Occurrence of ochratoxin A and citrinin in Czech cereals and comparison of two HPLC methods for ochratoxin A detection

The aims of the study were to obtain information about the occurrence of ochratoxin A (OTA) and citrinin (CIT) in cereals harvested in the Czech Republic and to compare two analytical procedures for detecting OTA. A total of 34 cereal samples, including two matrix reference materials (R-Biopharm, Germany), were analysed. The results were compared with the limit for raw cereal grains used as a foodstuff according to Commission Regulation No. 1881/2006, which allows a maximum OTA level of 5 µg kg^?1. Compared were two methods based on the high-performance liquid chromatography principle, one using the immunoaffinity columns OchraTest? (VICAM) and the second based on solvent partition (PART), both followed by fluorescence detection. The highest OTA contents were found in two barley samples. According to the method employed, the results for the first sample (malting barley) were VICAM = 31.43 µg kg^?1 and PART = 44.74 µg kg^?1. For the second sample (feeding barley) they were VICAM = 48.63 µg kg^?1 and PART = 34.40 µg kg^?1. Two samples of bread wheat had an OTA content approaching the legal limit (VICAM = 4.71 µg kg^?1 and PART = 6.03 µg kg^?1; VICAM = 4.12 µg kg^?1 and PART = 3.95 µg kg^?1). CIT was analysed using the PART method only, and its highest content (93.64 µg kg^?1) was found for the malting barley sample with high OTA content (44.74 µg kg^?1 as analysed using PART).     

两种同时检测茶叶中赭曲霉毒素A与桔霉素方法的比对

目的 对比两种同时检测茶叶中赭曲霉毒素A(ochratoxin A, OTA)与桔霉素(citrinin, CIT)的方法。方法 考察不同提取方法、溶剂、料液比的提取效果,比较适配体结合超滤法与免疫亲和柱纯化效果;对比高效液相色谱-荧光检测器(high performance liquid chromatography-fluorescence detector, HPLC-FLD)与超高效液相色谱-三重四极杆质谱法(ultra performance liquid chromatography-tandem mass spectrometry, UPLC-MS/MS)测定OTA与CIT的检出限、精密度、准确度等差异,评价方法的适用性。结果 采用70%甲醇超声与振荡的提取方法,在料液比为1:3的条件下,OTA、CIT的提取率最佳,回收率分别为99.49%的92.77%;经适配体结合超滤纯化后,CIT与OTA的回收率为82.07%～92.95%,与免疫亲和柱纯化方法效果类似; UPLC-MS/MS测定CIT和OTA的基质效应(20.0%、-20.0%)小于HPLC-FLD测定CIT和OT...     

化学发光免疫分析方法检测粮食谷物中赭曲霉毒素A残留

目的建立粮食谷物中赭曲霉毒素A(ochratoxin A,OTA)残留检测的化学发光免疫分析方法。方法人工改造OTA半抗原、制备包被原和多克隆抗体工作液,优化检测步骤,验证各项评价指标,比较化学发光免疫分析方法和高效液相色谱法(high performance liquid chromatography,HPLC)的实际样本检测结果的一致性,以此来实现化学发光免疫分析方法的建立。结果 50%抑制浓度(IC_(50))为0.278μg/L;OTA的最低检出限为5.0μg/kg,样本添加回收率在65.4%到115.8%之间,批内、批间相对标准偏差15%。同时,通过对实际样本检测,证明化学发光免疫分析方法具有显著的准确度和可靠性。结论基于化学发光免疫分析方法能够实现粮食作物中的OTA残留的快速检测,灵敏度高、成本低、时间短,为食品中霉菌毒素残留的快速检测提供了新的技术参考。     

Co-occurrence of aflatoxins, ochratoxin A and zearalenone in barley from a northern region of Spain

One-hundred and twenty-three barley samples from a region of Spain (Navarra) were analysed in order to evaluate the possible co-occurrence of aflatoxins (AFB1, AFG1, AFB2 and AFG2), ochratoxin A (OTA) and zearalenone (ZEA). The results indicated that 80% of the samples presented detectable, although very low levels, of two or more mycotoxins. The most frequent combinations were AFB1 and OTA; AFB1, ZEA and OTA; and AFB1 and ZEA. In general, the statistical study did not show significant differences between levels or incidence for the mycotoxins in different years of harvest, variety of barley, farming or origin. The calculated values for daily intake were low and the risk to consumers could be assumed to be very low. However, the co-occurrence of several mycotoxins, and therefore synergic or additive effects, should be taken into account when determining permitted levels or risk assessment.     

