Aflatoxins and ochratoxin A in export quality raisins collected from different areas of Pakistan

During 2012–2014, 170 samples of export quality raisins were collected from different vendors in Pakistan. The collected samples were analysed for the presence of aflatoxins (AFs) and Ochratoxin A (OTA) contamination using high-performance liquid chromatography technique. The limit of detection and limit of quantification of AFs/OTA were 0.12/0.10 and 0.36/0.30 µg kg^?1, respectively. Only 5% of the samples were contaminated with AFs, ranging 0.15–2.58 µg kg^?1 with a mean of 0.05 ± 0.26 µg kg^?1. None of the raisin samples exhibited AFs contamination above the maximum limit (ML = 4 µg kg^?1) as set by the European Union (EU). About 72% of the samples were contaminated with OTA, ranging 0.14–12.75 µg kg^?1 with a mean of 2.10 ± 1.9 µg kg^?1. However, in 95.3% of the tested samples, OTA level was lower than the ML of 10 µg kg^?1 as regulated by the EU. Apparently, a strict and continuous monitoring plan, including regulatory limits, improves food safety and quality for all types of commodities.     

Fumonisin B1 in cereal mixtures marketed in Brazil

The aim of this study was to detect and quantify fumonisin B₁ (FB₁) in cereal mixtures marketed in Brazil. Fifteen samples from different lots were acquired each month by internet from supermarkets during seven months, adding up to 105 analysed samples. The unit sample constituted of an original package with a minimum of 250 g. Extraction and clean-up of samples for FB₁ determination were carried out using immunoaffinity columns. Identification and quantification of FB₁ were performed by high performance liquid chromatography. Eighty-eight (83.8%) samples were contaminated with FB₁ and four (3.8%) presented levels above 500 µg kg^?1 (634, 703, 1269 and 1876 µg kg^?1). Maximum FB₁ + FB₂ levels allowed by Brazilian regulations will reach 1500 µg kg^?1 for corn flour in 2016 and 1000 µg kg^?1 for others corn products. This study showed that even at levels below the legislative limits, human exposure to this toxin can occur constantly.     

Fumonisins and fungi in dry soybeans (Glycine Max L.) for human consumption

This survey reports the occurrence of fumonisins (FBs) and fungi in dry soybeans sold for human consumption. The variation levels were 138–1495 µg kg^?1 and 178–552 µg kg^?1 for FB₁ and FB₂, respectively. In addition, potentially toxigenic fungi as Fusarium, Aspergillus and Penicillium genera were isolated in the samples. These can be considered as indicator-toxin and can produce considerable amounts of mycotoxins. Despite FB presence in the soybeans for human consumption, there is no legal regulation. Therefore, it is important to emphasise the need for frequent monitoring of these contaminants in soybeans.     

Fumonisin B2 production by Aspergillus niger from grapes and natural occurrence in must

Aspergillus niger has been recently found to produce fumonisin B₂ (FB₂). Thirty-one strains belonging to four Aspergillus species isolated from grape were evaluated for FB₂ production on agar plates. Four out of eight strains of A. niger produced FB₂ (29–293 µg g^?1). None of the strains of A. uvarum (n = 7), A. tubingensis (8) and A. carbonarius (8) produced detectable amounts of toxin. The capability to produce FB₂ was also confirmed by some A. niger strains artificially inoculated on grape berries. Natural occurrence of FB₂, at levels of 0.01 and 0.4 µg ml^?1, was found in two samples of must collected in Apulian cellars in 2007. This is the first report of FB₂ contamination in must. These findings suggest that there is a potential risk of exposure to FB₂ in the grape–wine chain for consumers and that A. niger may represent the major fumonisin-producing species among black Aspergilli occurring on grapes.     

Fumonisins B1, B2 and B3 in corn products,wheat flour and corn oil marketed in Shandong province of China

In this study a total of 522 samples were collected from Shandong province of China in 2014 and analysed for the occurrence of fumonisin B1 (FB₁), FB₂ and FB₃ by isotope dilution ultrahigh performance liquid chromatography–tandem mass spectrometry. Fumonisins were detected in 98.1% of the corn products, with the average total level of 369.2 μg kg^?1. The individual average values of FB₁, FB₂ and FB₃ in corn products were 268.3, 53.7 and 47.2 μg kg^?1, respectively. The simultaneous occurrence of FB₁, FB₂ and FB₃ was observed in 76.7% of the corn products. Especially, the results demonstrated that the difference in the contamination levels for fumonisins in these three types of corn products was apparent. In addition, 6.2% of the wheat flour samples were contaminated with FB₁, with concentrations ranging from 0.3 to 34.6 µg kg^?1. No FB₂ or FB₃ was detected in wheat flour. In corn oil samples no fumonisins were detected.     

