Determination of ochratoxin A in coffee by ELISA method and its relationship with the physical, physicochemical and microbiological properties / Determinación de ocratoxina A en café mediante método ELISA y su relación con las propiedades físicas, fisicoquímicas y microbiológicas

Background: Coffee is one of the most consumed beverages in the world; however, it may contain toxic compounds such as ochratoxin A (OTA). Objectives: Determine the OTA's presence in different types of coffee, intended for beverage preparation and marketed in Colombia through the application of the enzyme-linked immunosorbent assay (ELISA) and analyze its relationship with the physical, physicochemical and microbiological properties. Methods: 8 samples of coffee commercialized in the Colombian market were selected, in which the OTA content was determined by applying the ELISA method. Likewise, a microbiological analysis was performed, and physicochemical properties were determined, such as moisture content, aw, percentage total dissolved solids (%TDS), and extraction yield (%EY). Physical properties such as free-flow densities, compacted bulk densities (CBD), porosity, average particle size (ASP), and color. The data were treated with multivariate analysis using Principal Component Analysis (PCA) and Cluster Analysis (CA) to quantitatively investigate the relationships between the coffee samples concerning their physical, physicochemical properties, and OTA content. LSD test was applied with a significance level of 95 % and Pearson correlation test. Results:All the samples had OTA content, but only 2 exceeded the limits allowed by the regulations, with a maximum value of 15.449 µg/Kg, which represents 31.449 % of the tolerable daily intake according to the parameters defined by Joint FAO/WHO Expert Committee on Food Additives (JECFA). According to the PCA and CA, the samples were grouped harmonically according to the type of coffee associated with its commercial presentation and industrial process, OTA content, and ASP. OTA content was significantly and positively correlated (p< 0.05) with %EY, %TDS, ASP, porosity, CBD and moisture. Conclusions: The coffees marketed in Colombia showed a variable range of OTA, where soluble coffees had higher OTA contents than roasted coffees, and 25 % of the coffees analyzed do not meet the levels defined by Colombian regulations. The OTA content in coffee is related to properties that define the ability to extract solutes from coffee

Antecedentes: El café es una de las bebidas más consumidas en el mundo, sin embargo, puede contener compuestos tóxicos como la ocratoxina A (OTA). Objetivos: Determinar la presencia de OTA en diferentes tipos de café destinados a la preparación de bebida y comercializados en Colombia mediante la aplicación del ensayo inmunoabsorbente ligado a enzimas (ELISA) y analizar su relación con las propiedades físicas, fisicoquímicas y microbiológicas. Métodos: Se seleccionaron 8 muestras de café comercializado en el mercado colombiano, en las cuales se determinó el contenido de OTA mediante la aplicación del método ELISA. Así mismo se realizó análisis microbiológico y se determinaron propiedades fisicoquímicas como contenido de humedad, aw, porcentaje de sólidos disueltos totales (%TDS) y rendimiento de extracción (%EY); y propiedades físicas como densidad por caída libre, densidad compactada (CBD), porosidad, tamaño promedio de partícula (ASP) y color. Los datos fueron tratados con análisis multivariado empleando análisis de componentes principales (PCA) y análisis de conglomerados (CA) para investigar cuantitativamente las relaciones entre las muestras de café con respecto a sus propiedades físicas, fisicoquímicas y contenido de OTA. Se aplicó prueba LSD con un nivel de significación del 95 % y prueba de correlación de Pearson. Resultados: Todas las muestras presentaron contenido de OTA, pero solo 2 sobrepasaron los límites permitidos por la normatividad, con un valor máximo de 15.449 µg/Kg, el cual representa un 31.449 % de la ingesta diaria tolerable según los parámetros definidos por el Comité Mixto FAO/OMS de Expertos en Aditivos Alimentarios (JECFA). De acuerdo al PCA y CA, las muestras se agruparon armónicamente de acuerdo al tipo de café asociado a su presentación comercial y proceso industrial, contenido de OTA y ASP; el contenido de OTA se correlacionó significativa y positivamente (p < 0.05) con el %EY, %TDS, ASP, porosidad, CBD y humedad. Conclusión: Los cafés comercializados en Colombia presentan un rango variable de OTA, en donde los cafés solubles presentan contenidos de OTA mayores que los cafés tostados y el 25 % de los cafés analizados no cumplen con niveles definidos por la normatividad colombiana. El contenido de OTA en el café está relacionado con propiedades que definen la capacidad de extracción de solutos del café

Efecto antifúngico y antimicotoxigénico de Lactobacillus plantarum CRL 778 a diferentes valores de actividad de agua / Antifungal and antimycotoxigenic effect of Lactobacillus plantarum CRL 778 at different water activity values