Mycotoxins in breakfast cereals from the Canadian retail market: A 3-year survey

One hundred and fifty-six samples of breakfast cereals were collected from the Canadian retail marketplace over a 3-year period. The samples were analysed for the mycotoxins deoxynivalenol, nivalenol, HT-2 toxin, zearalenone, ochratoxin A, and fumonisins B₁ and B₂ to contribute to dietary exposure estimates in support of the development of Canadian guidelines for selected mycotoxins in foods. The samples included corn-, oat-, wheat- and rice-based cereals, as well as mixed-grain cereals, and were primarily from North American processors. Overall, deoxynivalenol was the most frequently detected mycotoxin —?it was detected in over 40% of all samples analysed. Fumonisins and ochratoxin A were each detected in over 30% of all samples. Zearalenone was detected in over 20% of all samples. Nivalenol and HT-2 toxin were each detected in only one sample. The survey clearly demonstrated regular occurrence of low levels of multiple mycotoxins in breakfast cereals on the Canadian market.     

Mycotoxins in breakfast cereals from the Canadian retail market: A 3-year survey

One hundred and fifty-six samples of breakfast cereals were collected from the Canadian retail marketplace over a 3-year period. The samples were analysed for the mycotoxins deoxynivalenol, nivalenol, HT-2 toxin, zearalenone, ochratoxin A, and fumonisins B₁ and B₂ to contribute to dietary exposure estimates in support of the development of Canadian guidelines for selected mycotoxins in foods. The samples included corn-, oat-, wheat- and rice-based cereals, as well as mixed-grain cereals, and were primarily from North American processors. Overall, deoxynivalenol was the most frequently detected mycotoxin  — it was detected in over 40% of all samples analysed. Fumonisins and ochratoxin A were each detected in over 30% of all samples. Zearalenone was detected in over 20% of all samples. Nivalenol and HT-2 toxin were each detected in only one sample. The survey clearly demonstrated regular occurrence of low levels of multiple mycotoxins in breakfast cereals on the Canadian market.     

Determination of ochratoxin A in organic and non-organic cereals and cereal products from Spain and Portugal

The objective of this work was to know the occurrence of OTA in organic and non-organic cereals and cereal products from Spain and Portugal. A method based on extraction with matrix solid phase dispersion (MSPD) using octylsilica (C₈) followed by liquid chromatography coupled with fluorescence detection (LC–FD) was used to determine OTA from the selected samples. Recoveries of OTA from the studied samples spiked at 10 ng/g level ranged from 78% to 89% with a standard deviation of 3.66. The limits of detection and quantification of this method were 0.05 and 0.19 ng/g, respectively. Furthermore, LC–FD after OTA methylation was used to confirm the identity of OTA in all positive samples. This procedure was applied to 83 organic and non-organic samples including rice, wheat, barley, rye, oats and maize from Spain and Portugal. OTA was detected in 22% of the samples, with concentrations ranging from 0.20 to 27.10 ng/g. From the total OTA contaminated samples (n = 18), 72% were organic cereal and 28% were non-organic cereal samples, with mean concentrations of 1.64 and 0.05 ng/g, respectively. The 66% and 34% of contaminated samples were from Spain and Portugal, respectively, with mean concentrations of 0.93 and 0.64 ng/g for each country. Six contaminated samples exceeded the maximum limits (ML) for OTA fixed by European Commission Regulation (5 μg/kg), among them three were from Spain and three from Portugal.     

Co-occurrence of aflatoxins,ochratoxin A and zearalenone in Capsicum powder samples available on the Spanish market

The aim of this study was to determine the co-occurrence of aflatoxins (AFs), ochratoxin A (OTA) and zearalenone (ZEA) in paprika and chilli samples purchased in Spain, using HPLC with fluorescence detection. The occurrence of mycotoxin in 64 paprika samples was 59% for AFs, 98% for OTA and 39% for ZEA, whereas in the 35 chilli samples, the contamination was 40% for AFs, 100% for OTA and 46% for ZEA. None of the samples had AFs levels higher than the legally allowable limits. Regarding the co-occurrence of mycotoxins, 75% of paprika samples and 65% of chilli samples contained more than one mycotoxin. Chilli samples generally had lower concentrations of AFB1, AFB2, total AFs and OTA than had paprika samples. The high incidence of OTA contamination suggests that additional legislation may be required to for these kinds of spices.     