Effects of different drying treatments on fungal population and ochratoxin A occurrence in sultana type grapes

This study aimed to determine the changes in mould and ochratoxin A (OTA) occurrence in sultanas under three different conventional drying conditions. Five different vineyards were chosen, and the three different treatments were applied to these grapes while drying. At the end of the drying process, total mould and black aspergilli (BA) populations in the samples varied from 2.45 to 5.61 log colony-forming units (CFU) g^–1 and from 0 to 4.92 log CFU g^–1, respectively. Significant increases (p < 0.05) occurred in mould loads depending on the extending drying period. However, independent of vineyard location, all the samples treated with cold dipping solution showed the lowest fungal loads. These results indicate that dipping solution treatment was the most effective drying method to minimise fungal infection of grapes. The expected results could not be achieved by drying grapes artificially contaminated with ochratoxigenic Aspergillus carbonarius spores. Seventy-one of 96 isolates (73.95%) obtained during drying were Aspergillus spp., and the remaining (n = 25, 26.05%) belonged to other genera, such as Penicillium, Trichoderma and Cladosporium. Grape juice-based agar medium was used to determine the realistic OTA production capacities of the isolated mould strains. The highest OTA production capacities were 809.70 ± 9.19, 87.58 ± 16.89 and 45.44 ± 18.78 ng g^–1 in 50% grape juice agar (GJ50), all five of which were from A. niger isolates. OTA was not present in any sample during the drying period; however, OTA was detected in two samples at 0.32 ± 0.15 and 0.52 ± 0.36 µg kg^–1 after the end of the drying process. The limit of detection (LOD) and limit of quantitation (LOQ) of the method used for detecting OTA in samples were 0.1 and 0.3 µg kg^–1, respectively.     

Fumonisins B1 and B2 in the corn-milling process and corn-based products,and evaluation of estimated daily intake

The distribution of fumonisins (FBs: FB₁ and FB₂) in the corn-milling process and in corn-based products, as well as daily intake estimates for the Brazilian population were evaluated. Among corn fractions samples, corn meal had the highest mean concentration of FB₁ (1305 µg kg^?1) and FB₂ (651 µg kg^?1) and a distribution factors of 452% and 256% in relation to corn grain, respectively. On the other hand, the distribution factor of FB₁ and FB₂ in corn flour was found to be 144% and 88% respectively, which demonstrates that fumonisins in this fraction were reduced compared with corn grain. As a result, almost half the corn meal samples (47%) would be non-compliant with future Brazilian regulation (2017) for fumonisins. However, corn-based products, such as corn flakes and popcorn, were in compliance with the regulation. The average probable daily intake and maximum probable daily intake of fumonisins estimated for the Santa Catarina state (Brazil) population were below the provisional maximum tolerable daily intake of 2 µg kg^?1 body weight day^?1 for all corn samples. Despite this, the adoption of practices to control the occurrence of fumonisins should be applied to the corn-milling fractions that may contain a higher concentration of this toxin, such as corn meal, often used for animal feed in Brazil.     

Fumonisins in corn (Zea mays L.) from Southern Brazil

A total of 232 samples of corn commercialised in Santa Catarina state, Southern Brazil (temperate zone climate), were evaluated from 2007 to 2012 for fumonisins (FBs: FB₁ and FB₂). Before performing this study, a FBs method with liquid chromatography and fluorescence detection (ex. 335; em. 440 nm) was validated first. FBs were detected in 46.6% (108 samples), with values ranging from 66 to 7832 µg kg^?1 for FB₁ and 110 to 1201 µg kg^?1 for FB₂. The number of contaminated corn samples for FB₁ and FB₂ varied and often presented contamination of FB₁ only. Per year of analysis, the numbers were: n = 22/8(FB₁/FB₂), 44/5(FB₁/FB₂), 25/12(FB₁/FB₂), 4(FB₁), 6(FB₁) and 7(FB₁) in 2007, 2008, 2009, 2010, 2011 and 2012, respectively. The contamination percentage was 42.3/15.4, 59.5/6.8, 43.8/21.1, 36.4, 35.3 and 33.3%, respectively, during these years. Consumers can be exposed to these mycotoxins and their health can be at risk through the consumption of contaminated corn.     