Ochratoxin A (OTA) is a mycotoxin produced by filamentous fungi with high impact Lactic acid bacteria; in food safety due to its toxicity. In the last decade, the presence of OTA was widely reported in different foods. In this study, the ability of Lactobacillus (L.) plantarum CRL 778 to control growth and OTA production by Aspergillus (A.) niger 13D strain, at different water activity (a w) values (0.955, 0.964, 0.971, 0.982, and 0.995) was determined in vitro. Both parame ters were significantly (p<0.05) reduced by the lactobacilli and the effect depended on a w. Greatest growth rate inhibition (46.9%) was obtained at a w = 0.995, which is the most suitable value for growth and production of antifungal metabolites (lactic acid, acetic acid, phenyllac-tic and hydroxyl-phenyllactic acids) by L. plantarum CRL 778. Besides, morphological changes and inhibition of melanin synthesis were observed in colonies of A. niger 13D in presence of L. plantarum CRL 778 at a w ranged between 0.971 and 0.995. In addition, maximum reduction (90%) of OTA production took place at a w = 0.971, while inhibition of fungi growth was more evident at a w =0.995. These findings suggest that L. plantarum CRL 778 could be used for control of ochratoxigenic fungal growth and OTA contamination in different fermented foods with a w values between 0.971 and 0.995.

Ocratoxina A (OTA) es una micotoxina producida por hongos filamentosos con un alto impacto en la seguridad alimentaria debido a su toxicidad. En la última década se ha reportado ampliamente a nivel mundial, la presencia de OTA en diversos alimentos. En este estudio se evaluó in vitro, la capacidad de Lactobacillus (L.) plantarum CRL 778 de controlar el crecimiento y la producción de OTA por Aspergillus (A.) niger 13D, a diferentes valores de actividad de agua (a w): 0.955, 0.964, 0.971,0.982 y 0.995). La cepa láctica redujo significativamente (p <0.05) ambos parámetros, siendo el efecto dependiente del valor de a w. La mayor inhibición del crecimiento (46.9%) se obtuvo a a w =0.995, valor más adecuado para el crecimiento y producción de metabolitos antifúngicos (ácido láctico, ácido acético, ácidos fenil-láctico e hidroxi-fenil láctico) por la cepa láctica. Además, se observaron cambios morfológicos en las colonias de A. niger 13D, crecidas en presencia de L. plantarum CRL 778 a valores de a w de 0.971 y 0.995. El porcentaje máximo de reducción en la producción de OTA (90%) por la cepa láctica se observó a un valor de a w = 0.971, mientras la inhibición del crecimiento fúngico fue mayor cuando a w = 0.995. Estos hallazgos sugieren que L. plantarum CRL 778 podría emplearse para el control de la contaminación por hongos ocratoxigénicos en alimentos con valores de aw comprendidos entre 0.971-0.995.

Efeito da ocratoxina A em suco de uva tinto sobre sobrevivência e geração de EROs em Caenoaerbiditis elegans / Effects of ochratoxin A in red grape juice on survival rate and ROs generation in Caenoaerbiditis elegans

O consumo do suco de uva vem crescendo ao longo dos anos em função das suas propriedades funcionais. No entanto, este produto não está livre de contaminantes como a ocratoxina A (OTA). O presente trabalho teve por objetivo avaliar o efeito da presença de OTA em suco de uva Concord sobre geração de espécies reativas de oxigênio (EROs) e taxa de sobrevivência em Caenoarbiditis elegans. Os vermes foram expostos por 30 minutos ao suco na presença de OTA (0, 1, 2 e 4 µg∙L-1). O suco em presença de OTA não afetou a sobrevivência do C. Elegans. A adição de suco livre de OTA reduziu a sobrevivência dos nematoides, embora não tenha influenciado a geração de EROs. Contudo o suco com a concentração mais elevada de OTA reduziu a geração de EROs. Assim, são necessários maiores estudos para entender os mecanismos de toxicidade da OTA neste modelo vivo.

Ocratoxinas y su potencial nefrótico / Ocratoxins and their nephrotoxic potential

Las ocratoxinas son metabolitos fúngicos que están presentes en una gran variedad de alimentos y sus subproductos. La nefrotoxicidad es su principal efecto tóxico, relacionado a su vez con distintos síndromes clínicos como la necrosis tubular o la nefropatía de los Balcanes. La mayor parte de la información que se conoce sobre estas sustancias proviene de reportes de casos, ensayos en animales o estudios experimentales in vitro. Este documento ofrece una visión general sobre las ocratoxinas, su mecanismo tóxico, su efecto nefrotóxico; así como un panorama sobre su regulación actual en Colombia.

Ochratoxins are fungal metabolites that are present in a wide variety of foods and their byproducts. Nephrotoxicity is its main toxic effect, related in turn to different clinical syndromes such as tubular necrosis or Balkan nephropathy. The information that is known about these substances comes from case reports, animal trials or in vitro experimental studies. This document offers an overview of ochratoxins, toxic mechanism, nephrotoxic effect, and a panorama of their current regulation in Colombia.