Co‐occurrence of citrinin and ochratoxin A in rice in Asia and its implications for human health

Citrinin (CIT) and ochratoxin A (OTA) are nephrotoxic mycotoxins, produced by several Aspergillus and Penicillium species and their co‐occurrence in rice may cause health effects in humans. Rice is an important food crop worldwide and is a major staple food in Asia which may be invaded by CIT and OTA producing fungal spores in the field, during harvest and storage. Humans are exposed to these mycotoxins through ingestion of contaminated rice and other food commodities. Yet, data on the combined presence to these food contaminants are still insufficient to estimate human exposure in Asia. This review describes the prevalence of CIT and OTA in rice in Asia and its implications on human health, which may help in establishing and carrying out proper management strategies against mould development on rice. From the health point of view, combined exposition of CIT and OTA should be a public concern as both are nephrotoxic and long‐term exposure can pose detrimental health effects. Thus, it is necessary for local farmers and food factories to implement strict measures and to improve methods for rice preservation during the distribution to consumers, particularly in the markets. Moreover, regular surveys for CIT and OTA occurrence in rice and human biomonitoring are recommended to reduce the health effects in Asian population. © 2017 Society of Chemical Industry     

Occurrence of aflatoxin B1, citrinin and ochratoxin A in rice in five provinces of the central region of Vietnam

The possible coexistence of three mycotoxins in rice, including aflatoxin B1 (AFB1), citrinin (CIT) and ochratoxin A (OTA), was investigated. The samples of rice were collected in large markets in five provinces of the central region of Vietnam. These toxins were extracted, purified and finally quantified by HPLC with fluorimetry detection. Contamination of AFB1 was found to be the most, followed by OTA, while contamination of CIT was insignificant. The coexistence of CIT with AFB1/OTA in rice was found in high percentage. Some samples overpassed the authorized limit by Europe in OTA and/or AFB1.     

葡萄酒中桔青霉素快速检测间接竞争ELISA方法的建立

该研究利用桔青霉素（CIT）结构中的羧基、活泼氢和羟基分别与牛血清白蛋白（BSA）、卵清蛋白（OVA）和血蓝蛋白（KLH）进行偶联制备全抗原。经小鼠免疫,采用间接竞争酶联免疫吸附（IC-ELISA）法筛选出具有最佳竞争性反应的配对抗血清和包被抗原,并构建快速检测葡萄酒中CIT的IC-ELISA法。结果表明,最优抗血清为免疫CIT-KLH（H）的血清,包被抗原为CIT-BSA（H）,基于此构建的IC-ELISA法的检测线性范围为0.1～100 μg/L（R2=0.969 88）,检测限为0.1 μg/L,桔青霉素抗血清半数抑制浓度（IC50）值为9.8 μg/L,抗血清对沙丁胺醇、莱克多巴胺、展青霉素和黄曲霉毒素B1无交叉反应性。该方法在葡萄酒中检测CIT的加标回收率为92.01%～119.33%,加标回收率试验结果的相对标准偏差（RSD）为2.35%～5.22%,体积分数14%的乙醇对IC-ELISA检测结果无显著性影响（P＞0.05）,表明可直接上样快速检测葡萄酒中残留的桔青霉素。     

Validation of the procedure for the simultaneous determination of aflatoxins ochratoxin A and zearalenone in cereals using HPLC-FLD