Improved sensitivity of ochratoxin A analysis in coffee using high-performance liquid chromatography with hybrid triple quadrupole-linear ion trap mass spectrometry (LC-QqQLIT-MS/MS)

A novel and sensitive method utilising high-performance liquid chromatography coupled to triple quadrupole-linear ion trap mass spectrometry (LC-QqQLIT-MS/MS) was developed in order to analyse the content of ochratoxin A (OTA) in coffee samples. The introduction of the triple-stage MS scanning mode (MS³) has been shown to increase greatly sensitivity and selectivity by eliminating the high chromatographic baseline caused by interference of complex coffee matrices. The analysis included the sample preparation procedure involving extraction of OTA using a methanol–water mixture and clean-up by immunoaffinity columns and detection using the MS³ scanning mode of LC-QqQLIT-MS/MS. The proposed method offered a good linear correlation (r² > 0.998), excellent precision (RSD < 2.9%) and recovery (94%). The limit of quantification (LOQ) for coffee beans and espresso beverages was 0.010 and 0.003 µg kg^?1, respectively. The developed procedure was compared with traditional methods employing liquid chromatography coupled to fluorescent and tandem quadrupole detectors in conjunction with QuEChERS and solid-phase extraction. The proposed method was successfully applied to the determination of OTA in 15 samples of coffee beans and in 15 samples of espresso coffee beverages obtained from the Latvian market. OTA was found in 10 samples of coffee beans and in two samples of espresso in the ranges of 0.018–1.80 µg kg^?1 and 0.020–0.440 µg l^?1, respectively. No samples exceeded the maximum permitted level of OTA in the European Union (5.0 µg kg^?1).     

Co-occurrence of aflatoxins,ochratoxin A and citrinin in “egusi” melon (Colocynthis citrullus L.) seeds consumed in Ireland and the United Kingdom

The natural co-occurrence of aflatoxins (AFs), ochratoxin A (OTA) and citrinin (CIT) in melon seed samples obtained from retailers and households in Ireland and the United Kingdom (UK) was evaluated. AFs and OTA were determined by HPLC with fluorescence detection while CIT was analysed by HPLC-MS/MS. AFB₁ was detected in all (100%) samples (mean = 9.7 μg kg^?1; range = 0.2–66.5 μg kg^?1). Mean total AFs was 12.0 μg kg^?1 (range = 0.3–82 μg kg^?1). Commercially retailed samples showed a significantly higher AFB₁ contamination (p < 0.05) than the household samples. OTA occurred in 3 (13.6%) samples, while 4 (18.2%) were contaminated with CIT at very low levels. In this study, 68% of the melon seed samples were contaminated above the 2 μg kg^?1 EU limit for AFB₁ in oilseeds. These results highlight the need for the development of strategies to reduce AF contamination in “egusi” for human consumption.     

Aflatoxins in animal feed in Iran

One hundred and forty-six samples of animal feed (barley, n = 60; wheat bran, n = 22; wheat dry pulp, n = 29; and canola meal, n = 35) were collected in 2011 from Mashhad (Khorasan, Iran). Aflatoxins (AFs) were determined in these samples after immunoaffinity column clean-up by high-performance liquid chromatography (HPLC) with fluorescence detection. Aflatoxin B₁ (AFB₁) contamination was found in 28 samples: in five of the barley samples (8.3%) at a mean level of 0.48 µg·kg^?1, in two wheat bran samples (9.0%) at a mean level of 0.88 µg·kg^?1, in 10 wheat dry pulp samples (34.5%) at a mean level of 0.30 µg·kg^?1 and in 11 canola meal samples (31.4%) at a mean level of 0.92 µg·kg^?1. AFB₁ levels were below the maximum levels of Iran regulations (5 µg·kg^?1) and the EU maximum limit (5 µg·kg^?1).     

Ochratoxin A in artisan salami produced in Veneto (Italy)

Fifty samples of artisan salami purchased in Veneto (Italy) were analysed for the determination of ochratoxin A (OTA). The analytical method, based on a sample preparation procedure with immunoaffinity columns (IACs), together with analysis by high-performance liquid chromatography with fluorescence detection (HPLC-FD), has guaranteed a high rate of recovery (about 97%), limit of detection (LOD) and limit of quantification (LOQ), respectively, of 0.06 µg kg^?1 and 0.20 µg kg^?1. OTA was detected in five samples, but only one exceeded the guideline value (1 µg kg^?1) established by the Italian Ministry of Health for pork meat and derived products. The results would seem to suggest that salami made with the traditional, non-industrial production method can be considered safe as regards contamination by OTA. However, the very high concentration observed in one sample proves that a high OTA contamination is also possible in this type of product. Thus, the controls of mycotoxin contamination must consider also salami.     

