Dissertations / Theses on the topic 'Aflatoxin contamination'

Mapping aflatoxin contamination in the field reveals that most toxin occurs in relatively few, highly contaminated, bolls. Several studies suggest that protection of early bolls from pink bollworm damage will eliminate many of these highly contaminated bolls. Early harvest will also help reduce aflatoxin contamination. However, the crop must still be carefully managed after harvest because toxin content of mature bolls can increase very rapidly.

Transgenic Bt cotton may have reduced susceptibility to aflatoxin contamination as a result of pink bollworm resistance. During 1995 and 1996, Bt cottonseed from several commercial fields in Arizona contained aflatoxin levels unacceptable for dairy use. Comparison of cottonseed with and without BGYF (bright-green-yellow fluorescence) from one highly contaminated (> 6,000 ppb aflatoxin Bj) Bt seed lot indicated that most contamination probably resulted from exposure of mature cotton to high humidity. Seed exhibiting BGYF was repeatedly detected in Bt cottonseed lots but, pink bollworm exit holes were not observed in the field. A field plot test in 1996 demonstrated high resistance among Bt cultivars to both pink bollworm damage and formation of BGYF seed cotton. These observations suggest that resistance to pink bollworm will result in reduced aflaaoxin contamination when pink bollworm pressure coincides with conditions conducive to Aspergillus flavus infection. However, Bt cultivars are not resistant to aflatoxin increases occurring after boll opening and large quantities aflatoxin can form during this period. If insect control provided by Bt cultivars leads growers to hold crops in the field longer, most advantages of Bt cotton in aflatoxin management may be lost. Combined use of Bt cultivars and atoxigenic strains of A. flavus may result in the most reliable control of aflatoxin contamination.

Aflatoxins are cancer-causing, immuno-suppressive mycotoxins that frequently contaminate important staples in Zambia including maize and groundnut. Managing aflatoxins begins with understanding the distribution of aflatoxins across the target region. Seventeen percent of crops from markets contained aflatoxin concentrations above allowable levels in Zambia, with the frequency of contamination in groundnut and maize highest in warmest regions of the country. Proper management of aflatoxin contamination requires a clear understanding of the etiologic agents of the observed contamination. Several species within Aspergillus section Flavi have been implicated as causal agents of aflatoxin contamination in Africa. In Zambia, A. parasiticus was the main etiologic agent of aflatoxin contamination of maize and groundnut, although fungi with S morphology also caused contamination. Aspergillus flavus L morphotype fungi were associated with reduced aflatoxins, suggesting natural biological control by atoxigenic strains may reduce aflatoxin contamination in Zambia. In addition to maize and groundnut, wild insects, fruits and fish are important sources of food and incomes in Zambia. Unfortunately, both insects and wild plants are susceptible to aflatoxin contamination. To evaluate the safety of wild insects and fruit, concentrations of aflatoxins and presence of aflatoxin-producers were assessed. Some species of wild fruits and insects were found to have unsafe levels of aflatoxins suggesting mitigation efforts should target these important foods of Zambia in addition to crops such as groundnut and maize. New lineages of aflatoxin-producing fungi have been described, and found associated with cases of aflatoxicoses in Kenya and elsewhere. Although A. parasiticus is highly frequent and an important etiologic agent of aflatoxin contamination, it is not known how this fungus is related to similar fungi elsewhere. A multigene phylogenetic analysis revealed at least two new groups divergent from known fungal species whose frequencies need to be modified if aflatoxin contamination of crops is to be reduced.

Aflatoxins are secondary metabolites produced by members of the fungal genus Aspergillus. Immunosuppressive and carcinogenic activities of these toxins negatively impact human health especially in developing countries. Severity of contamination is influenced by both fungal community structure and the environment to which the crop is exposed either prior to or after harvest. In 2004, a severe episode of lethal human aflatoxicosis occurred in the Eastern Province of Kenya. Analysis of fungal community structure revealed that this event was caused by a previously unknown fungal lineage closely resembling the S strain morphotype of Aspergillus flavus. Fungal communities associated with maize produced in affected regions of Kenya were invariably dominated by the new fungal lineage and its incidence was strongly correlated with maize aflatoxin content. Analyses of fungal communities of maize grown in adjacent Kenyan provinces showed that incidences of the new lineage are limited outside the Eastern Province where the aflatoxicoses outbreaks occurred. Multi-locus phylogenetic analyses suggest the newly identified Kenyan lineage is closely related to the B and G aflatoxin producing species A. minisclerotigenes, and more distantly related to both the A. flavus S strain and an unnamed taxon with similar morphology endemic in West Africa (strain SBG). Sequence analyses of the cypA aflatoxin biosynthesis gene identified a previously unknown 2.2 kb deletion unique to the Kenyan lineage and coherent with its phylogenetic placement. A polyphasic approach was used to study aflatoxin-producing fungal communities, with emphasis on occurrence of fungi with S strain morphology, in Sub-Saharan Africa. Four phylogenetically distinct groups of fungi with S strain morphology were identified with restrictions to West Africa (strain SBG) or Central and East Africa (A. flavus S strain, A. minisclerotigenes, the new lineage). Aflatoxin production in synthetic media was a poor predictor of aflatoxin production in viable maize grain. An in vitro assay was developed to predict the aflatoxin-producing potential of fungal isolates in maize. This screen was used to identify atoxigenic isolates of A. flavus with potential value for biological control within highly toxic Aspergillus communities associated with maize production in Kenya. These atoxigenic isolates have potential value for mitigating aflatoxin contamination in Kenya.

Aflatoxin (AFL) contamination of corn is a serious economic and food security issue. Although a variety of technical solutions for reducing AFL contamination of corn have been proposed, only a few have produced satisfactory results. A successful approach is a biocontrol strategy consisting of using non-flatoxigenic strains of Aspergillus flavus to replace indigenous AFL-producing isolates. The main objective of the present thesis was to investigate the dynamic and contamination of AFL/A. flavus in corn in Northern Italy. The study also investigated the role of the key-pest of corn, the European Corn Borer (ECB), on AFL contamination and dispersal of A. flavus propagules in corn. Finally, the study evaluated the feasibility of bioplastic-based granules entrapping a non-aflatoxigenic A. flavus strain for the biocontrol of this fungus in corn. The 2-year field study demonstrated the efficacy of the bioplastic formulation to reduce AFL contamination in corn. More precisely, although AFL contamination varied among the two years, application of 15 and 30 kg ha-1 of granules reduced AFL contamination to up 60 and 85% in 2009 and 2010 respectively. Microbiological analysis showed that the relative abundance of non-aflatoxigenic soil isolates significantly increased after 1 month from granules application (mid-May) and throughout the corn-growing season. These findings were consistent with data obtained using a bioplastic-based bait specifically developed to selectively isolate Aspergilli from soil and other environmental samples. In addition, field and laboratory evaluations showed that the level of damages produced by ECB larvae were not significantly correlated to A. flavus infestation and AFL contamination. Taking together, these findings demonstrated that AFL contamination of corn in Northern Italy was variable, but above the EU limit for human consumption. First proposed in the USA, this study showed the practical possibility of this formulation to be use for reducing AFL contamination in corn in the EU.

Aflatoxin (AFL) contamination of corn is a serious economic and food security issue. Although a variety of technical solutions for reducing AFL contamination of corn have been proposed, only a few have produced satisfactory results. A successful approach is a biocontrol strategy consisting of using non-flatoxigenic strains of Aspergillus flavus to replace indigenous AFL-producing isolates. The main objective of the present thesis was to investigate the dynamic and contamination of AFL/A. flavus in corn in Northern Italy. The study also investigated the role of the key-pest of corn, the European Corn Borer (ECB), on AFL contamination and dispersal of A. flavus propagules in corn. Finally, the study evaluated the feasibility of bioplastic-based granules entrapping a non-aflatoxigenic A. flavus strain for the biocontrol of this fungus in corn. The 2-year field study demonstrated the efficacy of the bioplastic formulation to reduce AFL contamination in corn. More precisely, although AFL contamination varied among the two years, application of 15 and 30 kg ha-1 of granules reduced AFL contamination to up 60 and 85% in 2009 and 2010 respectively. Microbiological analysis showed that the relative abundance of non-aflatoxigenic soil isolates significantly increased after 1 month from granules application (mid-May) and throughout the corn-growing season. These findings were consistent with data obtained using a bioplastic-based bait specifically developed to selectively isolate Aspergilli from soil and other environmental samples. In addition, field and laboratory evaluations showed that the level of damages produced by ECB larvae were not significantly correlated to A. flavus infestation and AFL contamination. Taking together, these findings demonstrated that AFL contamination of corn in Northern Italy was variable, but above the EU limit for human consumption. First proposed in the USA, this study showed the practical possibility of this formulation to be use for reducing AFL contamination in corn in the EU.

Transgenic Bt cotton varieties that are resistant to pink bollworm should sustain less feeding damage to bolls and cottonseed, compared to non-Bt varieties that are more susceptible to feeding damage by pink bollworm larvae. Prior to boll opening, the aflatoxin producing fungus Aspergillus flavus cannot penetrate undamaged cotton bolls. Thus resistance to pink bollworm could result in reduced aflatoxin contamination under high pink bollworm pressure. Cottonseed aflatoxin levels of Bt and non-Bt varieties were compared at various planting and harvest dates. Bt and non-Bt cotton varieties had similar cottonseed aflatoxin levels. Long season production systems favored high cottonseed aflatoxin levels, compared to short season production systems, regardles of the cotton variety grown.

Aflatoxin is a poisonous mold that has spread around the world, posing a threat to food security and the agricultural economy. A total of 4,5 million people are in danger of being exposed to aflatoxins on a long-term basis around the world. Acute toxicity is caused by consuming significant amounts of toxins in a short period of time, which can lead to death in the worst cases, whereas chronic toxicity is caused by consuming small quantities over a longer period of time, which can lead to low birth weight, immunosuppression, restricted growth in children, and liver cancer in the worst cases. The occurrence of aflatoxins in West Africa was recognized, studied, and investigated in this paper based on a literature review. The findings demonstrate that large levels of aflatoxins have been found in West African raw materials and food and the human body. Children under the age of five, pregnant women, and breastfeeding mothers are the most vulnerable to aflatoxins. Furthermore, the findings reveal that the majority of the population is unaware of aflatoxins and its health implications. Inadequate governmental systems, low societal development, lack of access to health care, a low educated population, climate variability and climate change, high levels of illiteracy, and a lack of laboratories are only a few of the many obstacles that the region has in limiting aflatoxins concentrations. Sorting procedures, the use of tarpaulins, and seminars have all helped raise awareness and knowledge and reduce contamination and consumption of aflatoxin-contaminated foods. This study argues that a multi-sectoral strategy is needed to promote food security and local education. Increased limitations and regulations and higher standards may be able to help limit aflatoxin contamination and exposure.

Contamination of peanuts by secondary metabolites of certain fungi, namely aflatoxins present a great health hazard when exposed either at low levels for prolonged times (carcinogenic) or at high levels at once (poisonous). It is important to develop an accurate and rapid measurement technique to trace the aflatoxin and/or source fungi presence in peanuts. Thus, current research focused on development of vibrational spectroscopy based methods for detection and separation of contaminated peanut samples. Aflatoxin incidence, as a chemical contaminant in peanut paste samples, was investigated, in terms of spectral characteristics using FTIR-ATR. The effects of spectral pre-processing steps such as mean-centering, smoothing the 1st derivative and normalizing were studied. Logarithmic method was the best normalization technique describing the exponentially distributed spectral data. Spectral windows giving the best correlation with respect to increasing aflatoxin amount led to selection of fat associated spectral bands. Using the multivariate analysis tools, structural contributions of aflatoxins in peanut matrix were detected. The best region was decided as 3028-2752, 1800-1707, 1584-1424, and 1408-1127 cm-1 giving correlation coefficient for calibration (R2C), root mean square error for calibration (RMSEC) and root mean square error for prediction (RMSEP) of 98.6%, 7.66ppb and 19.5ppb, respectively. Applying the constructed partial least squares model, 95% of the samples were correctly classified while the percentage of false negative and false positive identifications were 16% and 0%, respectively. Aspergillus species of section Flavi and the black fungi, A. niger are the most common colonists of peanuts in nature and the majority of the aflatoxin producing strains are from section Flavi. Seed colonization by selected Aspergillus spp. was investigated by following the chemical alterations as a function of fungal growth by means of spectral readouts. FTIR-ATR was utilized to correlate spectral characteristics to mold density, and to separate Aspergillus at section, species and strain levels, threshold mold density values were established. Even far before the organoleptic quality changes became visually observable (~10,000 mold counts), FTIR distinguished the species of same section. Besides, the analogous secondary metabolites produced increased the similarity within the spectra even their spectral contributions were mostly masked by bulk peanut medium; and led to grouping of species producing the same mycotoxins together. Aflatoxigenic and non-aflatoxigenic strains of A. flavus and A. parasiticus were further studied for measurement capability of FTIR-ATR system in discriminating the toxic streams from just moldy and clean samples. Owing to increased similarity within the collected spectral data due to aflatoxin presence, clean samples (having aflatoxin level lower than 20 ppb, n=44), only moldy samples (having aflatoxin level lower than 300 ppb, n=28) and toxic samples (having aflatoxin level between 300-1200 ppb, n=23) were separated into appropriate classes (with a 100% classification accuracy). Photoacoustic spectroscopy (PAS) is a non-invasive technique and offers many advantages over more traditional ATR system, specifically, for in-field measurements. Even though the sample throughput time is longer compared to ATR measurements, intact seeds can be directly loaded into sample compartment for analysis. Compared to ATR, PAS is more sensitive to high moisture in samples, which in our case was not a problem since peanuts have water content less than 10%. The spectral ranges between: 3600-2750, 1800-1480, 1200-900 cm-1 were assigned as the key bands and full separation between Aspergillus spp. infected and healthy peanuts was obtained. However, PAS was not sensitive as ATR either in species level classification of Aspergillus invasion or toxic-moldy level separation. When run for separation of aflatoxigenic versus non-aflatoxigenic batches of samples, 7 out of 54 contaminated samples were misclassified but all healthy peanuts were correctly identified (15 healthy/ 69 total peanut pods). This study explored the possibility of using vibrational spectroscopy as a tool to understand chemical changes in peanuts and peanut products to Aspergillus invasion or aflatoxin contamination. The overall results of current study proved the potential of FTIR, equipped with either ATR or PAS, in identification, quantification and classification at varying levels of mold density and aflatoxin concentration. These results can be used to develop quality control laboratory methods or in field sorting devices.
Ph. D.