Method validation for quantitative analysis of aflatoxins (AFs), ochratoxin A (OTA) and zearalenone (ZEA) in cereals using HPLC with fluorescence detector (FLD) is described. Mycotoxins were extracted with methanol?:?water (80?:?20) and purified with a multifunctional AOZ immunoaffinity column before HPLC analysis. The validation of the analytical method was performed to establish the following parameters: specificity, selectivity, linearity, limits of detection (LOD) and quantification (LOQ), accuracy, precision (within- and between-day variability), stability, robustness, measurement of performance, and measurement of uncertainty. Calibration curves were linear (r?>?0.999) over the concentration range, from the LOQ to 26, 40 and 400?ng/g for AFs, OTA and ZEA, respectively. LOD and LOQ were 0.0125 and 0.05?ng/g for aflatoxin B1 (AFB1) and G1 (AFG1), 0.0037 and 0.015?ng/g for aflatoxin B2 (AFB2) and G2 (AFG2), as well as 0.05 and 0.2?ng/g for OTA and 0.5 and 2?ng/g for ZEA, respectively. The mean recovery values were 77–104% for different concentrations of AFs, OTA and ZEA in spiked cereal samples. Both intra- and inter-day accuracy and precision were within acceptable limits. This method was successfully applied for the simultaneous determination of mycotoxins for 60 cereal samples collected from Malaysian markets. Fifty per cent of the cereal samples were contaminated with at least one of these mycotoxins, at a level greater than the LOD. Only one wheat sample and two rice samples were contaminated with levels greater than the European Union regulatory limits for AFs and OTA (4 and 5?ng/g). The means and ranges of mycotoxins obtained for the cereal samples were 0.4?ng/g and 0.01–5.9?ng/g for total AFs; 0.18?ng/g and 0.03–5.3?ng/g for OTA; and 2.8?ng/g and 2.4–73.1?ng/g for ZEA, respectively. The results indicate that the method is suitable for the simultaneous determination of AFs, OTA and ZEA in cereals and is suitable for routine analysis.     

Assessment of possible human exposure to ochratoxin A in Croatia due to the consumption of dry-cured and fermented meat products

Ochratoxin A (OTA) is a toxic secondary metabolite produced by the fungi of Aspergillus and Penicillium species. Data indicate a frequent OTA contamination of cereals and cereal products, and consequently also the contamination of meat and meat products. The aim of this study was to determine a possible level of meat product consumers’ exposure to OTA through the consumption of dry-cured and fermented meat products available on the Croatian market. Data showed the weekly OTA intake of 90% of male dry-cured ham consumers to be a maximum of 51.9 ng kg^–1 b.w., i.e., far below the tolerable weekly intake (TWI) of 120 ng kg^–1 b.w. weekly set out by the European Food Safety Authority (EFSA). OTA intake coming from the consumption of other meat products under study is lower and ranges from 0.1 to 42.1 ng kg^–1 b.w. weekly, dependent on the study. The study demonstrated that meat products in Croatia do not constitute a notable source of OTA in the human diet, so that the human health risk coming from the consumption of dry-cured and fermented meat products is negligible.     

Effects of ochratoxin A on micronucleus frequenncy in human lymphocytes

Ochratoxin A (OTA( a nephrotoxic and carcinogenic mycotoxin, was investigated to examine its potency to induce micronuclei (MN) in cultured human lymphocytes. Lymphocyte cultures were treated for the last 48 h with OTA at concentrations of 25 μM , 10 μM , 1 μM , 100 nM , 10 nM , 1 nM , and 100 pM and absolute ethanol. At the highest concentration, OTA was found to induce MN in cytokinesis‐blocked lymphocytes (p < 05). The 25 μM OTA concentration also led to a clear decrease in the percentage of binucleated cells, probably due to cytotoxicity. OTA at the other concentrations tested did not induce MN frequency. These results indicate that a high concentration of OTA is genotoxic in cultured human lymphocytes.     

Aflatoxins and ochratoxin A in flour: a survey of the Serbian retail market

This report presents data on the occurrence of aflatoxins (AF) and ochratoxin A in different types of flour marketed in Serbia. A total of 114 samples of wheat, buckwheat, rye, oat, barley, rice, millet and corn flour were collected in the period 2012–2016 and analysed using high performance liquid chromatography with fluorescence detection. Among flours other than corn, AFB1 was quantified only in rice, while ochratoxin A (OTA) was found in 29% of the samples. In corn flours the percentage of positive samples varied greatly over the years: AFB1 7.1–80.0%, OTA 30.0–40.6%, with a co-occurrence of 7.1–34.4%. Overall 5.2% of flours other than corn and 10.7% of corn flours exceeded the maximum levels (MLs) for AFB1 and/or OTA. The highest recorded levels were 8.80 μg kg^?1 of AFB1 (corn) and 23.04 μg kg^?1 of OTA (rye). Overall mean contamination levels of corn flours were 0.53 μg kg^?1 of AFB1 and 0.46 μg kg^?1 of OTA.     