Toxicological characteristics of Ochratoxin A and its impact on male reproduction / 中华男科学杂志

Ochratoxin A (OTA) is found not only nephrotoxic, teratogenic, neurotoxic, and immunotoxic, but also reprotoxic for human and animals. In the recent decade, more attention has been paid to the impact of OTA on human reproduction and the studies of its underlying mechanisms. Many studies show that OTA affects the function of the reproductive system by acting as an endocrine disrupter and, as a testicular toxin, decreases sperm quality and even induces testis cancer. This review summarizes the toxicological characteristics and toxicokinetic process of OTA as well as recent progress in the studies of various toxic effects of OTA and their underlying mechanisms, hoping to call the attention from more people to the toxicity of OTA to male reproductive health.

Efeitos da ocratoxina no desempenho do camarão-branco-do-pacífico (Litopenaeus vannamei, Bonne) / The effects of ochratoxina in performance pacific white shrimp (Litopenaeus vannamei Bonne)

A ocratoxina é um dos maiores grupos de micotoxinas; são metabólitos secundários produzidos principalmente por fungos dos gêneros Aspergillus e Penicillium. Possui propriedades tóxicas e nefrotóxicas, está relacionada à nefropatia endêmica dos Bálcãs, a tumores do trato urinário e foi classificada pela Agência Internacional de Pesquisa do Câncer (IARC) como pertencente ao grupo 2B, por ser possivelmente carcinogênica para humanos. O objetivo do presente estudo foi avaliar os efeitos da ocratoxina A (OTA) no desempenho do camarão-branco-do-pacífico (Litopenaues vannamei). O experimento foi feito simulando o manejo produtivo de uma fazenda de camarão marinho do litoral localizada em Luís Correia, Piauí. Foram utilizados cinco tratamentos com diferentes níveis de micotoxinas: T1- 100µg/kg de OTA; T2- 500µg/kg de OTA; T3- 1000µg/kg de OTA; T4- 100µg/kg de OTA e 500µg/kg afatoxina B1 e T5 - 0,0µg/kg de OTA. A produção de OTA foi realizada por meio da fermentação do milho, utilizando-se a cepa de Aspergillus ochraceus. Rações comerciais foram contaminadas com os núcleos de milho. A detecção e a quantificação de OTA dos núcleos, das rações comerciais e dos tecidos do camarão (cefalotórax e abdome) foram realizadas por cromatografia líquida de alta eficiência (CLAE). Para simular o sistema de criação da fazenda, os animais foram cultivados por um período de oito semanas, sendo 20 animais por caixa, recebendo alimentação duas vezes por dia. O menor ganho de peso observado foi no T2 e no T4 e os maiores ganhos de peso foram obtidos no T1 e no T5, que também apresentaram a melhor conversão alimentar. Após 56 dias de experimento, foi detectada OTA residual nas amostras de abdome apenas nos camarões do T1. Logo, camarões alimentados com rações contaminadas com OTA têm seu desempenho produtivo comprometido, o que gera impactos econômicos negativos para a indústria carcinicultora, além de ser um risco à saúde do consumidor, devido aos resíduos em sua musculatura.(AU)

Ochratoxin A is the second largest group of mycotoxins. It is a secondary metabolite produced mainly by fungi of the genera Aspergillus and Penicillium, and has toxic and nephrotoxic properties that are associated with the Balkan endemic nephropathy and urinary tract tumors. The International Agency for Research on Cancer (IARC) classifies it as group 2B: possibly carcinogenic to humans. The aim of this study was to evaluate the effects of ochratoxin A (OTA) on the performance of the Pacific white shrimp (Litopenaues vannamei). The experiment simulated the productive management of a shrimp farm located on the marine coast of Luis Correia, Piauí State. Five treatments with different levels of mycotoxins were used: T1- 100µg/kg of OTA; T2- 500µg/kg of OTA; T3 - 1000µg/kg of OTA; T4- 100µg/kg of OTA and 500µg/kg afatoxina B1 and T5 - 0.0µg/kg of OTA. OTA was produced by fermenting corn, using the Aspergillus ochraceus strain. Commercial feeds were contaminated with the corn kernels. OTA in the kernels, commercial feeds and shrimp tissues (cephalothorax and abdomen) were detected and quantified via high performance liquid chromatography (HPLC). To simulate the farming system, totaling 20 animals per box. The animals were fed twice a day and raised under these conditions for eight weeks. The shrimp gained weight during the weeks of the test, when subjected to different OTA treatments. The lowest weight gain was observed in T2 and T4 and the highest weight gains were in T1 and T5, which also presented the best feed conversion ratio. After 56 days, residual OTA was detected in samples of shrimp abdomen only in T1. Therefore, the productive performance of shrimp that are fed with OTA-contaminated feed is compromised, which has a negative economic impact on the shrimp industry, and is a health risk to consumers due to residues in the muscles.(AU)

Isolation and identification of toxigenic and non-toxigenic fungi in samples of medicinal plants from the market / Isolamento e identificação de fungos toxigênicos e não toxigênicos em amostras plantas medicinais do comércio

ABSTRACT: The consumption of preparations of medicinal plants has been increasing during the last decades in occidental societies. The presence of toxigenic fungi in a plant product may represent a potential risk of contamination, because of aflatoxins and ochratoxins. In this study, 12 samples of medicinal plants were analyzed in relation to the level of fungal contamination, and the presence of producers of ochratoxin A and aflatoxins was assessed by visualization of fungi using a cromatovisor in coconut milk. Most of the species found belong to the genus Cladosporium, Fusarium, Aspergillus and Penicillium. Species producing ochratoxin A were present in 2 samples (16.7%), Melissa and Hibiscus. Species producing aflatoxin were found in samples of Jacaranda decurrens (8.33%). This study suggests that herbs, if stored improperly, can provide the growth of fungi and should be examined before consumption.