Aflatoxin contamination of cottonseed from bolls damaged by the pink bollworm was compared with contamination of cottonseed from undamaged bolls. Cottonseed produced in pink bollworm damaged bolls was the predominant source of aflatoxin contaminated cottonseed.

Les micotoxines són metabòlits secundaris tòxics produïts per fongs filaments que poden contaminar una àmplia varietat de productes agrícoles tant en etapes precollita com en etapes postcollita. La gestió de la contaminació per micotoxines durant la cadena de producció de la llet és essencial per evitar la presència d'aflatoxina M1 (AFM1) en la llet com a conseqüència de l'exposició d'animals productors de llet a pinsos contaminats per aflatoxina B1 (AFB1). L'objectiu d'aquesta tesi va ser l'estudi de la contaminació per micotoxines en pinsos i ingredients per a pinsos per a vaques lleteres i la seva transferència a la llet. La presència d'aflatoxines i micotoxines de Fusarium es va avaluar mitjançant l'anàlisi de mostres de ració total barrejada (TMR) i diferents tipus d'ensitjats per a vaques lleteres. Mostres de llet procedents de vaques alimentades amb les mostres de TMR recollides es van analitzar per estimar la transferència de AFB1 en el pinso a AFM1 en la llet. Per saber si el tractament tèrmic afecta el contingut de AFM1 en la llet durant el seu processat, es van provar diferents tractaments tèrmics en llet contaminada natural i artificialment.
Las micotoxinas son metabolitos secundarios tóxicos producidos por hongos filamentos que pueden contaminar una amplia variedad de productos agrícolas tanto en etapas precosecha como en etapas poscosecha. La gestión de la contaminación por micotoxinas durante la cadena de producción de la leche es esencial para evitar la presencia de aflatoxina M1 (AFM1) en la leche como consecuencia de la exposición de animales productores de leche a piensos contaminados por aflatoxina B1 (AFB1). El objetivo de esta tesis fue el estudio de la contaminación por micotoxinas en piensos e ingredientes para piensos para vacas lecheras y su transferencia a la leche. La presencia de aflatoxinas y micotoxinas de Fusarium se evaluó mediante el análisis de muestras de ración total mezclada (TMR) y diferentes tipos de ensilados para vacas lecheras. Muestras de leche procedentes de vacas alimentadas con las muestras de TMR recogidas se analizaron para estimar la transferencia de AFB1 en el pienso a AFM1 en la leche. Para saber si el tratamiento térmico afecta al contenido de AFM1 en la leche durante su procesado, se probaron diferentes tratamientos térmicos en leche contaminada natural y artificialmente.
Mycotoxins are toxic secondary metabolites produced by filamentous fungi which can contaminate a wide variety of agricultural commodities either at pre-harvest or post-harvest stages. Through the milk supply chain, the management of mycotoxin contamination is essential in order to avoid the presence of aflatoxin M1 (AFM1) in milk as a consequence of the exposure of lactating animals to aflatoxin B1 (AFB1)-contaminated feed. The aim of this Thesis was to evaluate the mycotoxin contamination of feed and feed ingredients for dairy cows and their transference to milk. The occurrence of aflatoxins and Fusarium mycotoxins was evaluated through the analysis of total mixed ration (TMR) samples and different types of silages for dairy cows. Milk samples collecting from cows fed with the sampled TMR were analysed so as to estimate the transference of AFB1 form feed to AFM1 in milk. In order to know whether heat treatment affect to the AFM1 content in milk during processing, different heat treatments were tested in artificially and naturally contaminated milk.

Experiments were performed at the Yuma Valley Agricultural Center to determine how timely harvest of cotton may affect aflatoxin contamination of cottonseed As the cotton was held in the field between the final irrigation and harvest, the quantity of aflataxin in the crop increased. Significant reductions in aflatoxin contents of seed were realized by harvesting in early September.

The overall goal of our study is to find effective and environmentally sound methods to reduce pre -harvest aflatoxin contamination of cottonseed in Arizona. The specific objectives are: 1) to screen a large number of bacteria for their ability to destroy the aflatoxin producing fungus, Aspeigillus flavus; 2) to test the efficacy and consistency of the recovered antagonistic bacterial isolates to reduce aflatoxin contamination of cottonseed in field trials; 3) to study the survival and competitiveness of the antagonists on cotton plants under prevailing field conditions; 4) to find innovative procedures to enhance survivability and competitiveness of the antagonists on cotton plants; and 5) to test the potential of the bacterial antagonists to reduce the population of A. flavus in field soils. Over 800 bacterial isolates, recovered from cotton field soils, cotton leaves, stems, and immature as well as opened bolls, were tested for ability to inhibit the growth of A. flavus on cottonseed. Six isolates partially or totally inhibited the fungus. All of these effective isolates prevented the fungus fioin infecting simulated pink bollworm exit holes in immature bolls in the field.

The aim of the present work was to investigate aflatoxin levels in various food commodities and to study its production by Aspergillus parasiticus in culture to find out the possible ways to control it. Of 40 food samples collected from Abha region, Saudi Arabia, only 25% were contaminated with aflatoxins. Oil-rich commodities had the highly contaminated commodities by fungi and aflatoxins while spices were free from afatoxins. The optimal pH for the growth of A. parasiticus and its productivity of aflatoxin B, was found at 6.0, while the best incubation conditions were found at 30°C for 10 days. D-glucose was the best carbon source for fungal growth, as well as aflatoxin production. Corn steep liquor, yeast extract and peptone were the best nitrogen sources for both fungal growth and toxin production. (NH4)2HPO4 (1.55 gL-1) and NaNO2 (1.6 gL-1) reduced fungal growth and toxin production with 37.7% and 85%, respectively. Of ten amino acids tested, asparagine was the best for aflatoxin B, production. Zn2+ and Co2+ supported significantly both fungal growth, as well as aflatoxin B, production at the different tested concentrations. Zn2+ was effective when added to A. parasiticus growth medium at the first two days of the culture age. The other tested metal ions gave variable effects depending on the type of ion and its concentration. Water activity (a ) was an important factor controlling the growth of A. parasiticus and toxin production. The minimum aW for the fungal growth was 0.8 on both coffee beans and rice grains, while aW, of 0.70 caused complete inhibition for the growth and aflatoxin B, production. H202 is a potent inhibitor for growth of A. parasiticus and its productivity of toxins. Incubation with NaHCO3 and C6H5000Na converted aflatoxin B, to a water-soluble form which returned to aflatoxin B, by acid treatment. Black pepper, ciliated heath, cuminum and curcuma were the most inhibitory spices on toxin production. Glutathione, quinine, EDTA, sodium azide, indole acetic acid, 2,4-dichlorophenoxy acetic acid, phenol and catechol were inhibitory for both growth, as well as, aflatoxin B, production. Stearic acid supported the fungal growth and decreased the productivity of AFBI gradually. Lauric acid is the most suppressive fatty acid for both fungal growth and aflatoxin production, but oleic acid was the most potent supporter. Vitamin A supported the growth but inhibited aflatoxin B, production. Vitamins C and D2 were also repressive particularly for aflatoxin production. The present study included determining the activities of some enzymes in relation to aflatoxin production in A. parasiticus culture during 20 days. Glycolytic enzymes and pyruvate-generating enzymes seems to be linked with aflatoxin B, production. Also, pentose-phosphate pathway enzymes may provide NADPH for aflatoxin B, synthesis. The decreased activities of TCA cycle enzymes particularly from 4th day of growth up to 10th day were correlated with the increase of aflatoxin B, production. All the tested enzymes as well as aflatoxin B, production were inhibited by either catechol or phenol.

Aflatoxins (AFs) are a group of mycotoxins produced by moulds of Aspergillus genus which contaminate food commodities of tropical and sub-tropical countries. The aims of this study are to assess the extent of AF contamination of foods in the UK that originated in Asia and to identify components of Thai foods that may protect against the toxicity of AFs. Examination of 12 commercial, dried Asian foods showed that long grain rice, fragrant rice, peanuts, black beans and black pepper contained Aspergillus spp. which were identified as A. parasiticus (afiatoxigenic), A. versicolor, A. ustus, A.niger and A. ochraceus. These commodities contained undetectable AFs. Jasmine brown rice and crushed chilli contained 14.7 and 11.4 IJglkg of AFs, respectively, in the absence of Aspergillus. AFBl, the most toxigenic AFs was detected in crushed chilli (lO.7IJglkg) so Aspergillus was present at some stage of food production, particularly pre-harvest stage. Cross contamination during food processing is one of possible cause of AFs contamination in these commodities. These results indicate direct and indirect risks of exposure to AFs from these products since AFs fOlmation is possible in Aspergillus-contaminated crops and AFs can be carried throughout the food chain. Hence an alternative strategy to mitigate toxicity of ingested AFs is required. One possibility is by using food components to modulate the harm from ingested AFs by altering AF metabolism and mutagenesis. In this study, the effectiveness of the compounds in Thai culinary herb, fingerroot (Boesenbergia rotunda) were investigated. A crude methanol extract (ME) of fingerroot was analysed and found to contain ~ 15 putative flavonoids as major components of which pinocembrin, pinocembrin chalcone, cardamonin, pinostrobin, 4-hydroxypanduratin A and panduratin A were identified by HPLC and LC-MS. The ME significantly inhibited fonnation of mutagenic/carcinogenic metabolite of AFBl (AFB1-epoxide, AFBO) in a cell-free metabolic system (Model 1) and also suppressed mutagenicity of AFBl in Salmonella/microsomal assay (Model 2). These inhibitory properties of the ME might be related to its indigenous flavonoids which could modulate activities of AFB1 -metabolising enzymes (CYPIA2, 3A4). Purified flavonoids (cardamonin, apigenin, pinostrobin) were found to affect AFB 1 toxicity to some extent in both models, but their potencies were much lower than the ME particularly in the Salmonella model and evidence is provided that suggests that Reactive Oxygen Species rather than AFBO are the mutagenic entities in this assay. This also suggests that fingerroot contains effective metabolic modulators that were not identified. Consumption of fmgerroot could provide a combination of potential phytochemicals that protect against aflatoxin-mediated toxicity by altering AF metabolism at the initial stages of enzymatic activation.

A study was conducted in semi-arid Manyoni District, Central Tanzania, involving eight village communities to: 1) explore dietary and agricultural practices associated with aflatoxin risk; 2) assess aflatoxin contamination of village grains and groundnuts; and 3) assess village chicken susceptibility to aflatoxin exposure. To address the respective objectives, 1) Questionnaires were asked to 76 adult respondents randomly selected from households enrolled in a project, 2) A screening test was used on grain and groundnut samples collected peri-harvest (n=134) and ≥ 6 months in storage (n=157), followed by HPLC test on samples contaminated ≥10 µg/kg, and 3) 16 day old village chicks, randomly allocated to four groups, were fed diet with average aflatoxins ranging to 27, 102, 110, 358 µg/kg respectively, for 60 days. Most participating village farmers were naive to food toxins, with limited extension services, did not generally use irrigation, fertilisers and pesticides on crops, had inadequate harvesting, drying and storage techniques, and often consumed unpolished grains. Aflatoxin contamination of grains and groundnuts ranged to 198 µg/kg (mean=25.46 µg/kg) in post-harvest samples and 351 µg/kg (mean=50.83 µg/kg) in stored samples, above 10 µg/kg Tanzania MTL. Chronic exposure of village chickens to as low as 30 µg/kg aflatoxin containing feed was associated with organ lesions, low weight gain, low feed consumption, and low response to Newcastle disease vaccination.

Peanuts are an important crop grown in Egypt for either local consumption or export to European markets. The present study examined the importance of mycotoxigenic Aspergilli in Egyptian peanuts from five different regions (Alexandria, El-Beheira, El- Daqahliya, El-Sharqiya, Asyut) in two seasons (2007, 2008). This led to consideration of different potential strategies to control aflatoxigenic A. flavus strains and associated aflatoxin contamination of peanuts. The most common species in peanuts were from Aspergillus section Flavi, Aspergillus section Nigri and Aspergillus section Circumdati. Both qualitative (coconut cream agar) and quantitative analyses (HPLC) were used to analyse the potential mycotoxin production by strains isolated from peanuts. Of a total of 88 Aspergillus section Flavi strains examined, 90% were aflatoxigenic. Cont/d.