Aflatoxins in composite spices collected from local markets of Karachi,Pakistan

This survey was carried out to evaluate the occurrence of total aflatoxins (AFs; B₁+B₂+G₁+G₂) in unpacked composite spices. A total of 75 samples of composite spices such as biryani, karhai, tikka, nihari and korma masalas were collected from local markets of Karachi, Pakistan, and analysed using HPLC technique. The results indicated that AFs were detected in 77% (n = 58) samples ranging from 0.68 to 25.74 µg kg^?1 with a mean of 4.63 ± 0.95 µg kg^?1. In 88% (n = 66) samples, AFs level was below the maximum limits (ML = 10 µg kg^?1) as imposed by EU. Furthermore, 61% (n = 46) tested samples contained AFs level between 1 and 10 µg kg^?1, 9% (n = 7) exhibited AFs contamination ranged 10?20 µg kg^?1 and only 3% (n = 2) of the investigated samples contained AFs levels higher than the ML of 20 µg kg^?1 for total aflatoxins as set by the USA. It was concluded that there is need to establish a strict and continuous national monitoring plan to improve safety and quality of spices in Pakistan.     

Ochratoxin A in red pepper flakes commercialised in Turkey

The aim of this study was to investigate the presence of Ochratoxin A (OTA) in red pepper flakes commercialised in Turkey. A total of 75 samples were collected from different supermarkets and traditional bazaars in Istanbul during 2012–2013. OTA analysis was performed by high-performance liquid chromatography equipped with fluorescence detection after immunoaffinity column clean-up. The method was linear in the range 0.05–40 μg kg^?1 (r² = 0.9997). Twenty-seven out of 31 (87.1%) packed red pepper flake samples contained OTA at concentrations ranging from 0.6 to 1.0 μg kg^?1, whereas 100% of the unpacked red pepper flake samples contained OTA, in the range 1.1–31.7 μg kg^?1. Overall, only 4 unpacked samples (5.3%) contained OTA, with a mean value of 23.4 μg kg^?1, which is higher than the European Union limit. It is suggested that OTA content should be carefully considered in red pepper flake samples especially marketed in unpacked form.     

Quantification of mycotoxins in vegetable oil by UPLC-MS/MS after magnetic solid-phase extraction

ABSTRACT

The detection of mycotoxin contamination in foodstuffs is highly significant for public health. Herein we report an analytical method based on magnetic solid-phase extraction (MSPE) and UPLC-MS/MS for the simultaneous determination of mycotoxins, including fumonisins B₁ (FB₁), zearalenone (ZON) and ochratoxin A (OTA), in vegetable oil. Magnetic nanoparticles coated with double layers of silicon dioxide were synthesised and found to be an effective MSPE adsorbent for mycotoxins. The proposed MSPE procedure serves not only for sample clean-up but also for mycotoxin enrichment that enhances greatly the assay’s sensitivity. Under the selected MSPE conditions, linear matrix-matched calibration curves were obtained for mycotoxins in a concentration range from 0.178 to 625 μg kg^–1. The limits of detection were 0.210 μg kg^–1 for FB₁, 0.0800 μg kg^–1 for OTA and 1.03 μg kg^–1 for ZON. The proposed MSPE UPLC-MS/MS method was applied for the determination of mycotoxins in vegetable oil samples, including maize oil, rapeseed oil and soybean oil. ZON was detected in a maize oil at 101 μg kg^–1, which is below the European Union limit of 200 μg kg^–1 in foodstuffs.     

Aflatoxin B1 in betel nuts (Areca catechu L.) imported to Pakistan from different regions of South Asia

Aflatoxin B₁ (AFB₁) levels were evaluated in betel nuts (Areca catechu L.) being imported to Pakistan during 2010–2011. In total, 278 betel nut samples (India = 21, Indonesia = 51, Sri-Lanka = 34 and Thailand = 172) were received from the Department of Customs and were analysed by thin layer chromatography (TLC). All Indian origin betel nuts showed AFB₁ contamination ranging from 11.7–262.0 µg kg^?1 with a mean of 92.5 µg kg^?1. Among Indonesian and Sri Lankan shipments, 80.4% and 73.5% betel nuts were contaminated with AFB₁ ranging between 3.3–39.2 and 6.5–103.4 µg kg^?1 with a mean of 11.6 and 35.0 µg kg^?1, respectively. However, only 30.2% of Thailand origin samples showed AFB₁ contamination ranging 3.3–77.0 µg kg^?1 with a mean of 6.6 µg kg^?1. The widespread occurrence of AFB₁ increases the hazard associated with betel nuts. Thus, strict control is a pre-requisite for the production and import/export of psychoactive substances as betel nuts.     