RESUMO: O consumo das plantas medicinais vem aumentando nas últimas décadas nas sociedades ocidentais, porém, a presença de fungos toxigênicos nestas plantas pode representar um risco em potencial de contaminação devido à produção de aflatoxinas e ocratoxinas. Neste trabalho, 12 amostras de plantas medicinais foram analisadas em relação ao nível de contaminação por fungos, enquanto a presença de produtores de ocratoxina A e aflatoxinas foi avaliada pela visualização em cromatovisor dos fungos em meio de leite de coco. A maioria das espécies encontradas pertence aos gêneros Cladosporium, Fusarium, Aspergillus e Penicillium. Espécies produtoras de ocratoxina A estavam presentes em 2 amostras (16,7%), Melissa e Hibisco. Espécies produtoras de aflatoxina foram encontradas na amostra de Carobinha (8,33%). Este trabalho sugere que as ervas, sendo armazenadas inadequadamente, proporcionam o crescimento de fungos e, por isso, estes devem ser examinados antes do consumo.

Development of an aptamer/fluorescence dye PicoGreen-based method for detection of fumonisin B1 / 生物工程学报

Fumonisin B1 (FB1) is a carcinogenic mycotoxin found in commodities such as corn and corn-originated products. An aptamer-based method for detection of FB1 was developed using the fluorescent dye PicoGreen, which can recognize and bind double-stranded DNA. A peak fluorescence of PicoGreen was obtained in 15 min in the presence of FB1 aptamer, which formed a double-stranded hybridizer DNA with its complementary strand. The excitation and emission wavelengths for PicoGreen detection were 480 nm and 520 nm, respectively. The sensitivity of this aptamer/PicoGreen-based method was 0.1 μg/L. This method showed a good linearity for FB1 concentration ranging from 0.1 to 1 μg/L. The entire detection procedure for FB1 could be completed within 40 min. No cross reactions were observed with any other mycotoxins against aflatoxin B1, ochratoxin A, citrinin and zearalenone, demonstrating high specificity towards FB1 aptamer. Agreement between commercial, antibody-based enzyme-linked immunosorbent assay (ELISA) kit and aptamer method was excellent with a kappa value of 0.857. Taken together, this aptamer/PicoGreen-based method is more cost-effective, time-saving and useful than ELISA for detection of FB1.

Advancement of colloidal gold chromatographic technique in screening of ochratoxin A / 中国中药杂志

Ochratoxin A (OTA) is a toxic secondary metabolite mainly produced by Aspergillus and Penicillium species, existing in a variety of foodstuffs and Chinese medicines. OTA is difficult to be detected in practice because of the characteristics such as trace amounts, toxicity, existing in complex matrices. In the numerous detection technologies, colloidal gold chromatographic techniques are highly sensitive, specific, cost-effective and user-friendly, and are being used increasingly for OTA screening. Recently, with the development of aptamer technology and its application in chromatographic technique, a newly colloidal gold aptamer chromatographic technique has been developed. This review elaborates the structures and principles of both traditional and newly colloidal gold chromatographic techniques, focuses on newly colloidal gold aptamer chromatographic technique, summarizes and compares their use in rapid detection of OTA. Finally, in order to provide a reference for better research of related work, the development trends of this novel technique are prospected.

Efeitos in vitro de ocratoxina A, deoxinivalenol e zearalenona sobre a viabilidade celular e atividade de E-ADA em linfócitos de frangos de corte / In vitro effects of ochratoxin A, deoxynivalenol and zearalenone on cell viability and E-ADA activity in broiler chickens lymphocytes

Mycotoxins are a group of chemically diverse naturally occurring substances resulting from the secondary metabolism of pathogenic filamentous fungi. They are produced mainly by the genera Fusarium, Alternaria, Aspergillus and Penicillium which can contaminate grains and cereals such as wheat, corn and soy. According to the nature and the concentration levels, mycotoxins can induce toxic effects in food-production animals and humans. An in vitro study was conducted to evaluate the susceptibility of broiler chickens lymphocytes to different concentrations of ochratoxin A, deoxynivalenol and zearalenone. Each toxin was added to the cell medium at different concentrations (0.001, 0.01, 0.1 and 1μg/mL). Cell viability and ecto-adenosine deaminase activity were assessed at 24, 48 and 72 hours by colorimetric assays. Thus, it were used 0.7x10(5) lymphocytes/mL in RPMI 1640 medium, supplemented with 10% fetal bovine serum and 2.5 IU of penicillin/streptomycin per mL, incubated at 37°C in a 5% CO2 atmosphere. All the experiments were carried out in triplicate and the results were expressed as mean ± standard error of the mean. The results showed that OTA and DON induced lymphocyte proliferation and reduced enzymatic activity in vitro (P<0,05), whereas ZEA also promoted proliferation (P<0,05), but neither alteration on enzymatic activity (P>0,05). It was possible to correlate the results about viability cell and ecto-adenosine deaminase activity, suggesting that, at minimal concentrations, the evaluated mycotoxins do not stimulated the enzymatic activity, which has proinflammatory action and contributes for the immunosuppression process, thus, avoiding a decrease on the viability cell. This is the first in vitro study conducted with OTA, DON and ZON in broiler chickens lymphocytes evaluating these parameters.