A strain of Aspergillus flavus that does not produce aflatoxins was applied to soils planted with cotton at the Yuma Valley Agricultural Center in order to assess strain ability to competitively exclude aflatoxin producing strains during cotton boll infection and thereby prevent aflatoxin contamination of cottonseed. In both 1989 and 1990, the atoxigenic strain displaced other infecting strains during cotton boll development. Displacement was associated with significant reductions (75% to 82% in 1989, and 99% in 1990) in the quantity of aflatoxins contaminating the crop at maturity. Although frequency of infected locules differed between years, in both years displacement occurred without increases in the amount of developing boll infection. Currently, an Experimental Use Permit is being sought from the EPA for tests on commercial acreage

This study has (a) evaluated the biodiversity in toxigenic mycobiota associated with maize from subsistence farmers’ stores in five climatic regions of Lesotho in two seasons, (b) compared the effect of ecophysiological factors on interactions between atoxigenic (AFL-) and toxigenic (AFL+) Aspergillus strains and control of aflatoxin B1 (AFB1) contamination of maize, (c) examined the mechanism of action of AFL- strains in relation to mycelial growth rate, sporulation, germination rate, germ tube extension, C-source utilisation patterns and hydrolytic enzymes and (d) examined ecophysiological approaches to enhance competitiveness of the atoxigenic strains.

Chilean red chili peppers contaminated with aflatoxins were reported in a previous study. If the development of gallbladder cancer (GBC) in Chile is associated with a high level of consumption of aflatoxin-contaminated red chili peppers, such peppers from other countries having a high GBC incidence rate may also be contaminated with aflatoxins. We aimed to determine whether this might be the case for red chili peppers from Bolivia and Peru. A total of 7 samples (3 from Bolivia, 4 from Peru) and 3 controls (2 from China, 1 from Japan) were evaluated. Aflatoxins were extracted with acetonitrile:water (9:1, v/v) and eluted through an immuno-affinity column. The concentrations of aflatoxins B1, B2, G1, and G2 were measured using high-performance liquid chromatography (HPLC), and then the detected aflatoxins were identified using HPLC-mass spectrometry. In some but not all of the samples from Bolivia and Peru, aflatoxin B1 or aflatoxins B1 and B2 were detected. In particular, aflatoxin B1 or total aflatoxin concentrations in a Bolivian samples were above the maximum levels for aflatoxins in spices proposed by the European Commission. Red chili peppers from Bolivia and Peru consumed by populations having high GBC incidence rates would appear to be contaminated with aflatoxins. These data suggest the possibility that a high level of consumption of aflatoxin-contaminated red chili peppers is related to the development of GBC, and the association between the two should be confirmed by a case-control study.

Stink bugs, including native brown stink bug (Euschistus servus) and invasive brown marmorated stink bug (Halyomorpha halys), cause damage to a variety of crops including field corn (Zea mays). Frequency and size of stink bug infestations have increased in corn fields in the Mid-Atlantic U.S., and there are growing concerns that these infestations may contribute to reductions in grain quality including increased mycotoxin concentrations. Prior research on native and invasive stink bugs has focused on understanding their biology, the damage they cause, and elucidating effective and economic management strategies. However, few studies examined the potential for stink bugs to facilitate fungal infection and mycotoxin contamination of corn grain. Thus, the objectives of this research were to: 1) assess the relationship between invasive brown marmorated stink bug (H. halys) feeding injuries and fumonisin contamination of field corn in the Mid-Atlantic U.S., 2) determine if stink bugs are a vector for mycotoxigenic Fusarium spp. in corn, and 3) evaluate the efficacy of pesticides for mitigating stink bug feeding injury and associated mycotoxin contamination in field corn. A correlation between H. halys feeding injury and fumonisin concentrations was identified, and the ability of H. halys to increase F. verticillioides infection and fumonisin concentrations in corn was demonstrated in field experiments. Fusarium species including fumonisin-producing F. verticillioides and F. proliferatum were isolated from field-collected stink bugs, and in laboratory experiments, E. servus was able to transmit F. verticillioides to non-infected corn ears after feeding on F. verticillioides-infected corn. In field studies, both fungicide and insecticide reduced stink bug-associated mycotoxin concentrations in corn, but levels of control were inconsistent. Thus, additional tactics that target both the stink bug and Fusarium should be implemented to mitigate risks of mycotoxin contamination in corn.
Doctor of Philosophy
Native and invasive stink bugs can severely damage crops including field corn. Frequency and size of stink bug infestations in Mid-Atlantic U.S. corn fields have increased, and there is growing concern that this may contribute to reductions in grain quality. Insect feeding injury is a risk factor for fungal infection and mycotoxin contamination in corn. Mycotoxins are toxic chemicals produced by certain fungi that have detrimental health effects on animals including livestock and humans. The relationship between stink bug feeding injuries and mycotoxin contamination in corn grain is not well understood, and management strategies to minimize the risk of mycotoxin contamination in corn need to be identified. The main goal of this research was to characterize interactions between stink bugs and mycotoxin-producing fungi and identify tactics for controlling both the insect pest and pathogen. Specific objectives were to: 1) assess the relationship between invasive brown marmorated stink bug (H. halys) feeding injuries and fumonisin contamination of field corn in the Mid-Atlantic U.S., 2) determine if stink bugs are a vector for mycotoxin-producing Fusarium spp. in corn, and 3) evaluate the efficacy of pesticides for mitigating stink bug feeding injury and associated mycotoxin contamination in field corn. Results from this work indicated that stink bugs have the ability to cause feeding injuries which facilitate invasion of mycotoxin-producing Fusarium species, leading to increases in mycotoxin concentrations in corn grain. Studies also demonstrated that stink bugs can vector Fusarium species during feeding and increase Fusarium infection of corn resulting in subsequent mycotoxin contamination. Field studies indicated that pesticide applications targeting both the stink bugs and mycotoxigenic fungi may be needed to minimize risk of mycotoxin contamination in corn. However, under low pest pressure, application of pesticides is unlikely to be profitable.

Le aflatossine si possono trovare come contaminanti naturali in una varietà di prodotti alimentari tra cui: cereali, noccioline,arachidi, pistacchi, patate dolci, burro di arachidi, banane, vino, spezie, latte e latticini, ecc. Perciò è molto importante avere metodi semplici e abbastanza rapidi per l’analisi di queste sostanze. Le tecniche immunologiche hanno il vantaggio rispetto alle tecniche classiche (cromatografia) di non richiedere elevata purificazione e concentrazione, inoltre sono rapidi, semplici da adoperare, sensibili e specific. La loro specificità è dovuta all’uso di anticorpi monoclonali o policlonali selettivi per quellla tossina. Lo scopo di questa tesi di dottorato è stato quello di realizzare metodi per la determinazione del’aflatossina M1 e B1, che possano essere semplici da adoperare, rapidi e privi della necessità di una procedura di estrazione molto complicata/elaborata. La tesi è stata strutturata nei sequenti capitoli: Capitolo I. Descrive alcuni aspetti generali riguardanti la tossicità, struttura chimica, la contaminazione, la legislazione e metodi di detossificazione delle aflatossine. Capitolo II. Presenta lo stato del arte nel campo dei metodi analitici della determinazione di aflatossine, tra cui un sottocapitolo dedicato ai kit comerciali disponibili al momento. Capitolo III. Descrive le applicazioni sperimentali realizzate durante questa tesi, divise in due metodi per la determinazione dell’aflatossina M1 in latte (un metodo immunologico accopiato con l’analisi amperometrica in flusso e un altro metodo che utilizza delle piastre a 96-pozzetti elettrochimiche accopiate con la tecnica amperometrica ad impulsi intermitenti). La seconda parte è stata focalizzata sull’ottimizzazione di un metodo spettrofotometrico immunoenzimatico per la determinazione dell’aflatossina B1 in campioni di mais utilizzando un nuovo coniugato AflatossinaB1-fosfatasi alcalina (AFB1-AP) realizzato nel nostro laboratorio, non essendo comercialmente disponibile. In più, è stato realizzato anche uno studio della stabilità dell’aflatossina M1 in campioni freschi di latte contaminati artificialmente e conservati a 4°C (latte liofilizzato) ed a -30°C (latte liquido) per 3 mesi. Questo studio è stato realizzato applicando la tecnica amperometrica ad impulsi intermitenti accopiata con l’utilizzo delle piastre a 96-pozzetti elettrochimiche. Capitolo IV. Sottolinea tutte le applicazioni analitiche per la determinazione dell’aflatossina M1 e B1 in campioni reali con le loro charatteristiche, vantaggi e svantaggi.
Aflatoxins can be found as natural contaminants in a variety of food ingredients such as, cereals and cereals products, peanuts, nuts, almonds, pistachios, hazelnut and other dried fruits, coconut, cocoa, sweet potato, peanut butter, bananas, wine, spices, milk and milk products, etc. It is, therefore, important to have available, simple and quantitative methods for aflatoxin analysis. Rapid methods based on immunochemical techniques usually have the advantage of not requiring clean-up or analyte-enrichment steps, are speed, ease of operation, sensitive and specific by using the specific monoclonal or polyclonal antibodies produced against the toxin. The aim of this Ph.D. thesis was to develop methods for AFM1 and AFB1 determination which should be simple of operation, rapid and not requiring a complex clean-up procedure. The thesis has been structured in the following chapters: • Chapter I describes some general aspects regarding the toxicity, chemical structure, contamination, legislation and detoxification methods regarding aflatoxins. • Chapter II presents the state of the art in the field of analytical methods for aflatoxins determination, including commercial kits available at the moment of the writing of this thesis. • Chapter III describes the experimental applications carried out during this thesis, which are divided in two methods for AFM1 determination in milk samples (a Flow Injection Immunoassay with amperometric detection and an electrochemical multichannel microplate coupled with Intermittent Pulse Amperometry) and a second part which is focused on a spectrophotometric enzyme immunoassay for AFB1 determination in corn samples using a new conjugate Aflatoxin B1-alkaline phosphatase (AFB1-AP) prepared in our laboratory, being not commercial available. Moreover, a study of stability of AFM1 in raw milk samples stored at 4°C (lyophilized milk) and -30°C (liquid milk) for 3 months, was carried out. The study was realized with the electrochemical multichannel plate with IPA technique. • Chapter IV underlines all analytical applications for AFM1 and AFB1 determination in real samples with its performance, advantages and drawbacks.

This thesis deals with the reduction of mycotoxin contamination level during soybean fermentation (Thua-Nao). Beside this work, isolation, characterization, and ochratoxin A production ability of toxigenic fungi from French grapes were also study. Results of this latter part showed that Aspergillus carbonarius and Aspergillus niger are the most ochratoxin A producer in wine grape from France. Furthermore, Aspergillus japonicus can produce a little bit quantity of ochratoxin A in wine grape too. Regarding to the main part of the work, 23 isolates of Bacillus spp. were isolated from Thai Thua-Nao. An Aspergillus flavus aflatoxin producing strain was also isolated from Thua-Nao whereas an Aspergillus westerdijkiae was chosen as an OTA producing reference strain. The objectives were to find an efficient Bacillus strain for: Growth inhibition of Aspergillus flavus and Aspergillus westerdijkiae NRRL 3174. - Limitation of aflatoxin B1 production. ; - Mycotoxins, aflatoxin B1 and ochratoxin A detoxification. Among the results, Bacillus CM 21, which was identified later by ITS sequencing as Bacillus licheniformis, showed the highest ability on inhibition of growth of both Aspergillus strains and both of mycotoxins removal (decrease of 74% of AFB1 and 92.5% of OTA). Another Bacillus strain, MHS 13, inhibiting both Aspergillus growth and detoxifying 85% of AFB1 was identified as Bacillus subtilis. Finally, culture supernatant and cellular extract from both interested Bacillus strains were tested for aflatoxin B1 and ochratoxin A degradation ability in order to know their degradation mechanisms. Moreover, study on optimal condition for aflatoxin B1 and ochratoxin A degradation were also conducted. All results indicated that OTA was significantly degraded by culture supernatant from Bacillus licheniformis CM 21 (p lower than 0.0001) in OTalpha. The percentage of OTA degradation was 97.5% and the optimal activity of its culture supernatant was found at pH 7.0 and 37°C with 24 h culture incubation time and 2 h contact time. Moreover, OTA was also significantly degraded by culture supernatant from Bacillus subtilis MHS 13 (p lower than 0.0017) at pH 5.0 and 37°C with 48 h culture incubation time and 2 h contact time. The proposed degradation mechanism should be extracellular and carboxypeptidase A probably responsible for this degradation since no activity was found for the intracellular extract. However, AFB1 could be degraded by neither culture supernatant nor cellular extract from both of these microorganisms. Hence, the AFB1 detoxification mechanism may be due to non-enzymatic mechanism.

