Reduction of in-shell Brazil nut (Bertholletia excelsa H.B.K.) aflatoxin contamination by ozone gas application during storage

The susceptibility of the in-shell Brazil nut mycoflora and aflatoxins (AFLs) contamination to ozone (O3) gas during storage is reported. In-shell Brazil nuts obtained from retail market were submitted to O3 gas atmosphere at different concentrations immediately before to be stored. Samples were collected just after the gas exposure and every 30 days during the storage period to carry on mycological tests and AFLs analysis. A sensorial evaluation by descriptive quality analysis was carried out to check treated nuts sensory attributes according to consumer acceptance after gas exposure. The O3 treatment applied within 5 h at 31 mg/L was able to successfully inhibit the viability of fungi of the nut-contaminating microflora and so the toxigenic Aspergillus species from the day of application. AFLs were totally degraded in all samples whatever O3 concentration applied. No significant changes on sensory attributes were observed that could affect nut acceptability  after the O3 treatments and storage conditions applied in the present experiment. This procedure is tentatively applied at an Amazon State nut factory for checking its potential in mycotoxin risk contamination of in-shell Brazil nuts safeguarding under the Amazon region environment. Keywords: In-shell Brazil nut, Ozone, Mycoflora, Aflatoxin, Storage, Sensory evaluation

Ochratoxin A and filamentous fungi in red wine grapes from Santa Catarina, Brazil

The quality of wines has been evaluated traditionally according to sensorial properties. Recently, safety issues have been raised, such as pesticide residues and mycotoxins, with the introduction of new agricultural practices and the development of analytical methods with higher sensitivity. Ochratoxin A (OTA) is such a mycotoxin, produced by some Aspergillus and Penicillium species and is one of the most recent safety issues for wine. The mycobiota of, and the occurrence of OTA in Southern Brazilian grapes are not known. The presence of these contaminants was assessed by collecting 30 samples of grapes, from 16 vineyards, from the two most important wine subregions in the State of Santa Catarina, Brazil. The mycobiota was evaluated by plating 10 grapes from each sample in Dichloran Rose Bengal Chloramphenicol Agar and Sabouraud Dextrose Agar, supplemented with chloramphenicol. Production of OTA by black Aspergillus strains was estimated after growing in Czapeck Yeast Agar. OTA was analysed in 9 grape samples by chromatography with immunoaffinity clean-up, as stipulated by the European regulation. Three hundred and eighty seven strains were isolated. The dominant genera were Cladosporium (found in 86.7% of plated berries), Alternaria (80.0%), Botrytis (70.0%), Aspergillus (66.7%), and Penicillium (63.3%). Sixteen A. niger aggregate strains (26 % of total Aspergillus strains) were isolated, and OTA was not detected from any of these strains. No A. carbonarius was isolated. OTA was found in 6 grape samples, with a range of values from 0.16 μg/Kg to 0.77 μg/Kg. In conclusion, no OTA producing black Aspergillus strains were found in grapes, although some grape samples contain the mycotoxin. The fungal source of OTA requires further investigation

Effect of oxygen reducing atmospheres on the quality and safety of stored shelled Brazil nut packs

High moisture content, relative humidity, temperature and environment rich in oxygen (O2) are the main factors for tree nuts to get infected by fungi and so aflatoxins (AFLs) contaminated. During storage and commercialization dry Brazil nuts packs need to maintain their safety and quality. Modified atmospheres in storage (macro-environment) and packaging (micro-environment) have been used to prolong food shelf life by reducing O2 concentration with inhibitory gases or, more recently, by adding O2 absorber pads. This work reports the application of O2 atmosphere reducing methods on stored shelled Brazil nut packs aiming fungi and AFL degradation as well as hygienic conditions improvements. The methods applied were: (a) ozone - O3, (b) carbon dioxide - CO2 and (c) O2 absorber pads with and without vacuum. Nuts were submitted to microbiological tests (fungi, aflatoxigenic strains, yeast and bacteria), moisture content and AFLs analysis. From all O2 reducing atmosphere evaluated, the best performance was obtained with O3. A reduction on fungi growth (1.8 x 104 cfu.g-1 to 2.6 x 10 cfu.g-1) and yeast destruction after the first month of storage were registered. Also O3 was the only nut treatment that was able to degrade AFLs. None of the spiked (AFLs: 15 ppb) nut samples O3 treated had AFLs detected up to the LOQ of the method (0.36 μg.kg-1 for AFB1+AFB2+AFG1+AFG2) i.e., much lower than the allowed by the European Union regulation (MRL: 4 and 2 ppb for total and AFB1, respectively), thus producing safer nuts. All other treatments stabilized and/or inhibited microorganisms growth. Add CO2 and O2 pads played an important role on nut quality. Further study will be carried out in order to adjust O3 concentration and application conditions for longer period of storage