Micotoxinas representam um vasto grupo de contaminantes químicos naturais originados a partir do metabolismo secundário de fungos filamentosos patogênicos. Elas são produzidas, principalmente, pelos gêneros Fusarium, Alternaria, Aspergillus e Penicillium, os quais podem contaminar grãos e cereais, como trigo, milho e soja. Conforme sua natureza e níveis de concentração, micotoxinas podem induzir efeitos tóxicos em animais de produção e humanos. Um estudo in vitro foi realizado para avaliar a susceptibilidade das células linfocitárias de frangos de corte a diferentes concentrações de ocratoxina A, deoxinivalenol e zearalenona. Cada micotoxina foi adicionada ao meio celular em diferentes concentrações (0,001; 0,01; 0,1 e 1μg/mL). A viabilidade celular e atividade de ecto-adenosina desaminase foram analisadas em 24, 48 e 72 horas através de ensaios colorimétricos. Para isso, foram utilizados 0,7x10(5) linfócitos/mL em meio RPMI 1640, suplementado com 10% de soro fetal bovino e 2,5 UI de penicilina/estreptomicina por mL, incubados em atmosfera de 5% de CO2 a 37 °C. Todos os experimentos foram realizados em triplicata e os resultados foram expressos como média e erro padrão da média. Os resultados obtidos demonstraram que tanto ocratoxina A como deoxinivalenol induziram proliferação linfocitária e baixa atividade enzimática in vitro (P<0,05), enquanto zearalenona também induziu proliferação (P<0,05), mas nenhuma alteração na atividade enzimática (P>0,05). Foi possível correlacionar os dados referentes à viabilidade celular e atividade de ecto-adenosina desaminase, sugerindo que, em concentrações mínimas, as micotoxinas testadas não estimularam a atividade da enzima, que possui ação pró-inflamatória e contribui para o processo de imunossupressão e, portanto, evitando um decréscimo na viabilidade celular. Este é o primeiro estudo feito com OCRA, DON e ZEA sobre linfócitos de frangos de corte em cultivos in vitro na avaliação desses parâmetros.

Niveles de Ocratoxina A y Aflatoxinas totales en cafés de exportación de Panamá por un método de ELISA

Se realizó un estudio de las condiciones del procesamiento del café de exportación en 15 beneficios, ubicados en Chiriquí, región occidental de Panamá. Además se analizaron 21 muestras de café procesado (grano verde), provenientes de los beneficios. Las muestras fueron analizadas microbiológicamente y se cuantificaron las Aflatoxinas totales (B1, B2, G1 y G2) y Ocratoxina A (OTA), mediante el método de inmunoafinidad ELISA. Se determinó un límite de detección de 0,017 ng/mL, para la Ocratoxina A, lo que equivale a una concentración de 0,829 μg/kg en la muestra, y un límite de detección de 0,027 ng/mL, para las Aflatoxinas totales, lo que equivale a una concentración de 1,350 μg/kg de Aflatoxinas totales. En la muestra, se encontró que cuatro de las 21 (19%) resultaron positivas a la presencia de Ocratoxina A y tres, a la presencia de Aflatoxinas totales (14%). Las muestras presentaron niveles de Ocratoxina A en el rango de 4,90-37,73 μg/kg; sólo tres de ellas superaron el límite máximo permitido por la Unión Europea, para la concentración de Ocratoxina, que es de 5,0 μg/kg. Las Aflatoxinas totales se encontraron en el rango de 1,51- 1,93 μg/kg, por debajo de los 10 μg/kg, que es el límite máximo permitido en el café por la Unión Europea. Los resultados nos indican que el procesamiento de café producido en Panamá cumple satisfactoriamente con los estándares internacionales de manejo poscosecha, lo que conduce a una baja incidencia de hongos productores de micotoxinas y niveles muy bajos de micotoxinas.