Dissertação composta por 02 artigos.
CNPq; Capes
O leite é uma das principais fontes de nutrientes da dieta humana e é um alimento que acompanha o ser humano durante toda a vida, tanto como leite de consumo como através de seus derivados. Entretanto, são inúmeras as formas e os tipos de contaminação que acometem o leite, destacando-se, dentre os contaminantes de ordem química, as aflatoxinas. Dentre os métodos de análise de aflatoxinas em leite e derivados, destaca-se a Cromatografia Líquida de Alta Eficiência (CLAE), principalmente devido a sua versatilidade, rapidez e acuracidade das medidas quantitativas. Entretanto, trata-se de um sistema complexo em que as propriedades dos constituintes da fase móvel são afetadas por mudanças nas condições de processo nas quais são realizados os experimentos. Normalmente as condições de análise por CLAE são determinadas empiricamente, pelo método “tentativa e erro” em que inúmeras tentativas são realizadas sem um estudo mais detalhado do sistema. Diante disso, a primeira etapa deste estudo objetivou a otimização de multirrespostas em CLAE, por meio da seleção das condições ótimas como a composição da fase móvel, sua vazão no sistema cromatográfico e a temperatura da coluna, a fim de identificar, separar e quantificar simultaneamente a aflatoxina M1 (AFM1) e a aflatoxina B1 (AFB1). Para tanto, realizou-se planejamento experimental de misturas para três componentes com restrições combinado com um planejamento fatorial 22 para as variáveis de processo (temperatura da coluna e vazão). Após a definição dos modelos para as variáveis dependentes, foi realizada busca das condições ótimas usando o método simplex sequencial e as funções de desejabilidade de Derringer e Suich. As variáveis avaliadas foram: composição da fase móvel (acetonitrila, metanol e solução aquosa de ácido acético 1%), vazão da fase móvel e temperatura da coluna. Os parâmetros cromatográficos obtidos como respostas foram: tempo e fator de retenção para ambas as aflatoxinas, fator de separação, resolução da coluna e altura dos picos. Após a validação do planejamento, foi realizada a validação analítica do método otimizado através das figuras analíticas de mérito: linearidade, precisão, exatidão e limites de detecção e quantificação. O planejamento realizado foi capaz de produzir modelos confiáveis que possibilitaram a estimativa das melhores condições atendendo aos múltiplos objetivos. Na validação analítica do método cromatográfico, os parâmetros analíticos avaliados ficaram dentro dos intervalos de confiança, podendo o método ser considerado exato e preciso, apresentando limites de quantificação de 0,3 e 0,5 μg L-1 para AFM1 e AFB1, respectivamente e linearidade com R2 > 0,99 para ambas as aflatoxinas. A segunda etapa do estudo objetivou a avaliação da interação entre as AFM1 e AFB1 com proteínas lácteas, tendo em vista que estudos demonstram que aquelas, especialmente a AFM1, localizam-se predominantemente nas frações proteicas. Entretanto, esses estudos não avaliaram a interação entre aflatoxinas e as proteínas do leite, mas apenas baseiam-se na sua quantificação nas frações proteicas do leite. Portanto, buscou-se por meio deste estudo avaliar a possível interação entre as AFM1 e AFB1 com as frações proteicas do leite. Análises por Espectroscopia na região de infravermelho com transformada de Fourier (FTIR) foram realizadas para avaliação de possíveis modificações na estrutura secundária das proteínas lácteas quando fortificadas com as aflatoxinas e Calorimetria Exploratória Diferencial (DSC). Para tanto, preliminarmente foram avaliados os espectros obtidos com padrões de caseína e β-lactoglobulina em solução tampão fosfato-salino (PBS) e em solução modelo, assim como no leite propriamente dito, integral e desnatado. Na segunda etapa, regiões espectrais específicas foram avaliadas por meio de técnicas de deconvolução e curve-fitting. Os resultados indicam que a solução PBS foi mais adequada para o estudo da interação entre as AFB1 e AFM1 e proteínas lácteas avaliadas, β-lactoglobulina e caseína. Foram observadas alterações nas estruturas secundárias e essas sugerem que, embora possivelmente ocorram interações de caráter hidrofílico entre β-lactoglobulina e as aflatoxinas (especialmente com AFM1), ocorram também interações de caráter hidrofóbico (especialmente de AFB1) com os pacotes hidrofóbicos da β-lactoglobulina. Já com a caseína, as alterações promovidas nas estruturas secundárias proteicas foram mais discretas, porém deslocamentos de picos foram observados indicando alterações estruturais da proteína, especialmente na presença de AFB1, o que sugere que ocorram interações químicas entre os componentes avaliados. As alterações espectrais, mais evidentes com a fração β-lactoglobulina, do que com a fração caseína sugerem que, embora a quantificação de aflatoxinas seja comumente superior na fração caseína, não se pode afirmar que por esse motivo ocorram interações mais tangíveis entre aflatoxinas e caseína do que entre aflatoxinas e β-lactoglobulina. Possivelmente a quantificação em maior percentual de aflatoxinas na fração caseína é atribuída ao fato dessa proteína encontrar-se, no leite, em percentual superior às proteínas do soro. Outra hipótese levantada pelo estudo é a possibilidade das aflatoxinas avaliadas encontrarem-se “mascaradas” por estarem conjugadas com a β-lactoglobulina e não sendo, portanto, detectadas pelos métodos analíticos convencionais ocasionando sua subestimação nessa fração.
Milk is one of the main sources of nutrients in the human diet and is a food that accompanies the human being throughout life, both as drinking milk as through its derivatives. However, there are countless forms and types of contamination that affect milk, especially among the contaminants of chemical order, aflatoxins. Among the methods of analysis of aflatoxins in milk and dairy products, the High Performance Liquid Chromatography (HPLC) stands out mainly due to its versatility, speed and accuracy of quantitative measurements. However, it is a complex system in which the properties of the constituents of the mobile phase are affected by changes in process conditions under which the experiments are performed. Typically the HPLC analysis conditions are determined empirically, using the "trial and error" in which numerous attempts are made without a more detailed study of the system. Therefore, the first step of this aimed to optimize multiresponses in HPLC, by selecting the optimal conditions as the mobile phase composition, its flow into the chromatographic system and the column temperature in order to identify, separate and quantify aflatoxin M1 (AFM1) and aflatoxin B1 (AFB1) simultaneously. Therefore, it was carried out experimental a mixture design for three components with restrictions combined with a 22 factorial design to the process variables (flow and column temperature). After defining the models for the dependent variables, a search of the optimum conditions was made using the sequential simplex method and the Derringer and Suich desirability functions. The variables evaluated were: mobile phase composition (acetonitrile, methanol and aqueous solution of acetic acid 1%), the mobile phase flow rate and column temperature. The chromatographic parameters obtained as responses were time and factor of retention for both aflatoxins, separation factor, column resolution and height of the peaks. After the design validation, analytical validation was performed through the analytical figures of merit: linearity, precision, accuracy and limits of detection and quantification. The experimental design carried out was able to produce reliable models that allowed better conditions estimation regarding multiple objectives. In the analytical validation of the chromatographic method, the analytical parameters evaluated were within the confidence interval, the method can be considered accurate and precise showing quantitation limits of 0.3 and 0.5 μg L-1 for AFB1 and AFM1, respectively, and linearity with R2> 0.99 for both aflatoxins. The second stage of the study aimed to evaluate the interaction between the AFB1 and AFM1 with dairy proteins, considering that studies show that those, especially AFM1, are located predominantly in the protein fractions. However, these studies did not evaluate the possibility of interaction between aflatoxins and dairy proteins, but only based on its quantification in the dairy protein fractions. Therefore, we sought through this study to evaluate a possible interaction between AFM1 and AFB1 with dairy protein fractions. Analyzes were performed by Fourier transformation infrared spectroscopy (FTIR) to assess possible changes in the secondary structure of dairy proteins when spiked with aflatoxins and Differential Scanning Calorimetry (DSC). For this purpose, preliminarily the spectra obtained were evaluated with standard casein and β-lactoglobulin in phosphate buffer saline solution (PBS) and a bovine milk model solution as well as in actual milk, whole and skim. In the second step, specific spectra bands were assessed through deconvolution and curve-fitting techniques. The results show that PBS was more suitable for the interaction study between aflatoxins B1 and M1 and the dairy proteins evaluated, β-lactoglobulin and casein. Changes in secondary structures suggest that although possibly occurring interactions with hydrophilic characters between β-lactoglobulin and aflatoxins were observed (especially with AFM1) also occur interactions with hydrophobic character (especially AFB1) with the hydrophobic β-lactoglobulin packages. Already with the casein, the changes introduced in protein secondary structure were more discreet but peak shifts were observed indicating structural changes of the protein, especially in the presence of AFB1, which suggests that chemical interactions occur between the components evaluated. The spectral changes, more evident with the β-lactoglobulin fraction than the casein fraction, suggests that although the quantification of aflatoxins is commonly higher in the casein fraction, it’s not possible to ensure that for this reason occur more tangible interactions between aflatoxins and casein than between aflatoxins and whey proteins (β-lactoglobulin). The greater quantify percentage of aflatoxins in the casein fraction, apparently, is attributed to the fact that this protein is found, in milk, in superior percentage to the whey proteins. Another hypothesis is the possibility of the aflatoxins evaluated are "masked" by being combined with β-lactoglobulin and not, therefore, being detected by conventional analytical methods leading to their underestimation in this fraction.
5000

L'objectif de cette etude etait de definir les mecanismes de contamination fongique et mycotoxique sur deux types de fruits (la noix et la figue) afin d'aboutir a des mesures de prevention raisonnee. Pour la noix, l'etude s'est deroulee en deux etapes. Tout d'abord, l'etude de la mycoflore, au cours de trois annees successives, a permis d'isoler 338 souches appartenant a 17 genres ; 14% des souches isolees produisaient: les aflatoxines (af b1, b2, g1 et g2), l'acide kojique, l'acide penicillique, la fumonisine b1 (fb1), la citrinine, la patuline et la zearalenone ; des essais concernant leur stabilite durant la conservation ont montre que seules l'af, la fb1 et la zearalenone peuvent representer un risque mycotoxique ; enfin, la comparaison de 11 fruits secs (oleagineux et riches en glucides) a montre que le cerneau est favorable a la production d'af. La seconde etape, concernant l'etude depuis le verger jusqu'a la conservation, a montre que la contamination n'est effective qu'apres contact avec le sol, le lavage et l'enoisage. Des contaminations avec des souches toxinogenes, a certaines etapes critiques, ont confirme l'absence de risque au niveau du verger ; en revanche, la contamination fongique et aflotoxinique de la figue s'effectue a ce stade. Pour la noix, le risque mycotoxique existe quand le delai de contact au sol et celui precedant le sechage sont prolonges. Des simulations de conservation du cerneau a 5 activite en eau (a#w) ont permis de determiner une a#w minimale de 0,80 empechant tout developpement fongique. Cette strategie d'etude, decrite pour le cas particulier de la noix, peut etre transposee a d'autres substrats susceptibles d'etre contamines par les mycotoxines. Enfin, la fluorescence jaune, associee a la presence d'af et la fluorescence permet d'eliminer les figues a haut risque

