1992; Pryor et al., 2000). To date studies carried out have used the rat (Foote et al., 1970; Gradelet et al., 1998) and the mule duckling model (Cheng et al., 2001) to show the protective effect of these carotenoids against AFB1 exposure. Of the well known carotenoids, lycopene and beta- carotene occur in abundance in fruits and vegetables and are safe for human consumption. Aflatoxin B1 frequently induces mutations of the p53 gene which is linked to HCC. Although there is much evidence from epidemiological studies linking the beneficial aspects of carotenoids to the prevention of cancer, the cellular and molecular mechanisms need to be understood in order to implement large scale intervention strategies to prevent AFB1 induced carcinoma. The use of chemical or dietary interventions to alter the susceptibility of humans to the actions of carcinogens and to block, retard or reverse carcinogenesis is an emerging chemoprotective strategy for disease prevention (Abdulla and Gruber, 2000; Kensler et al., 2003; Bingham and Riboli, 2004). Chemoprotection by natural products involves maintaining cellular integrity, preventing DNA alterations, activation of p53 suppressor protein and apoptosis. The aim of this study was thus to investigate the cellular and molecular mechanisms by which beta-carotene and lycopene may prevent the AFB1-induced toxic changes in human hepatocytes. In order to achieve this aim, the following objectives were set out: i. To optimise an in vitro system for the evaluation of AFB1 damage to cultured hepatocytes. ii. To determine the biochemical protection offered by beta-carotene and lycopene to AFB1-exposed hepatocytes, by measuring the mitochondrial activity, cell viability and ROS levels using appropriate enzyme assays and flow cytometry. iii. To determine the cellular protection offered by beta-carotene and lycopene to AFB1-exposed hepatocytes, by studying the morphological changes at the structural and ultrastructural levels using phase contrast light and electron microscopy respectively. iv. To determine the molecular protection offered by beta-carotene and lycopene to AFB1-exposed hepatocytes, by detecting apoptotic bodies as genomic markers and measuring the levels of p53 protein and AFB1-N7-guanine adducts produced.

Le micotossine sono metaboliti tossici prodotti da funghi in grado di svilupparsi sulle derrate alimentari. Diverse strategie sono state considerate per risolvere questo problema studiando la crescita di funghi/produzione di micotossine. Questa tesi è focalizzata sullo sviluppo/validazione di modelli matematici per prevedere la contaminazione di micotossine (deossinivalenolo, fumonisine e aflatossine) in cereali (mais/grano) sulla base di dati meteorologici. Il primo capitolo fornisce un’introduzione sulla teoria dei modelli e sui pat-sistemi modelizzati. Il secondo si concentra sulla presenza di tricoteceni e zearalenone nel frumento coltivato in Italia. Nel cap.3 sono stati confrontate le differenze predittive di modelli empirici/meccanicistici per la contaminazione di deossinivalenolo nel grano. Nel cap.4 è stata descritta la contaminazione da fumonisine e aflatossine in mais coltivato in Italia. I capitoli 5 e 6 analizzano il pato-sistema mais-Aspergillus flavus, il primo si concentra sulla sporulazione di A. flavus , il secondo sullo sviluppo di un modello per prevedere la contaminazione da aflatossina. Un altro modello meccanicistico per prevedere la presenza di fumonisina nel mais è descritto nel cap.7. L'ultimo capitolo riassume l'attività svolta nel progetto europeo MYCORED in cui sono stati coinvolti diversi paesi in tutto il mondo che hanno fornito i dati necessari per la validazione dei modelli. / Mycotoxin are toxic secondary metabolite produced by fungi able to colonize crops and thus posing a potential menace to human/animal health. Several strategies have been considered to mitigate the problem studying the variables related to mould growth and mycotoxin production. This thesis focuses on the development and validation of mechanistic models to predict mycotoxins (deoxinivalenol, fumonisins and aflatoxins) contamination in cereals (maize/ wheat) based on meteorological data. The first chapter introduce modelling theory, and patho-systems analysed. Chapter 2 focuses on trichothecenes and zearalenone occurrence in wheat produced in Italy. Predictive performance of empirical and mechanistic models for deoxnivalenol contamination in wheat were discussed in chapter 3. Chapter 4 described fumonisins and aflatoxins occurrence in maize grown in Italy. Chapters 5 and 6 analised the patho-system maize-Aspergillus flavus; the former focuses on the dynamics of A. flavus sporulation the lalatter on the development of a mechanistic model to predict aflatoxin produced by A. falvus. Another mechanistic model for Fusarium ear rot and fumosin production in maize (chapter 7). The last chapter summarised the activity done in the European project MYCORED in which several countries worldwide were involved and wheat and/or maize samples collected with data necessary for model validation.