Levels of Ochratoxin A and total Aflatoxins in Panamanian exportation coffee by an ELISA Method. A study about processing conditions of exportation coffee in 15 benefits located in Chiriquí, western region of Panama, was conducted. In addition, 21 samples of processed coffee (green beans), from the benefits, were analyzed. The samples were microbiologically tested in order to quantify total aflatoxins (B1, B2, G1 and G2) and Ochratoxin A (OTA), using the immunoaffinity ELISA method. A detection limit of 0.017 ng/mL, was determined for Ochratoxin A, which is equivalent to a concentration of 0.829 μg/kg, and a detection limit of 0.027 ng/mL, for total aflatoxins, which is equivalent to a concentration of 1.350 μg/kg. It was found that four (19%) out of the 21 samples were positive to the presence of Ochratoxin A and three (14%) to the presence of total aflatoxins. Samples showed levels of Ochratoxin A in the range 4.90 - 37.73 μg/kg; only three of them exceeded the maximum limit allowed by the European Union, for the concentration of Ochratoxin, which is of 5.0 μg/kg. Total aflatoxins were found in the range 1.51 - 1.93 μg/kg, below 10μg/kg which is the maximum limit allowed for coffee by the European Union. The results indicate that the processing of coffee produced in Panama successfully meets international standards for postharvest handling, which leads to a low incidence of mycotoxins and very low levels of mycotoxin- producing fungi.

Determination of ochratoxin A in human urine by HPLC-FLD after cleaned-up by molecularly imprinted polymer solid phase extraction column / 药学学报

A method was developed for the determination of ochratoxin A (OTA) in human urine by HPLC-FLD after molecularly imprinted polymer solid phase extraction (MIP-SPE) column. After the pH being adjusted to 2.5 with 0.1 mol x L(-1) HC1, sample was cleaned up with MIP-SPE column for ochratoxin A, the analyte was analyzed by high performance liquid chromatography coupled with fluorescence detection (HPLC-FLD), and finally all the positive results were confirmed by LC-MS/MS. Recoveries from urine samples spiked with OTA at levels ranging from 2 to 20 ng x mL(-1) were 90.6%-101.9%, and RSDs were 0.1%-1.6%. Sixty-five volunteers living in Beijing took part in the study, of which 5 were found containing OTA in their urine and the highest value was 0.091 ng x mL(-1). The MIP-SPE column was firstly applied to purify and concentrate OTA in human urine, this method is simple, rapid and reliable and can be used to determine the contents of OTA in human urine.

Aislamiento de cepas de Aspergillus sección Nigri en la yerba mate comercializada en Posadas (Misiones, Argentina) y evaluación de su potencial ocratoxigénico / [Isolation of Aspergillus section Nigri strains in yerba mate in Posadas (Misiones, Argentina) and evaluation of their ochratoxigenic potential].

The objectives of the present work were to investigate the isolation frequency of genus Aspergillus in canchada yerba mate (YMCH) and elaborated yerba mate (YME) (Ilex paraguariensis) and the proportion of section Nigri isolates, as well as to determine ochratoxin A production by Aspergillus species section Nigri. Three hundred twenty eight Aspergillus strains from 20 samples of YMCH and 1306 Aspergillus strains from 36 samples of YME were isolated; of the total, 279 from the first group of strains and 1215 from the latter group, belonged to section Nigri. For the detection of ochratoxin A production, the strains were cultivated on Czapeck yeast extract agar and the toxin was detected by thin layer chromatography under UV light. Uniserate species predominance was observed in the 1494 strains of Aspergillus section Nigri obtained (Aspergillus japonicus var. japonicus and Aspergillus japonicus var. aculeatus), whereas none of the strains analysed showed ochratoxin A production in vitro at the detection level of the methodology employed.

Pesquisa de fungos produtores de ocratoxina A em granola comercializada / Ochratoxin A-producing fungi in commercial granola

Este trabalho efetuou a identificação das espécies fúngicas presentes na granola e analisou a capacidade das cepas produzirem ocratoxina A. As amostras foram adquiridas no comércio do município de Teresina-Piauí, no total de 60 amostras de quatro diferentes marcas. Foram realizadas as metodologias de contagem,isolamento e identificação das espécies fúngicas; e as cepas da seção Nigri foram testadas quanto à capacidade de produção de ocratoxina A. Em 11 das amostras analisadas não houve o crescimento fúngico,e nas amostras em que houve os valores chegaram a 5,17 log10 UFC/g. Houve diferença significativa (p <0,05) entre as diferentes marcas de granola analisadas. Os gêneros fúngicos mais frequentemente isolados foram Cladosporium (46,9 %), seguido de Aspergillus spp. e seus teleomorfos (37,4 %), e do gênero Penicillium spp. (5,4 %). A amostra de granola da marca A apresentou contagens bem mais elevadas do que as demais, o que indica que possivelmente tenha ocorrido falhas em alguma(s) etapa(s) do processo de industrialização. Todas as cepas isoladas de Aspergillus seção Nigri não apresentaram capacidade de produção de ocratoxina A.