A contaminação do amendoim com aflatoxinas é objeto de estudo de vários trabalhos de pesquisa. Entretanto, não consta em trabalhos publicados no país, o estudo da distribuição da contaminação com aflatoxinas entre embalagens de produtos processados que apresentam grãos inteiros e grãos triturados, assim como a estimativa da ingestão de aflatoxinas oriundas do consumo de produtos de amendoim. O objetivo deste trabalho foi investigar a ocorrência e a distribuição da contaminação com aflatoxinas entre embalagens de produtos de amendoim comercializados em estabelecimentos da cidade de Piracicaba – SP, além de caracterizar as amostras quanto a dados qualitativos e quantitativos e condições do ambiente no local da amostragem, assim como estimar a ingestão de aflatoxinas através de produtos de amendoim. A análise qualitativa visual das embalagens não constata esta como fonte de favorecimento da contaminação por aflatoxinas. Quantitativamente, a análise das faixas dos pesos líquidos dos produtos corresponderam aos pesos declarados nas embalagens dos produtos. A atividade de água, mostraram valores normalmente não altos o suficiente para permitirem crescimento fúngico e somente em alguns produtos específicos, os valores poderiam proporcionar o crescimento fúngico. Quanto aos parâmetros temperatura e umidade relativa, observou-se que a temperatura em vários locais amostrados poderia favorecer o crescimento fúngico, enquanto que a umidade relativa não demonstrou valores favoráveis. Os dados de contaminação com aflatoxina mostraram que os produtos comercializados, em alguns casos, ainda apresentam contaminação acima do permitido pela legislação brasileira e que estabelecimentos de diferentes portes apresentaram mesma freqüência e nível de contaminação. O estudo da distribuição da contaminação mostrou que pode ocorrer uma distribuição bastante diferente entre embalagens do mesmo lote, também no produto processado a partir de amendoim triturado, e a detecção da contaminação com aflatoxinas nesses produtos, mostra ser de mais fácil detecção, do que em produtos processados que utilizam amendoim inteiro ou parcialmente inteiro. A avaliação do consumo de produtos de amendoim mostrou níveis de consumo diferentes dos obtidos através de dados de literatura, ressaltando a importância de se trabalhar com dados realísticos de consumo para estimar a ingestão diária provável de aflatoxina AFB1. A estimativa da ingestão diária provável (IDP) de aflatoxina AFB1, mostrou ser inferior a ingestão diária aceitável (IDA) proposta na literatura, ressaltando que as (IDP) relatadas neste estudo foram somente devido ao consumo de produtos de amendoim e não em relação a dieta completa da população.
The peanut contamination with aflatoxins has already been objective of several researches. However there is no published article dealing with the aflatoxin contamination distribution among packages of processed peanut products, containing hole or smashed grains of peanut, neither with the estimate of ingestion of aflatoxins related to peanut products. The objective of this research study was to investigate the occurrence and the widespread-distribution of contamination with aflatoxins in packages of processed peanut products available on the market in Piracicaba city – state of São Paulo and to characterize the samples in terms of: the aspect of the package by visual inspection, the actual weight in comparison to the weight printed on the labels, the water activity of the peanut products and the ambient conditions (temperature and relative humidity) in the sampled stores, as well as to estimate the ingestion of aflatoxin AFB1 through peanut products. In general, based on the visual inspection of the aspect of the peanut products sampled in this study, the packages were not considered a potential source of contamination with aflatoxins. Also, the check of the actual net weight of the products in comparison to the declared weight on the product labels didn’ t show harmfull weight variations to the consumers. The analytical results for water activity showed that in general the values of aw of the products were not high enough to allow the fungic growth-development and the values of water activity could allow the fungic growth. With respect to the temperature and relative humidity it was observed that the temperature of many stores could favor the fungic growth while that relative humidity wasn‘t show values favorable to the fungic growth. The results of contamination with aflatoxins showed that there were some cases which showed contamination with aflatoxin above accepted levels by the Brazilian legislation and that establishment of different size showed one same frequency and level of contamination by aflatoxin. The research of the distribuition of contamination showed that can occur on distribuition enough different among packages of the same lot, also in processed product from peanut products that presented grains crushed, and detection of contamination with aflatoxin in these products, showed to be the more easy detection that in processed products that used grains not crushed or entire grains. The evaluation the consumption the peanut products showed levels differents the levels obtained across the literature datas, to stand up an importance the if to work with real datas of consumption for estimated the probable diary ingestion the aflatoxin AFB1. The estimated of probable diary ingestion (IDP) the aflatoxin AFB1, showed be lower than acceptable diary ingestion (IDA) to proposal by literature, to stand up that as (IDP) reported in this reasearch were only due the consumption the peanut products and not in the statement the completed diet of population.

Le présent travail vise à expliquer les facteurs favorisant la contamination par les aflatoxines dans la filière arachide en Haïti. Les résultats obtenus à partir d’une revue de la littérature, d’observations et d’entretiens avec des acteurs de la filière dans les départements Nord et Nord-Est du pays ont permis de relever différentes pratiques favorables à la contamination des produits telles que : l’absence de rotation culturale, la récolte précoce ou tardive, le séchage et le stockage inadéquats, la faible rigueur dans la sélection des arachides pendant et après la récolte. D’autres pratiques néfastes comme le mouillage des arachides et le mélange incontrôlé voire délibéré d’arachides de bonne et de mauvaise qualités augmentent substantiellement les risques de contamination pendant la commercialisation. Les résultats des tests d’aflatoxines réalisés sur certains produits ont montré des taux élevés allant de 22 ppb à 36 864 ppb pour 55 des 100 échantillons collectés sur le terrain, ce qui témoigne du grave problème auquel est confrontée la filière. Plusieurs facteurs à la base des mauvaises pratiques et de la contamination des produits ont été analysés. Ces facteurs sont d’ordre organisationnel, socioéconomique, institutionnel, politique, technologique et environnemental. Nous avons ainsi pu construire un schéma systémique qui montre comment ces multiples facteurs se conjuguent pour entrainer des pratiques qui favorisent la contamination par les aflatoxines et fourni des pistes d’intervention pour une amélioration de la qualité des produits dans la filière.
Le présent travail vise à expliquer les facteurs favorisant la contamination par les aflatoxines dans la filière arachide en Haïti. Les résultats obtenus à partir d’une revue de la littérature, d’observations et d’entretiens avec des acteurs de la filière dans les départements Nord et Nord-Est du pays ont permis de relever différentes pratiques favorables à la contamination des produits telles que : l’absence de rotation culturale, la récolte précoce ou tardive, le séchage et le stockage inadéquats, la faible rigueur dans la sélection des arachides pendant et après la récolte. D’autres pratiques néfastes comme le mouillage des arachides et le mélange incontrôlé voire délibéré d’arachides de bonne et de mauvaise qualités augmentent substantiellement les risques de contamination pendant la commercialisation. Les résultats des tests d’aflatoxines réalisés sur certains produits ont montré des taux élevés allant de 22 ppb à 36 864 ppb pour 55 des 100 échantillons collectés sur le terrain, ce qui témoigne du grave problème auquel est confrontée la filière. Plusieurs facteurs à la base des mauvaises pratiques et de la contamination des produits ont été analysés. Ces facteurs sont d’ordre organisationnel, socioéconomique, institutionnel, politique, technologique et environnemental. Nous avons ainsi pu construire un schéma systémique qui montre comment ces multiples facteurs se conjuguent pour entrainer des pratiques qui favorisent la contamination par les aflatoxines et fourni des pistes d’intervention pour une amélioration de la qualité des produits dans la filière.
This work aims to explain the factors that facilitate aflatoxins contamination in the peanut sector in Haiti. The results obtained from by literature review, observations and interviews with actors in North and North-East areas of the country have revealed several practices favorable to the contamination of products such as : lack of crop rotation, early or late harvest, inadequate drying and storage, poor rigor in peanut selection during and after harvest. Other harmful practices such as wetting peanut and uncontrolled or even deliberate mixing of good and poor-quality peanuts substantially increase the risk of contamination during marketing. Aflatoxin test results on some products showed high rates ranging from 22 ppb to 36 864 ppb in 55 out of 100 field-collected samples, indicating the serious quality problem in the chain. Several factors sustaining the bad practices and product contamination were analyzed. Dimensions are organizational, socio-economic, institutional, political, technological and environmental. We have thus been able to build a systemic diagram that shows how these multiple factors combine to lead practices that strengthen aflatoxins contamination and provided paths of intervention for improving products quality in the commodity chain.
This work aims to explain the factors that facilitate aflatoxins contamination in the peanut sector in Haiti. The results obtained from by literature review, observations and interviews with actors in North and North-East areas of the country have revealed several practices favorable to the contamination of products such as : lack of crop rotation, early or late harvest, inadequate drying and storage, poor rigor in peanut selection during and after harvest. Other harmful practices such as wetting peanut and uncontrolled or even deliberate mixing of good and poor-quality peanuts substantially increase the risk of contamination during marketing. Aflatoxin test results on some products showed high rates ranging from 22 ppb to 36 864 ppb in 55 out of 100 field-collected samples, indicating the serious quality problem in the chain. Several factors sustaining the bad practices and product contamination were analyzed. Dimensions are organizational, socio-economic, institutional, political, technological and environmental. We have thus been able to build a systemic diagram that shows how these multiple factors combine to lead practices that strengthen aflatoxins contamination and provided paths of intervention for improving products quality in the commodity chain.

53% du lait produit à Veracruz est utilisé pour la fabrication des fromages, mais ce lait n’est soumis à aucune analyse de la qualité, de sorte que le contenu des polluants tels que les pesticides et les mycotoxines pourraient se concentrer sur les fromages. Les pesticides, tels que les organochlorés (POC), peuvent augmenter l'incidence du cancer et jouer le rôle de perturbateur endocrinien ; tandis que les mycotoxines, telles que les aflatoxines (AF): l'aflatoxine M1 (AFM1) et l'aflatoxine M2 (AFM2), sont considérées comme cancérigènes. La consommation de fromage avec ces toxines peut mettre en danger la santé du consommateur, cependant, il y a peu de rapports sur la concentration de ces composés et la consommation de fromages par la population de Veracruz. L'objectif de ce travail était d'évaluer le risque pour la santé dû à la consommation de fromage (frais et Oaxaca) contaminé par les POC, AFM1 et AFM2. Pour atteindre cet objectif, des fromages frais et Oaxaca ont été échantillonnés au hasard dans 40 points de vente dans la ville de Veracruz, au cours de trois années différentes (2014, 2015 et 2016), couvrant les périodes sèches et pluvieuses. La concentration de POC (n = 20) a été quantifiée par CG-MS et celle des AF par HPLC. Pour obtenir des données sur la consommation de fromage, deux questionnaires ont été utilisés: un pour la fréquence d'achat (n = 100) et un journal alimentaire pour 7 jours (n = 309 pour chaque type de fromage). L'évaluation des risques par exposition de la population à l'AFM1 a été évaluée à l'aide de la combinaison des fonctions de densité de probabilité (PDF) de la concentration AF, de la consommation de fromage Oaxaca et du poids corporel (méthodologie probabiliste). Seulement dans 5% des échantillons de fromage Oaxaca, les POC ont été détectés mais dans des valeurs inférieures à la limite de quantification (LOQ = 0,01 mg kg-1), ce qui montre que l'exposition des consommateurs aux POC est faible. La consommation moyenne de fromage frais et Oaxaca était de 50,9 g personne-1 d-1 et 47,8 g personne-1 d-1, respectivement. Seulement dans 37% des échantillons de fromage Oaxaca, la concentration d'AFM1 et dans 10% pour l'AFM2, était supérieure à la valeur établie par la Commission européenne de 0,05 μg kg-1. L'exposition des enfants (5,9 ug de poids corporel AFM1 1 kg d-1 et 0,5 pg de poids corporel 1 AFM2 kg d-1) était supérieure à celle observée chez les adultes (2,32 g AFM1 kg de poids corporel -1 d-1 et 0,2 μg de AFM2 kg de poids corporel-1 d-1). La prise en compte d'un TDI de 1 ng kg-1 pc par jour-1 ,les résultats indiquent que la santé de 72,8% et 51% des enfants et des adultes respectivement population de la ville de Veracruz, est à risque de manger du fromage Oaxaca contaminés par l'AFM1 et environ 13% et 3% respectivement de la population des enfants et des adultes, en raison de la consommation de AFM2. La présente étude est la première enquête sur l'exposition de la population de Veracruz à des fromages contaminés par AFM1 et AFM2
About 53 % of the milk produced in Veracruz is used for the manufacture of artisanal cheeses, however this milk is not subjected to any quality analysis, so the content of contaminants such as pesticides and mycotoxins could be found in cheeses. Pesticides, such as organochlorines (POCs), can increase the incidence of cancer and act as an endocrine disruptor; Additionally mycotoxins, such as aflatoxin M1 (AFM1) and aflatoxin M2 (AFM2), are considered carcinogenic. The consumption of cheese with these toxins can put the health of the consumer at risk; however, there are only reports on the concentration of these compounds in cheeses, and cheese data consumption by the population of Veracruz are not available. The objective of this work was to evaluate the health risk due to the consumption of contaminated cheese (fresh and Oaxaca) with POC's and AFM1. In order to attain this objective, fresh and Oaxaca cheeses were randomly sampled at 40 groceries in Veracruz city, in three different years (2014, 2015 and 2016), covering periods of drought and rain. The concentration of POC's (n= 20) was quantified by CG-MS and that of AFs by HPLC. Cheese intake data were obtained from a purchase frequency questionnaire (n = 100) and from a 7 days food diary questionnaire (n = 309 for each type cheese). The risk assessment for exposure of the population to AFM1 was calculated by the combination of probability density functions (PDF) of AFM1 concentration, Oaxaca cheese consumption and body weight (probabilistic methodology). POCs were detected only in 5 % of the samples at concentrations above the quantification limit (LOQ = 0.01 mg kg-1), which showst that consumer exposure is low. The average consumption of fresh and Oaxaca cheese was 50.9 g person-1 d-1 and 47.8 g person-1 d-1, respectively. Only in 37 % of Oaxaca cheese samples, the concentration of AFM1 and in the 10 samples of AFM2 was higher than the value established by the European Commission of 0.05 μg kg-1. The exposure of children (5.9 μg AFM1 kg body weight-1 d-1 and 0.5 μg AFM2 kg body weight-1 d-1) was higher than found in adults (2.32 μg AFM1 kg body weight-1 d-1 and 0.2 μg AFM2 kg body weight-1 d-1). Taking into account a TDI of 1 ng kg-1 pc d-1 the results indicated that health that the 72.8 % And 51% of the adult and child population, respectively, of the population of Veracruz city is at for consuming contaminated Oaxaca cheese with AFM1 and about 13% and 3% of the infant and adult population, respectively, due to the consumption of AFM2. The present study is the first research on the exposure of Veracruz population concerning contaminated cheeses with AFM1 and AFM2
El 53 % de la leche producida en Veracruz se utiliza para fabricación de quesos artesanales, sin embargo esta leche no es sometida a ningún análisis de calidad, por lo que el contenido de contaminantes como plaguicidas y micotoxinas podrían concentrarse en los quesos. Los plaguicidas, como los organoclorados (POC’s), pueden aumentar la incidencia de cáncer y funcionar como disruptor endócrino; mientras que las micotoxinas, como las aflatoxinas (AFs): Aflatoxina M1 (AFM1) y Aflatoxina M2 (AFM2), son consideradas cancerígenas. El consumo de queso con estos tóxicos puede poner en riesgo la salud del consumidor, sin embargo, existen pocos reportes de la concentración de estos compuestos y el consumo de quesos por la población de Veracruz. El objetivo de este trabajo fue evaluar el riesgo a la salud por el consumo de queso (fresco y Oaxaca) contaminado con POC’s, AFM1 y AFM2. Para alcanzar este objetivo, se muestrearon aleatoriamente quesos fresco y Oaxaca en 40 puntos de venta de la ciudad de Veracruz, en tres años diferentes (2014, 2015 y 2016), abarcando periodos de secas y de lluvias. La concentración de POC’s (n = 20) se cuantificó por CG-MS y la de AFs por HPLC. Para la obtención de datos de ingesta de queso, se utilizaron dos cuestionarios: uno de frecuencia de compra (n = 100) y un diario de alimentos de 7 días (n = 309 para cada tipo de queso). La evaluación del riesgo por exposición de la población a AFM1 se evaluó a través de la combinación de las funciones de densidad de probabilidad (PDF) de la concentración de AFs, del consumo de queso Oaxaca y del peso corporal (metodología probabilística). Sólo en el 5 % de las muestras de queso Oaxaca se detectaron POC’s pero en valores inferiores al límite de cuantificación (LOQ = 0.01 mg kg-1), lo que muestra que la exposición del consumidor a los POC’s es baja. El consumo promedio de queso fresco y Oaxaca fue de 50.9 g persona-1 d-1 y 47.8 g persona-1 d-1, respectivamente. Sólo en el 37 % de las muestras de queso Oaxaca, la concentración de AFM1 y en el 10 % para la AFM2, fue mayor al valor establecido por la Comisión Europea de 0.05 μg kg-1. La exposición de los niños (5.9 μg de AFM1 kg de peso corporal-1 d-1 y 0.5 μg de AFM2 kg de peso corporal-1 d-1) fue mayor que la encontrada en adultos (2.32 μg de AFM1 kg de peso corporal-1 d-1 y 0.2 μg de AFM2 kg de peso corporal-1 d-1). Teniendo en cuenta una TDI de 1 ng kg-1 pc día-1 los resultados indicaron que la salud del 72.8 % y el 51 % de la población infantil y adulta, respectivamente, de la ciudad de Veracruz, está en riesgo por consumir queso Oaxaca contaminado con AFM1 y cerca del 13 % y 3 % de la población infantil y adulta respectivamente, debido al consumo de AFM2. El presente estudio es la primera investigación sobre la exposición de la población veracruzana a quesos contaminados con AFM1 y AFM2