Design of a sampling plan to detect ochratoxin A in green coffee

The establishment of maximum limits for ochratoxin A (OTA) in coffee by importing countries requires that coffee-producing countries develop scientifically based sampling plans to assess OTA contents in lots of green coffee before coffee enters the market thus reducing consumer exposure to OTA, minimizing the number of lots rejected, and reducing financial loss for producing countries. A study was carried out to design an official sampling plan to determine OTA in green coffee produced in Brazil. Twenty-five lots of green coffee (type 7 - approximately 160 defects) were sampled according to an experimental protocol where 16 test samples were taken from each lot (total of 16 kg) resulting in a total of 800 OTA analyses. The total, sampling, sample preparation, and analytical variances were 10.75 (CV = 65.6%), 7.80 (CV = 55.8%), 2.84 (CV = 33.7%), and 0.11 (CV = 6.6%), respectively, assuming a regulatory limit of 5 µg kg^-1 OTA and using a 1 kg sample, Romer RAS mill, 25 g sub-samples, and high performance liquid chromatography. The observed OTA distribution among the 16 OTA sample results was compared to several theoretical distributions. The 2 parameter-log normal distribution was selected to model OTA test results for green coffee as it gave the best fit across all 25 lot distributions. Specific computer software was developed using the variance and distribution information to predict the probability of accepting or rejecting coffee lots at specific OTA concentrations. The acceptation probability was used to compute an operating characteristic (OC) curve specific to a sampling plan design. The OC curve was used to predict the rejection of good lots (sellers' or exporters' risk) and the acceptance of bad lots (buyers' or importers' risk).     

Simple liquid chromatography assay for analyzing ochratoxin A in bovine milk

Ochratoxin A (OTA) is a mycotoxin with teratogenic and carcinogenic properties. Animal intake of feedstuffs contaminated with OTA may cause that some residues may be found in bovine milk, therefore, its analysis requires a highly sensitive, simple and precise technique.     

Effect of activated carbon on ochratoxin A reduction in “Pedro Ximenez” sweet wine made from off-vine dried grapes

The application effect of an activated carbon on Ochratoxin A (OTA) contents was studied in small-scale winemaking trials in musts from sun-dried grapes Pedro Ximenez variety. For this experiment a 2 × 3 × 4 factorial arrangement was used in the same batch, the factors being adsorbent (control and activated carbon), time (1, 2, and 3 h), and adsorbent dose (control, 0.12, 0.24, and 0.36 g/L). Immunoaffinity columns to sample clean-up and enzyme-linked immunosorbent technique were utilized for OTA determination. The two-parameter Freundlich equation was used to evaluate the adsorption data and the K _F values obtained ranging from 0.11 to 2.68. Activated carbon at 0.24 g/L dose was the most effective treatment to reduce OTA, up 70% in sweet wines. The mean percentages reduction of color parameters (intensity and tone) and total polyphenol index of the wines were 9.8, 3.5, and 4%, respectively. The study demonstrates that this treatment can be successfully applied as a suitable technique to reduce OTA contents of the musts and sweet wines during the early winemaking processes.     

Preliminary data on the presence of mycotoxins (ochratoxin A, citrinin and aflatoxin B1) in black table olives "Greek style" of Moroccan origin

Many mould strains, in particular Aspergillus and/or Penicillium, are able to develop on olive and produce ochratoxin A (OTA) and/or citrinin (CIT) and/or aflatoxin B (AFB) after harvest, during drying and storage of olives. The development of fungi on olives is responsible for the reduction of nutritional quality of olive because they can disturb the synthesis of the fatty acids. OTA, CIT and AFB are particularly dangerous for health, inducing cancer of urinary tracts or liver carcinoma. In this study, ten olive samples bought at retailer and at supermarket in Morocco were analyzed for their OTA, CIT and AFB contents. These three mycotoxins were extracted simultaneously by a method based on solvent partition validated in-house, then separated by HPLC coupled to a fluorescence detector. All olive samples contain OTA ranging from LOQ to 1.02 microg/kg. Respectively, 50 and 25% from retailer and supermarket samples were contaminated by more than 0.65 microg/kg. In addition, 80% of olive samples contained CIT above LOD, and 100% of olive tested contained AFB above 0.5 microg/kg. As simultaneous presence of these toxins increases toxic risks, it is thus essential to have a good control of the conservation of olives after harvest.