The present study aimed at identifying the fungal species occurring in granola, and to verify the abilityof the strains in producing ochratoxin A. Sixty granola samples of four different brands were purchasedin commercial establishments located in the city of Teresina – Piauí state. The fungi were counted,isolated and the species were identified, and the section Nigri strains were tested for detecting their abilityin producing ochratoxin A. No fungal growth was found in 11 of the analyzed samples. In samplesshowing fungal growth, it was as high as 5.17 log10 CFU/g. A significant difference (p<0.05) amongthe analyzed granola brands was found. The most frequently isolated fungus genus was Cladosporium(46.9 %), followed by Aspergillus spp and its teleomorphs (37.4 %), and Penicillium spp. (5.4 %). Thegranola brand A showed the highest counting among the analyzed brands, which demonstrated that apossible failure at some stages of the industrialization process might be occurred. None of the isolatedAspergillus section Nigri strains showed the ability in producing ochratoxin A.

Ochratoxigenic fungi associated with green coffee beans (Coffea arabica L.) in conventional and organic cultivation in Brazil

The genera Aspergillus comprises species that produce mycotoxins such as aflatoxins, ochratoxins and patulin. These are cosmopolitan species, natural contaminants of agricultural products. In coffee grains, the most important Aspergillus species in terms of the risk of presenting mycotoxins belong to the genera Aspergillus Section Circumdati and Section Nigri. The purpose of this study was to assess the occurrence of isolated ochratoxigenic fungi of coffee grains from organic and conventional cultivation from the South of Minas Gerais, Brazil, as well as to evaluate which farming system presents higher contamination risk by ochratoxin A (OTA) produced by fungi. Thirty samples of coffee grains (Coffea arabica L.) were analysed, being 20 of them of conventional coffee grains and 10 of them organic. The microbiological analysis was done with the Direct Plating Technique in a Dichloran Rose Bengal Chloramphenicol Agar (DRBC) media. The identification was done based on the macro and micro morphological characteristics and on the toxigenic potential with the Plug Agar technique. From the 30 samples analysed, 480 filamentous fungi of the genera Aspergillus of the Circumdati and Nigri Sections were isolated. The ochratoxigenic species identified were: Aspergillus auricoumus, A. ochraceus, A. ostianus, A. niger and A. niger Aggregate. The most frequent species which produces ochratoxin A among the isolated ones was A. ochraceus, corresponding to 89.55%. There was no significant difference regarding the presence of ochratoxigenic A. ochreceus between the conventional and organic cultivation systems, which suggests that the contamination risk is similar for both cultivation systems.

Diseño de un programa de Análisis de Peligros y Puntos de Control Crítico en el proceso productivo de cacao en polvo en una industria alimentaria

El Análisis de Peligros y Puntos de Control Crítico (HACCP) es una herramienta para la Gestión de Inocuidad de los alimentos que permite identificar los peligros físicos, químicos y biológicos asociados al proceso a través de toda la cadena productiva. Este trabajo tiene por finalidad diseñar el Programa de HACCP para el proceso de producción de cacao en polvo en una industria de alimentos venezolana. Previamente se evaluó el cumplimiento de las Buenas Prácticas de Manufactura (BPM) y los Procedimientos Operativos Estándar de Saneamiento (POES), elementos básicos para el establecimiento del HACCP. Se visitaron las instalaciones de varios proveedores a objeto de observar el cumplimiento de las Buenas Prácticas Agrícolas (BPA). Para el desarrollo del programa HACCP se aplicaron los siete principios básicos del mismo y las cinco tareas preliminares, conforme a la metodología descrita por el Codex Alimentarius.Conducido el análisis de peligros, se identificaron tres puntos de control críticos en la línea de proceso: descascarillado (control de ocratoxina A), fase de tostado (control de Salmonella) y detección de partículas metálicas. Se establecieron los Límites Críticos, los Procedimientos de Vigilancia, las Acciones Correctivas, los Procedimientos de Verificación y de Documentación, recomendándose implementar el Programa HACCP en la industria procesadora de cacao en polvo con la realización de los ajustes correspondientes en los casos donde sea necesario. Recientemente la ocratoxina A (OTA),ha sido relacionada con el cacao en grano. Aunque se ha señalado que el descascarillado es una medida de control efectiva para este peligro químico, se recomienda estudiar la prevalencia de OTA en el cacao producido en el país y validar la etapa del descascarillado como control de micotoxinas.

Design of an HACCP program for a cocoa processing facility. The HACCP plan is a food safety management tool used to control physical, chemical and biological hazards associated to food processing through all the processing chain. The aim of this work is to design a HACCP Plan for a Venezuelan cocoa processing facility.The production of safe food products requires that the HACCP system be built upon a solid foundation of prerequisite programs such as Good Manufacturing Practices (GMP) and Sanitation Standard Operating Procedures (SSOP). The existence and effectiveness of these prerequisite programs were previously assessed.Good Agriculture Practices (GAP) audit to cocoa nibs suppliers were performed. To develop the HACCP plan, the five preliminary tasks and the seven HACCP principles were accomplished according to Codex Alimentarius procedures. Three Critical Control Points (CCP) were identified using a decision tree: winnowing (control of ochratoxin A), roasting (Salmonella control) and metallic particles detection. For each CCP, Critical limits were established,the Monitoring procedures, Corrective actions, Procedures for Verification and Documentation concerning all procedures and records appropriate to these principles and their application was established. To implement and maintain a HACCP plan for this processing plant is suggested. Recently OchratoxinA (OTA) has been related to cocoa beans. Although the shell separation from the nib has been reported as an effective measure to control this chemical hazard, ochratoxin prevalence study in cocoa beans produced in the country is recommended, and validate the winnowing step as well.