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Coordena??o de Aperfei?oamento de Pessoal de N?vel Superior - CAPES
Mycotoxins are natural contaminants produced by filamentous fungi and they are widely present in food. In wheat grains, the presence of trichothecenes produced by Fusarium spp. and also aflatoxins, produced by Aspergillus spp. are a serious public health problem because they are toxic metabolites to man and animals that remain stable during the industrial processes to which wheat is subjected when manufacturing derived products. One way to reduce such contaminants is to use ozone (O3) in food processing. Due to its high potential as an oxidant, O3 may react with mycotoxins and reduce their toxicity due to molecular degradation. This research aimed to evaluate the effects of different conditions of ozonation on the i) reduction in mycotoxins levels (deoxynivalenol and total aflatoxins); ii): reduction in total fungal count; iii) the effects on chemical, mineral and technological profiles and; iv) the possible differences sensorial profile of the flour obtained from the ozonized grains. In a second phase of this project, a methodology for determination of D3G (deoxynivalenol-3-glucoside), a masked form of DON, using High Performance Liquid Chromatography with photodiode array detector (HPLC-PDA) was optimized and in-house validated. Results obtained from ozonation study showed that O3 reduced total fungal count in approximately 3.0 cycles log CFU/g of wheat grain and deoxynivalenol and total aflatoxins contamination up to 64.3% and 48.0%, respectively. The gaseous ozonation can be applied without negatively changing the chemical, technological and sensory characteristics of the grains and can be considered an excellent method for remediation of fungal and mycotoxin contaminations. Also, the method optimized and in-house validated for determination of D3G by HPLC-PDA showed adequate results and, it could be considered an alternative to mass spectrometry determination of D3G in wheat grains.
Micotoxinas s?o contaminantes naturais, produzidos por fungos filamentosos e, podem ocorrer em altos n?veis nos alimentos. Nos gr?os de trigo, a presen?a de tricotecenos, um grupo de micotoxinas produzidas por Fusarium spp. e, aflatoxinas, produzidas por Aspergillus spp., representam um importante problema de sa?de p?blica por serem t?xicas ao homem e animais e muito est?veis aos processos no qual o trigo ? submetido para obten??o de produtos industrializados. Uma forma de reduzir a contamina??o dos alimentos por micotoxinas ? atrav?s do uso do oz?nio (O3) no processamento do alimento. Devido ao alto potencial oxidante do O3, esse pode degradar as mol?culas das micotoxinas, tendo como consequ?ncia a elimina??o ou redu??o de seus efeitos t?xicos. Essa pesquisa teve como objetivos principais avaliar os efeitos de diferentes condi??es de ozoniza??o na i) redu??o de micotoxinas (desoxinivalenol e aflatoxinas) em gr?os de trigo; ii) redu??o nos n?veis de fungos filamentosos; iii) influ?ncia nos par?metros qu?micos, perfil de minerais e par?metros tecnol?gico dos gr?os e da farinha obtida ap?s o processamento e; iv) influ?ncia nas caracter?sticas sensoriais da farinha elaborada a partir dos gr?os ozonizados. Em uma segunda etapa do projeto, um m?todo para determina??o de uma forma modificada do desoxinivalenol (DON), o desoxinivalenol-3-glicos?deo (D3G) foi otimizada e validada intralaboratorialmente, utilizando cromatografia l?quida de alta efici?ncia com detector de arranjo de diodos (CLAE-DAD). Os resultados obtidos dos ensaios de ozoniza??o demonstraram que o O3, nas condi??es experimentais utilizadas, reduziu a contagem de fungos totais em cerca 3,0 logs UFC/g de gr?os e a contamina??o por desoxinivalenol e aflatoxinas totais em at? 64,3 % e 48,0 %, respectivamente. O processo de ozoniza??o n?o influenciou de modo negativo a qualidade qu?mica, tecnol?gica e sensorial dos gr?os de trigo, podendo ser utilizado como um excelente m?todo para remedia??o da contamina??o dos gr?os por fungos e micotoxinas. Resultados adequados tamb?m foram obtidos na valida??o do m?todo de determina??o de D3G por CLAE-DAD, demonstrando que o m?todo ? confi?vel para a determina??o dessa forma mascarada do DON em gr?os e trigo e, pode ser utilizado como um m?todo alternativo a espectrometria de massas para tal an?lise.

CNPQ
O leite é uma das principais fontes de nutrientes da dieta humana e é um alimento que acompanha o ser humano durante toda a vida, tanto como leite de consumo como através de seus derivados. O leite pode apresentar contaminação oriunda de diversas fontes, relacionados desde a ordenha até o processo de beneficiamento do mesmo. Dentre esses contaminantes, destacam-se as micotoxinas, que são metabólitos tóxicos produzidos por fungos em condições de estresse. Estudos correlacionam a ocorrência de micotoxinas em leite e derivados lácteos, com ênfase na aflatoxina M1 (AFM1) que é regulamentada pela legislação vigente. Apesar da maior parte da literatura afirmar que a aflatoxina B1 (AFB1) é completamente convertida em AFM1, resultados preliminares tem sugerido que isso não é verdadeiro, o que vem a justificar que a mesma passe, também, a ser analisada, uma vez que a toxicidade da AFB1 é maior que a da AFM1. Na área de leite e derivados lácteos, estudos têm sugerido que as aflatoxinas, em especial a AFM1, localizam-se predominantemente nas frações proteicas. Porém, a elucidação da interação entre AFM1 e fração proteica permanece sem conclusão definitiva, o que sugere um campo amplo de estudo nesta área. Ainda, tendo em vista que a concentração de aflatoxinas é predominante na fração proteica e que pode haver uma interação química entre esta fração e aflatoxinas, os tratamentos térmicos aos quais o leite é submetido podem causar alterações significativas nas estruturas dessas proteínas e, por conseguinte, nas interações entre proteínas e AFM1 e AFB1. Portanto, o entendimento da natureza das ligações entre as AFM1 e AFB1 às proteínas do leite é imprescindível para a compreensão da biodisponibilidade dessas micotoxinas em animais e seres humanos. Desta forma, o objetivo do trabalho foi verificar o efeito do tratamento térmico e da digestibilidade sobre a interação proteína-aflatoxina. Para isso, foram contaminados amostras de solução de caseína bovina e leite desnatado com AFB1 (200 µg.L-1) e AFM1 (20 µg.L-1), posteriormente submetidas a pasteurização e digestão in vitro. As determinações realizadas consistiram na quantificação das AFLB1 e M1 por Cromatografia à Líquido de Ultra Alta Eficiência, Espectroscopia na região do infra vermelho (FTIR), Calorimetria Exploratória Diferencial (DSC) e Espectroscopia de fluorescência. Não houve redução da concentração de AFB1 e AFM1 após o tratamento térmico e a bioacessibilidade foi de 91,2% 70,5% e 90,9% e 69,7%, respetivamente, para caseína bovina e leite desnatado. Avaliando quantitativamente as estruturas secundárias da solução de caseína bovina e leite desnatado contaminados com AFB1 e AFM1 as principais mudanças ocorreram nas estruturas β-volta, β-anti e β-folha. Com a análise de DSC observou-se picos exotérmicos para ambos os ensaios, com variação de entalpia (∆H) após cada ensaio. Em relação aos espectros de fluorescência, foi possível verificar que as AFB1 e AFM1 promoveram redução da fluorescência original das proteínas lácteas. Pelos resultados obtidos, é possível concluir que a concentração das AFB1 e M1 não foram reduzidas pela temperaturaempregada, provavelmente pela interação proteínas do leite-aflatoxina, resultando numa bioacessibilidade relativamente elevada.
Milk and derivates are one of the main sources of nutrients of the human diet and is a food that follow's man during all of your life. The milk may be contaminated from several sources, related since milking until the beneficiation process. Among these contaminants, highlights are the mycotoxins, which are toxic metabolites produced by fungus in conditions of stress. Studies related to the occurrence of mycotoxins in milk and milk products, with the emphasis on aflatoxin M1 (AFM1) which is ruled by Brazilian law. While most of the literature stating that aflatoxin B1 (AFB1) is completely converted into AFM1, preliminary results have suggested that this is not true, justifying that it passes also to analyze because the toxicity of AFB1 is greater than the AFM1. In the milk and dairy products, studies have suggested that aflatoxins, in particular AFM1, are located predominantly in protein fractions. However, the elucidation of the interaction between AFM1 and protein fraction remains without definitive conclusion, suggesting a large broad field of study in this area. Also considering that the concentration of aflatoxins are prevalent in the protein fractions and can have a chemical interaction between these fractions and aflatoxins, the thermal treatments that the milk is subjected can cause significant changes in these proteins structures and consequently in interactions between proteins and AFM1 and AFB1. Therefore, the knowledge of nature of the links between AFM1 and AFB1 to milk proteins is indispensable for understanding the bioavailability of these mycotoxins in animals and humans. Thus, the aim of this study was to investigate the effect of thermal treatment and the digestibility of the interaction protein-aflatoxin. For this, samples of bovine casein solution and skimmed milk were contaminated with AFB1 (200 μg.L-1) and AFM1 (20 μg.L-1), and then submitted to pasteurization and in vitro digestion. The carried analyzes consisted in quantifying AFB1 and AFM1using high performance liquid chromatography, Fourier transform infrared spectroscopy (FTIR), Differential Scanning Calorimetry (DSC) and fluorescence spectroscopy. There was no reduction in the concentration of AFB1 and AFM1 and after the thermal treatment the bioaccessibility was 91.2% and 70.5%, 90.9% and 69.7%, respectively, for bovine casein and skim milk. In quantitative assessments of the secondary structures of bovine casein and skim milk contaminated with AFB1 and AFM1 the major changes occurred in the β-turn structures, β-anti and β-sheet. With the DSC analysis was observed exothermic peaks for both tests, the change of enthalpy (∆H) after each test. Regarding the fluorescence spectra, we found that AFB1 and AFM1 promoted reduction of the original fluorescence of milk proteins. From the results, it may be concluded that the concentration of AFB1 and M1 not been reduced by the temperature applied, probably by the interaction between milk protein and aflatoxin, resulting in a relatively high bioaccessibility.