Comparison of methods by TLC and HPTLC for determination of aflatoxin M1 in milk and B1 in eggs Comparação de metodologia para análise de aflatoxina M1 em leite e aflatoxina B1 em ovos por CCD e CCDAE

Milk and egg matrixes were assayed for aflatoxin M1 (AFM1) and B1 (AFB1) respectively, by AOAC official and modified methods with detection and quantification by thin layer chromatography (TLC) and high performance thin layer chromatography (HPTLC). The modified methods: Blanc followed by Romer, showed to be most appropriate for AFM1 analysis in milk. Both methods reduced emulsion formation, produced cleaner extracts, no streaking spots, precision and accuracy improved, especially when quantification was performed by HPTLC. The use of ternary mixture in the Blanc Method was advantageous as the solvent could extract AFM1 directly from the first stage (extraction), leaving other compounds in the binary mixture layer, avoiding emulsion formation, thus reducing toxin loss. The relative standard deviation (RSD%) values were low, 16 and 7% when TLC and HPTLC were used, with a mean recovery of 94 and 97%, respectively. As far as egg matrix and final extract are concerned, both methods evaluated for AFB1 need further studies. Although that matrix leads to emulsion with consequent loss of toxin, the Romer modified presented a reasonable clean extract (mean recovery of 92 and 96% for TLC and HPTLC, respectively). Most of the methods studied did not performed as expected mainly due to the matrixes high content of triglicerides (rich on saturated fatty acids), cholesterol, carotene and proteins. Although nowadays most methodology for AFM1 is based on HPLC, TLC determination (Blanc and Romer modified) for AFM1 and AFB1 is particularly recommended to those, inexperienced in food and feed mycotoxins analysis and especially who cannot afford to purchase sophisticated (HPLC,HPTLC) instrumentation.Aflatoxinas M1 (AFM1) e B1 (AFB1) foram analisadas em leite e ovos respectivamente, por diferentes métodos oficiais da AOAC e modificações usando detecção por cromatografia em camada delgada (CCD) e CCD de alta eficiência (CCDAE). Os métodos modificados: Blanc e Romer, apresentaram-se mais apropriados para análise de AFM1 em leite. Ambos reduziram formação de emulsão, produziram extratos limpos, sem formação de caudas. Inclusive, a precisão e acuidade aumentaram, especialmente quando a quantificação foi realizada por CCDAE. O uso de solventes ternários, no método de Blanc, foi vantajoso. Este solvente extrai AFM1 diretamente da primeira fase (extração), deixando outros compostos na camada binária, evitando emulsão, reduzindo assim, perda da toxina. O RSD% foi muito baixo com 16 e 7%, respectivamente. Quanto ao ovo, AFB1 e extrato final, ambos os métodos necessitam mais estudos. Embora esta matriz induza a formação de emulsão com conseqüente perda de toxina, o método de Romer modificado apresentou extrato razoavelmente limpo com recuperação de 92 e 96% para CCD e CCDAE, respectivamente. A maioria dos métodos estudados não apresentou o desempenho esperado porque as amostras possuem conteúdo elevado de trigliceridios, colesterol, caroteno e proteínas. Embora, a maioria das metodologias para AFM1 seja baseada em CLAE; o uso de CCD para de determinação de AFM1 e AFB1 é particularmente recomendada para aqueles, inexperientes em análises de alimentos e micotoxinas e especialmente, laboratórios que não podem adquirir equipamento sofisticado

Mycoflora and deoxynivalenol in whole wheat grains (Triticum aestivum L.) from Southern Brazil.

Fungi species Fusarium graminearum is related to deoxynivalenol (DON) formation. The aim of this study was to evaluate mycoflora and DON occurrence in 53 whole wheat grains samples collected in Southern Brazil during the 2012 crop. Wheat grains showed adequate values of water activity ranging from 0.48-0.72, within the required limits of moisture content, ranging from 9.1-13.9%. In addition low counts for fungal colonies, ranging from 10 to 8.2x102 were found. For Fusarium genera there was predominance of F. verticillioides (34%) and F. graminearum (30.2%). For Aspergillus species, 37.7% A. flavus was determined. Regarding the Penicillium species, P. digitatum (49%) was the most found species. DON was detected in 47.2% (25 out of 53) of the samples analyzed, with levels from 243.7 to 2281.3 ?g kg-1 (mean: 641.9 ?g kg-1)