Determination of multiple constituents in Wuling capsules by HPLC simultaneously / 中国中药杂志

<p><b>OBJECTIVE</b>To establish a method for the content determination of 5-methylmellein, 5-hydroxymellein, 5-carboxylmellein and genistein in Wuling capsules simultaneously by HPLC.</p><p><b>METHOD</b>Four components were determined by HPLC on a Kromasil KR100-5C18 column (4.6 mm x 250 mm, 5 microm) with acetonitrile and 0. 2% phosphoric acid as mobile phase in a gradient elution. The flow rate was 1.0 mL x m min(-1), and the column temperature was set at 30 degrees C.</p><p><b>RESULT</b>5-hydroxymellein showed a good linear relationship at the range of 7.86-157.2 ng, the average recovery was 101.0% with RSD 1.3%. 5-carboxylmellein showed a good linear relationship at the range of 9.57-191.4 ng, the average recovery was 98.6% with RSD 1.5%. Genistein showed a good linear relationship at the range of 28.80-576.0 ng, the average recovery was 98.6% with RSD 1.8%. 5-methylmellein showed a good linear relationship at the range of 21.46-429.2 ng, the average recovery was 99.2% with RSD 1.8%.</p><p><b>CONCLUSION</b>The established method is feasible and the repeatability is good. The method can be used for quality control of Wuling capsules.</p>

Simultaneous determination of aflatoxin B1, B2, G1, G2 and ochratoxin A in Glycyrrhiza uralensis by HPLC-fLD after immunoaffinity column with online post-column photochemical derivatization / 中国中药杂志

<p><b>OBJECTIVE</b>To develop a method for the simultaneous determination of aflatoxin B1, B2, G1, G2 and ochratoxin A in Glycyrrhiza uralensis by HPLC-FLD after immunoaffinity column with online post-column photochemical derivatization.</p><p><b>METHOD</b>Sample was extracted with MeOH: H2O (80:20) and cleaned up by immunoaffinity column. The toxins were separated by reversed-phase HPLC and the mobile phase was consisted of methanol and 0.5% acetic acid solution with gradient elution. The determination was carried out by fluorescence detector after photochemical derivatization.</p><p><b>RESULT</b>The detection limits of aflatoxin G2, G1, B2, B1 and ochratoxin A were 0.02, 0.06, 0.015, 0.03 and 0.25 microg x kg(-1), respectively. The recoveries of analytes were from 76.0% to 103% and the relative standard deviations (RSDs) were below 13%.</p><p><b>CONCLUSION</b>The method is a simple, accurate and can be used to determine the contents of aflatoxin B1, B2, G1, G2 and ochratoxin A in G. uralensis simultaneously.</p>

Determinación de ocratoxina-a por HPLC con detección por fluorescencia (HPLC-FL): un nuevo método estandarizado para muestras de trigo / Ochratoxin-a determination by HPLC with fluorescence detection(HPLC-FL): a new standardization method for wheat samples

El objetivo principal de este estudio fue evaluar la presencia de Ocratoxina-A (OTA) en los granos del trigo y harina del trigo realizadas por un nuevo método de determinación que usa la cromatografía líquida de alta resolución (CLAR) acoplada al descubridor delfluorimetrio. El experimento usó seis muestras de grano de trigo del lugar del almacenamiento diferente a la industria local de Chapeco (SC), Brasil Sur, en agosto, 2008. El extracto de OTA era llevado a cabo usando el acetonitrila: agua (120:80 vlv) como solventes. Después el suprenadante fue filtrado, y aplicado en la columna del inmunoafinidad específica a OTA. Además, la columna se lavó con agua y la toxina era el eluido con el metanol. La determinación del OTA se realizó por detección de fluorescencia acoplado al aparato de HPLC. Los volúmenes de OTA en los granos del trigo y harina del trigo eran entonces los determínate y los resultados mostraron una concentración de OTA menor que los límites exigidos por la legislación internacional.

The main objective of this study was to evaluate the presence of Ochratoxin A (OTA) in wheat grains and wheat flour samples using a new high performance liquid chromatography (HPLC) method. The experiment used six wheat grain samples from different industry storage place from Chapeco (SC), South Brazil, on August 2008. The OTA extraction was carried out using acetonitrile: water (120:80 v/v) as solvent. Thereafter, the supernatant was filtered, and applied on OTA-specific immunoafinity column to HPLC Furthermore, the column was washed with water and the toxin was eluted with methanol. The OTA wheat grains and wheat flour concentration were analyzed by a fluorescence detector coupled to the HPLC apparatus. The results showed a smaller OTA concentration than the limits set by international legislation.