>Magister Scientiae - MSc
Subsistence farmers may contribute significantly to food production, food security, and employment in South Africa. However poor storage practices and contamination with mycotoxins, particularly fumonisins and aflatoxins impacts adversely on production, food safety and food security. Mycotoxins are toxic natural food-borne compounds which frequently contaminate agricultural produce worldwide. They are hazardous to humans and animals and result in significant production losses for farmers. This study focused on former Bantustans in Northern South Africa, namely Vhembe District Municipality (Limpopo) and Gert Sibande District Municipality (Mpumalanga). The aim was to assess mycological and mycotoxin contamination of crops grown by subsistence farmers. A semi-structured questionnaire was administered to randomly thirty-nine households. Data on demographics, storage practices and production during period of 2011 and 2012 cropping seasons were collected. One hundred and fifteen (115) crop samples (maize, beans and peanuts) were collected for analysis. Standard mycological methods and validated mycotoxin analysis methods (HPLC and LC- MS/MS) were used. It was found that maize was the staple food in both provinces, with a significant difference (p = 0.0184) in its production between the two districts; Vhembe produced 0.6 tonnes compared to 2.4 tonnes in Gert Sibande. The majority of the farmers for storage used traditional open wooden cribs (15/20) and steel tanks (5/20) while VDM farmers used sealed store houses 5/19 and 15/19 used polystyrene sacks. Aflatoxin occurrence was low with <1% of GSDM samples contaminated compared to 11% of VDM samples. No significant difference (p > 0.05) was observed in the aflatoxin contamination in VDM samples between the year 2011 and 2012. Samples from VDM households had higher Aspergillus fungal infection (maximum incidence 69%) compared to GSDM (27%) over both seasons. The most frequently isolated Fusarium species in VDM samples was F. verticillioides (92%; 93%), and F. subglutinans (97%; 80%) in GSDM samples over seasons 2011 and 2012, respectively. Highest levels of fumonisins (FB1+ FB2) ranged between 1010 μg/kg and 12168 μg/kg with less than 30% extremely contaminated above the regulated limit in 91% of samples from Limpopo over both seasons (2011 and 2012). Fumonisin levels between the two seasons in VDM showed no significant difference (p>0.05). Only three (less than 5%) from 68% GSDM contaminated maize samples were above the FB1 and FB2 limit. In 2011, there were two highly contaminated maize samples (1762 μg/kg and 4598 μg/kg) with the other samples less than 600 μg/kg, whereas in season two (2012) all samples were below 200 μg/kg, except one highly contaminated sample (26115 μg/kg). None of the beans and peanuts from Mpumalanga was contaminated with mycotoxins above the recommended limit, but from Limpopo 1/5 peanuts was found contaminated with aflatoxin G1 (41 μg/kg). Natural occurrence and contamination of both fumonisin and aflatoxin in stored home-grown maize from VDM was significantly (p < 0.0001) higher than GSDM over both seasons. In general, Limpopo farmers’ experience lower harvests and greater mycotoxin contamination of agricultural produce. This may be attributed in part to poor storage practices and environmental and climatic conditions in that agro-ecological zone.

Les mycotoxines sont des métabolites secondaires toxiques produits par des moisissures appartenant principalement aux genres Aspergillus, Pénicillum, Fusarium, naturellement présentes dans l’air ambiant et le sol. Parmi elles, certaines (les aflatoxines, l’ochratoxine A, le désoxynivalenol. . . ) présentent des risques importants (activités mutagènes, cancérigènes, tératogènes et immuno-toxinogènes) pour la santé publique et des organismes internationaux ont fixé des valeurs toxicologiques de références (DJT, DHTP) représentant la quantité que le consommateur peut consommer tous les jours de sa vie sans courir de risque pour sa santé. Pour caractériser le risque, il est nécessaire de connaître outre les valeurs toxicologiques de références, les quantités auxquelles le consommateur est exposé. Cette étude permet de mesurer pour la première fois au Liban, l’exposition du consommateur (Beyrouthin, Adulte 25-54 ans) à trois familles de mycotoxines, les ochratoxines, les trichothécènes et les aflatoxines par une approche de la distribution simple à partir de la méthode de la ration totale, basée sur le panier de la ménagère. Les résultats obtenus montrent que les mycotoxines sont présents, d’une façon générale dans les aliments consommés par les sous groupes de la population libanaise étudiée, à des taux conformes aux normes nationales et européennes en vigueur. Cependant, des cas de dépassement des limites maximales de certaines mycotoxines (Aflatoxine Mi, DON, OTA) dans respectivement certains aliments analysés (lait et boissons à base de lait, pains et biscottes et biscuits et viennoiseries) ont été observés. Compte tenu du niveau d’exposition à OTA et au DON, il y a risque de dépassement des Doses Journalières Tolérables respectives. De plus, il semble nécessaire que soit portée une attention particulière à l’exposition aux aflatoxines de certains groupes de populations tels que les forts consommateurs pour lesquels le risque d’être exposés à un niveau d’aflatoxine Bi et Mi n’est pas nul. Le café, les produits céréaliers (pains et biscottes, biscuits et viennoiseries) et les boissons alcoolisées sont parmi les principaux contributeurs de l’exposition à IOTA, le lait et les boissons à base de lait et le yaourt pour l’AFM1, les produits céréaliers (pains et biscottes) pour l’AFB1 et les produits céréaliers (pains et biscottes et riz et produits à base de riz) pour le DON. Ceci souligne la nécessité de l’établissement des plans de contrôle et de surveillance pour ces substances dans les différentes denrées alimentaires afin de protéger la santé des consommateurs
Mycotoxins are toxic secondary metabolites produced by fungi belonging mainly to the genera Aspergillus, Penicillium, Fusarium, naturally present in ambient air and soil. Some of them (aflatoxins, ochratoxin A, deoxynivalenol. . . . ) pose significant risks (activities mutagenic, carcinogenic, teratogenic and immuno-toxigenic) to public health. International agencies have set reference toxicological values (TDI, PTWI) representing the amount of food that consumers can eat all the days of his life without running any risk to their health. To characterize the risk, it is also necessary to know the toxicological reference values, the quantities which the consumer is exposed. This study provides a measure for the first time in Lebanon, consumer exposure (for Beirut consumer, Adults 25-54) to three families of mycotoxins, ochratoxin, trichothecenes and aflatoxins by a simple distribution approach of total diet study, based on household basket. The results show that mycotoxins are present, generally in food consumed by subgroups of the Lebanese population studied, at levels consistent with national and European regulations. However, an exceeding of maximum limits of certain mycotoxins (Aflatoxin M1, DON, OTA) respectively in certain analyzed foods (milk and milk-based beverages, breads and crackers, cookies and pastries) were observed. Given the level of exposure to OTA and DON, there is a risk of exceeding the tolerable daily intake respectively. Moreover, it seems necessary that special attention must be paid to exposure to aflatoxins in certain population groups such as high consumers for whom the risk of being exposed to a level of aflatoxin B1 and M1 is not zero. Coffee, wheat based products (breads, crackers, cookies and pastries) and alcoholic beverages are among the main contributors to OTA exposure, milk and milk beverages and yogurt to AFM1, wheat based products (breads and crackers) to AFB1 and wheat based products (breads, crackers and rice products and rice) to DON. This underlines the need for establishing control and surveillance plans for these substances in different foodstuffs to protect consumers’ health

Un des dangers chimiques les plus importants relevés par l'OMS concerne la contamination des céréales par les mycotoxines et notamment les aflatoxines et les fumonisines. La réglementation recommande des contaminations maximales d'aflatoxines dans les céréales inférieures à 20 mg/kg ; cependant on relève couramment des taux supérieurs à 200 mg/kg dans le maïs au Mexique. Bien qu'il ait été évalué que le processus de nixtamalisation détruit plus de 90 % des fumonisines et de 80 à 95 % des aflatoxines, le taux résiduel peut encore être élevé : certaines publications rapportent des concentrations jusqu'à 100 mg/kg dans les tortillas, ce qui représente un risque avéré vue la grande consommation de tortillas au Mexique (325g/j). Le JECFA (2001) a établi une dose maximale acceptable de 2μg/kg pc/j pour la fumonisine et recommande de réduire l'exposition aux aflatoxines au plus faible niveau possible en considérant que le seuil de 1 ng/kg pc/j ne devrait pas être atteint. Au cours de cette thèse 3 enquêtes aléatoires et représentatives ont été menées dans 40 tortillerias de la ville de Veracruz. La consommation de maïs de la population a été évaluée à partir d'un questionnaire de consommation. L'analyse des mycotoxines a été réalisée par HPLC-FD par utilisation de colonnes à immunoaffinité selon la réglementation européenne (CIRAD-Montpellier). L'analyse des données obtenues a été effectuée selon une méthode probabiliste permettant de construire une fonction de distribution de probabilités à partir de la méthode de Monte Carlo (UBO). La représentativité de la population a été validée à partir d'évaluation de quotas de population après échantillonnage aléatoire initial. La contamination des tortillas a été mesurée à 0.54-1.96 mg/kg pour les aflatoxines et à 65-136 mg/kg pour les fumonisines. La consommation moyenne de tortillas a été mesurée à 148 g de maïs par jour. L'exposition de la population aux aflatoxines apparaît alors comprise entre 0,94 et 3,14 ng/kg pc/j et celle aux fumonisines entre 146 et 315 ng/kg pc/j. Les échantillons les plus contaminés proviennent des tortillerias réalisant elles-mêmes leur procédure de nixtamalisation. L'analyse des résultats montre que 60 % de la population de Veracruz serait à risque selon les préconisations du JECFA. L'exposition aux fumonisines atteint 5 % de la dose maximale acceptable, du fait d'une relativement faible contamination du maïs à cette mycotoxine. Les résultats montrent donc un risque sanitaire pour la population de la ville de Veracruz. Une extension de ce travail à la totalité de l’Etat de Veracruz, incluant la population rurale, devrait être menée du fait du risque probablement accru de cette dernière catégorie de population en lien avec sa plus forte consommation de maïs
One of the chemical hazards that WHO has reported more frequently is cereals contamination with mycotoxins, mainly aflatoxins and fumonisins. NOM-188-SSA1-2002 establishes that aflatoxin concentration in grain should not exceed 20 mg kg-1 ; however, there are reported concentrations > 200 mg kg-1 in maize. Although it has been documented that nixtamalizacion removes more than 90% of fumonisins and between 80 and 95% of aflatoxins, the residual amount could be important, finding reports concentrations higher than 100 mg kg-1 of aflatoxin in tortilla, representing a risk due to the high consumption of tortillas in Mexico (325 g d-1). The JECFA (2001) establishes a maximum intake of 2 mg kg-1 pc d-1 for fumonisin and aflatoxin recommends reducing “as low as reasonably achievable” levels. 3 random and representative sampling in Veracruz city, each in 40 tortillerias, were made. Corn intake and weight of the population were estimated using a consumption questionnaire. Mycotoxins analysis were performed by HPLC-FD using immunoaffinity columns according to European standard UNE-EN ISO 14123 : 2008 for aflatoxins and UNE-EN 13585 : 2002 for fumonisin in the CIRAD (Montpellier, France). Statistical analysis were performed under a probabilistic approach in collaboration with the University of Bretagne Occidentale (Brest, France), building probability density function (PDF) and using the Monte Carlo method. PDF parameters of the weight of the population was 74.15kg for men (which coincides with reported by CANAIVE) and 65.83kg for women ; the pollution aflatoxin tortilla was 0.54 – 1.96mg kg-1 and fumonisin from 65.46 – 136.00mg kg-1 ; the tortilla consumption was 148.3g of corn per person per day ; the daily intake of aflatoxins was 0.94 – 3.14ng kg-1 bw d-1 and fumonisin of 146.24 – 314.99ng kg-1 bw d-1. Samples with higher aflatoxin contamination came from tortillerias that make the nixtamalization in situ. In assessing exposure it was found that up to 60% of the population could be consuming more than the recommended by JECFA (2001) for aflatoxin dose (1ng kg-1 bw d-1). Exposure to fumonisins intake was < 5% due to low contamination by these mycotoxins. The results suggest that the population of the city of Veracruz could be in food risk by eating contaminated corn tortillas AFT. It is advisable to extend this study in rural communities, where risk factors could increase

M.Sc. (Nanoscience)
Ochratoxin A and Aflatoxin B1 are important food contaminates as they are known to be mutagenic, genotoxic, nephrotoxic, hepatotoxic, immunosuppressive and teratogenic to both animals and humans. These mycotoxins are associated with the contamination of food stuff such as grapes, maize, red pepper, meat, milk, beans and processed products from contaminated raw material. Current physical, biological and chemical methods employed to improve the safety of food often compromise the nutritional value and result in huge losses. The alternative to these treatments are addition of supplements with protective properties to reduce the toxicity of mycotoxins or prevent their formation. The work presented in this dissertation reports an attempt to develop such materials to prevent damage caused by ochratoxin A and aflatoxin B1. This was done through the synthesis; characterisation and cytotoxicity study of chitosan nanoparticles with methanolic plant extracts (L. leonurus, M. longifolia and A. montanus). Inhibition of cellular damage due to mycotoxins for possible application in prevention of cellular damage by mycotoxins also presented. Chitosan nanoparticles were synthesised using an ionic gelation method with sodium triphosphate as the cross linker. The methanolic medicinal plants extracts were incorporated into the chitosan solution before synthesising nanoparticles, and nanoparticle synthesis initiated by the addition of sodium triphosphate solution. The synthesised products were characterised using zetasizer, transmission electron microscopy, x-ray diffraction and Fourier-transform infrared spectroscopy. The extracts’ antioxidant ability was evaluated before incorporation into chitosan using 2, 2-diphenyl- 1-picrylhydrazy (DPPH) radical scavenging assay. This assay was performed using UVvis spectroscopy. The cytotoxicity of the synthesised nanoparticles was assessed using a Vero cell line and by evaluating the cell viability with an MTS assay. The nanoparticles were successfully synthesised and showed the presence of different functional groups as expected. Plain chitosan nanoparticles were roughly spherical shaped and had smooth surfaces, nanoparticles containing extracts similarly were spherical in shape as well but had rougher surfaces when visualised under TEM. All nanoparticles had positive zeta potentials between 26 – 28 mV. The average particle sizes ranged between 31 – 65 nm as measured using TEM and average particle sizes obtained using zetasiser was 78 – 190 nm. The cytotoxicity studies of plain nanoparticles and nanoparticles with extract showed that the synthesised nanomaterials were not toxic even at concentration of 500 μg/ml and less than 20% of the Vero cells were affected under these conditions.

Peanuts contribute significantly to food security in western Kenya due to their high nutritional value and cash crop potential. However, the crop is highly susceptible to aflatoxin contamination. Yet little information is available on the extent of contamination in the region. This study explores the level and extent of contamination of peanuts by aflatoxins, Aspergillus section Flavi, Rhizopus and Penicillium spp. in western Kenya. A survey of 769 households was carried out in the Busia and Homa bay districts of Kenya. Information on peanut pre- and post-harvest practices was collected through person-to-person interviews. Aflatoxin levels of samples collected from each household were determined by indirect competitive ELISA method. Isolation of Aspergillus section Flavi, Penicillium and Rhizopus spp. was done on Modified Dichloran Rose Bengal (MDRB) agar, while identification of specific fungal species was done on Czapek yeast extract agar (CYA). Screening isolates of A. flavus and A. parasiticus for aflatoxin production was done in high sucrose yeast extract (YES) liquid medium, and the aflatoxin types identified on TLC plates, using analytical grades of aflatoxin B1, B2, G1 and G2 as reference standards. Common household preparation techniques (roasting, making peanut paste and boiling peanuts) were evaluated for effectiveness in reducing aflatoxin levels in peanuts. The boiling procedure was modified to test the effect of magadi (locally available salt used mainly to soften legumes, vegetables or maize while cooking), ammonium persulphate and sodium hypochlorite during soaking. Magadi, sodium bicarbonate and locally prepared ash was subsequently used to boil the nuts after soaking. Aflatoxin levels ranged from zero to 7525 ìg/kg. Most samples were safe to consume, based on the European Union and Kenya Bureau of Standards tolerance levels, with 63.7 per cent of all samples having undetectable levels, and only 7.54 per cent being contaminated based on KEBS standards. Peanuts from the Busia district, which has more of Lower Midland 1 (mean annual rainfall of 1600-1800 mm) and Lower Midland 2 (mean annual rainfall of 1300-1700 mm) agro-ecological zones had significantly (÷2=14.172; P=0.0002) higher levels of aflatoxin compared to the Homa bay district, that has more of the drier Lower Midland 3 agroecological zone (mean annual rainfall of 900-1500mm). Improved cultivars had significantly (÷2=9.748; P=0.0018) lower levels of aflatoxin compared to local cultivars. Over 60 per cent of all samples had A. flavus S-strain, A. flavus L-strain and A. niger. A. flavus S-strain was positively correlated with aflatoxin levels. As expected, grading of peanuts post-harvest significantly reduced the incidence of A. flavus S- and L-strains, while peanuts collected from farmers who belonged to producer marketing groups had a significantly lower incidence of A. flavus S- and L-strains, A. niger and Rhizopus spp. The incidence of A. flavus L-strain, A. niger and Rhizopus spp. was significantly higher in local landraces compared to the improved cultivars. Over 60 per cent of isolates produced Aflatoxin B1. Intermediate processes such as sorting and dehusking led to a significant decline in levels of aflatoxin. Soaking peanuts in water, magadi, NaOCl and ammonium persulphate significantly reduced aflatoxin levels by 27.7, 18.4, 18.3 and 1.6 per cent respectively; while boiling the peanuts in magadi, local ash, baking powder and water reduced aflatoxin levels by 43.8, 41.8, 28.9 and 11.7 per cent respectively. Using magadi during boiling increased the acceptability of the boiled peanuts while reducing the aflatoxin levels. The impact of aflatoxin levels in peanuts studied in this research is within safe limits except a few samples, and therefore aflatoxin contamination of peanuts at household level is not a serious threat. Contamination by aflatoxin and post-harvest fungi can be reduced by focusing on improved control strategies for wetter and more humid zones such as planting improved peanut cultivars and controlling pre-harvest pest damage. Conventional household peanut preparation techniques should be explored as possible aflatoxin management strategies in Kenya. The aflatoxin binding properties of locally available salts such as magadi and locally prepared ash should be further investigated.
Thesis (Ph.D.)-University of KwaZulu-Natal, Pietermaritzburg, 2010.

Text in English
Fungal species and their mycotoxins are the most predorminant contaminants of dried agricultural products in sub-Saharan Africa (SSA) and the main species of fungi that can synthesize mycotoxins are Aspergillus, Fusarium and Penicillium. In Africa, aflatoxin is labelled as a great threat to human and animal health due to its high contamination levels reported of aflatoxins in foods. The aim of this study was to survey fungi and aflatoxin contamination of small-scale processed foodstuffs sold at informal retail outlets in the Johannesburg metropolis, South Africa. A total of 270 food samples (10 starch and legume based foods, 11 meat and fish based foods, 22 spices and local condiments, 14 dried fruits and vegetables) were collected from retailers; and analysed four (4) times in different seasons of spring, summer, autumn and winter. Out of the 270 samples analysed, only 27.8% were contaminated with fungal. Of all the six categories of foods analysed, roots and tubers (60.0%), nuts and seeds (40.0%), dried vegetables (37.1%), and the Meat and Insect foods (33.3%) respectively, had the most contaminated samples with fungal respectively. The least contaminated food groups were the fish foods (10.0%) and spices and local condiments (16.7%) respectively. Twenty percent of the 270 dried food analysed were contaminated by Aspergillus species out of which 61.1% of the contaminated samples had fungal counts above 103 cfu/g. Aspergillus niger was the most predominant Aspergillus species identified in all the categories of food samples analysed. Fruits and vegetables (24.4%) and the nuts and seeds (20.0%) food groups had the highest number of samples contaminated with aflatoxin. Peanut flour and Cardamom had the most incidence of aflatoxin. AFB1, AFB2 & AFG1 were the most prominent aflatoxin types recovered from the food samples. Almost all the food samples in which aflatoxin were identified had aflatoxin values above 10μg/ml.
Life and Consumer Sciences
M.Sc. (Life Science)

M.Tech. (Biomedical Technology)
Aflatoxins (AFs) are naturally occurring secondary metabolites produced principally by Aspergillus flavus and Aspergillus parasiticus in food and feed commodities worldwide. Contaminations of compound feeds by AFs do not only affect animal health, but the economy as well. It is for this purpose that a study was carried out to establish the quality of South African feeds with respect to AF-producing fungi, establish a correlation between levels of AFs and determinant gene (nor-1) responsible for producing these toxins. To this end, compound feeds (n=92) from various feed manufacturers in South Africa were sampled and analysed for aflatoxigenic fungi (Aspergillus flavus and Aspergillus parasiticus) and nor~1 genes using the conventional identification and real time- polymerize reaction (RT-PCR) methods, respectively. Data obtained revealed that 66.5 and 53.1% of samples were positive for A. flavus and A. parasiticus, respectively. Aflatoxins levels in similar samples were estimated by high performance liquid chromatography (HPLC) following an immune-affinity clean-up and multi mycotoxin extraction procedures. Accordingly, levels established ranged from 0.06 – 77.97 ppb (mean: 16.8 ppb) with feeds for poultry being the main contaminating substrate and no correlation (overall R2=0.093) was established between the concentrations of AFs and those of nor~1. The cytotoxic effect of some selected AF extracts from these feeds on human lymphocyte cells was performed in comparison to that of AFB1 standard. Data obtained from the cytotoxic assay revealed that cell viability was affected significantly (P<0.001) by both the dose and duration of exposure, which was much more noticeable when cells were exposed to AFB1 standard than for individual extracts. In conclusion, even though none of the feeds analysed contained levels of AFs above regulatory limits established in South Africa, such feeds when consumed on a continuous basis may pose some serious health problems especially when AFs is found in co-contamination with such significant mycotoxins as ochratoxins (OTs) and fumonisins (FBs). Thus, the continuous need to limit AFs levels in feed commodities from South Africa is imperative.

碩士
亞洲大學
保健營養生技學系碩士班
98
The advantages of concentrated chinese medicinal preparations (CCM) was conveniented to consume and saved time in preparing traditional decoction. In recent years, many domestic businesses of CCM products often add raw medicine powder to manufacture excipients, but owing to the numerous problems of adding raw medicine powder, like the possible contents of more pesticide residue, heavy metals and bacterial infection. The method used commercial immunoaffinity columns for clean-up and HPLC with fluorescence detection for quantification of aflatoxins (AF). The samples were extracted with 25 mL 80% methanol, filtered and applied to an AflaTest immunoaffinity column. The column was washed with 10 mL purified water. AF was eluted with methanol and quantified by reversed-phase HPLC with fluorometric detection (excitation wavelength 365 nm, emission wavelength 440 nm) using methanol–acetonitrile–water (19:18:63, v/v) as mobile phase. The regression equations of AFB1 was Y=0.0016X+0.1364 (r=0.9999). The intraday and interday relative standard deviations of AFB1 were at the levels of 0.00-0.05% and 0.01-0.05%, respectively. Detection limit of AFB1 was 0.1 ng/mL based on a signal-to-noise ratio of 3:1. The average AFB1 recoveries from spiked AFB1-free CCM varied from 0.00-83.75%, and RSD ranged from 0.00 to 0.05%. The method was applied to 122 samples. AF was detected in 25 of samples, measurable at 1.15-14.9 ng/g. In conclusion, this study had shown that the HPLC method could be applied successfully to analyze AF occurred in the CCM. Risk assessment calculated of AFB1 for either the hepatitis B carriers or healthy population does not present any immediate risd based on the investigation.

Mycotoxins are poisonous secondary metabolites commonly produced by fungi and are involved in human disease conditions known as mycotoxicoses. There is evidence to show that food eaten by the rural Black population of Southern Africa is contaminated with mycotoxins. A tenuous relationship exists between the occurrence of mycotoxins in foods and certain disease conditions in humans. In order to verify this relationship, efforts have, in the past, been made to detect mycotoxins and their metabolites in physiological fluids and tissues. The difficulty with this approach is that mycotoxins in the body have short half lives, being rapidly excreted or metabolised to other forms. More recently it has been shown that aflatoxin B1, as its activated epoxide, can conjugate with macromolecules such as nucleic acids and proteins. These survive for much longer than the free toxins and by suitable methods can be isolated and measured. This allows for a much better estimate of exposure of the individual to aflatoxin. This study reviews and evaluates screening methods for the detection and analysis of mycotoxin contamination in rural foodstuffs such as maize and groundnuts. Methods for the production of aflatoxin-lysine and protein adducts are motivated and developed then used in the identification of naturally occurring adducts in humans. Isolation and quantitative analysis techniques are proposed to routinely screen patients for evidence of aflatoxin exposure.
Thesis (M.Med.)-University of Natal, Durban, 1995.

Mycotoxins are toxic secondary metabolites produced by filamentous fungi. Aflatoxins and fumonisins are among the most toxic mycotoxins. They are a significant risk factor for a cocktail of chronic health conditions including cancer of the liver, oesophagus and kidney, teratogenicity, neural tube defects, interference with lipid metabolism, a weakened immune system and a negative impact on micronutrient absorption in both man and animals. This study compared natural contamination of peanuts, peanut butter and sorghum from Gaborone, Botswana and Bulawayo, Zimbabwe with aflatoxins and fumonisins. In total 34 peanut samples, 34 sorghum samples and 11 peanut butter samples were collected randomly from retail shops and informal markets in the two cities. Fungal contamination was determined using standard mycology methods. Aflatoxin and fumonisin contamination was determined using HPLC-FLD. A. flavus/parasiticus species were detected in 66% and 100% of randomly analysed peanut samples from Bulawayo and Gaborone respectively and 27% (3/11) of peanut butter samples from Bulawayo. 67% of randomly analysed sorghum samples from Bulawayo showed A. flavus/parasiticus and Fusarium species contamination while none of the randomly analysed sorghum samples from Gaborone showed any fungal contamination. Furthermore aflatoxins were not detected in any of the sorghum samples; however 61% (11/18) of the Bulawayo sorghum samples showed fumonisin contamination (Range: 8 – 187 ng/g). Three of the peanut samples from Bulawayo were contaminated with aflatoxins (range: 6.6 – 622 ng/g) and no aflatoxins were detected in Gaborone peanuts. All 11 peanut butter samples from Bulawayo were contaminated with aflatoxins (Mean: 73.5 ng/g, Range: 6.8-250 ng/g) and AFB1 was the most prevalent. These preliminary results indicate that peanut butter and peanuts from Bulawayo are contaminated with high levels of aflatoxins. Stricter policing of regulations should be implemented to ensure compliance by manufacturers and public health interventions implemented in vulnerable communities.
Life & Consumer Sciences
M. Sc. (Life Sciences)