Aflatoxin M1 in raw milk from different regions of São Paulo state – Brazil

A total of 635 raw milk samples from 45 dairy farms, from three regions of São Paulo state – Brazil, were evaluated during 15 months for aflatoxin M₁ (AFM₁). AFM₁ was determined by high performance liquid chromatograph with fluorescence detection. AFM₁ was detected (>0.003 µg kg^?1) in 72.9%, 56.3% and 27.5% of the samples from Bauru, Araçatuba and Vale do Paraíba regions, respectively. The mean AFM₁ contamination considering all the samples was 0.021 µg kg^?1. Furthermore, the concentration of AFM₁ was quite different among Bauru (0.038 µg kg^?1), Araçatuba (0.017 µg kg^?1) and Vale do Paraíba (<0.01 µg kg^?1) regions. Only three samples (0.5%) had higher contamination than the tolerated limit in Brazil (0.50 µg kg^?1) and 64 samples (10.1%) had a higher contamination than the maximum limit as set by the European Union (0.050 µg kg^?1). The estimated AFM₁ daily intake was 0.358 and 0.120 ng kg^?1 body weight per day for children and adults, respectively.     

Determination of mycotoxins in biscuits,dried fruits and fruit jams: an assessment of human exposure

ABSTRACT

A reliable, fast and simple method using UHPLC-MS/MS was developed for the determination of aflatoxins B₁ (AFB1), G₁ (AFG1), B₂ (AFB2) and G₂ (AFG2), ochratoxin A (OTA), deoxynivalenol (DON), zearalenone (ZEA), HT-2 toxin and T-2 toxin in crude extracts of biscuits with fruit filling, cookies, dried fruits and fruit jams. The method was successfully demonstrated on 39 samples of biscuits with fruit filling, 34 cookies, 14 dried fruits and 10 fruit jams. The mycotoxins detected in biscuits samples were ZEA, OTA, T-2 and AFB1 with an average concentrations of positive samples of 2.64, 4.10, 8.13 and 1.32 µg kg^?1, respectively; while the mycotoxins detected in jam samples were AFB1, OTA, T-2 and AFB2 with an average concentrations of positive samples of 2.00, 17.7, 4.37 and 1.15 µg kg^?1, respectively. The results showed that the majority of samples were in compliance with relevant regulations. However in eight samples of biscuits and three samples of fig jam the contents of OTA were higher than the existing OTA limits. The combined dietary exposure of selected mycotoxins was estimated for the first time for children, adolescents and adults. The estimated combined dietary exposures were all lower than the proposed value assumed to predict a possible risk scenario.     

Mycotoxin contamination of home-grown maize in rural northern South Africa (Limpopo and Mpumalanga Provinces)

The aim of this study was to assess mycotoxin contamination of crops grown by rural subsistence farmers over two seasons (2011 and 2012) in two districts, Vhembe District Municipality (VDM, Limpopo Province) and Gert Sibande District Municiality (GSDM, Mpumalanga Province), in northern South Africa and to evaluate its impact on farmers’ productivity and human and animal health. A total of 114 maize samples were collected from 39 households over the two seasons and were analysed using a validated liquid chromatography-tandem mass spectrometry mycotoxins method. Aflatoxin B₁ (AFB₁) occurrence ranged from 1 to 133 µg kg^?1 in VDM while AFB₁ levels in GSDM were less than 1.0 µg kg^?1 in all maize samples. Fumonisin B₁ levels ranged from 12 to 8514 µg kg^?1 (VDM) and 11–18924 µg kg^?1 (GSDM) in 92% and 47% positive samples, respectively, over both seasons. Natural occurrence and contamination with both fumonisins and aflatoxins in stored home-grown maize from VDM was significantly (p < 0.0001) higher than from GSDM over both seasons.     

Effect of post-harvest treatments on the occurrence of ochratoxin A in raw cocoa beans

Cocoa beans are the principal raw material for chocolate manufacture. Moulds have an important place in the change in the quality of cocoa beans due to their role in the production of free fatty acids and mycotoxins, namely ochratoxin A (OTA). This study investigated the impact of the key post-harvest treatments, namely the fermentation and drying methods on OTA contamination of raw cocoa beans. Analytical methods for OTA detection were based on solid–liquid extraction, clean-up using an immunoaffinity column, and identification by reversed-phase HPLC with fluorescence detection. Of a total of 104 randomly selected cocoa samples analysed, 32% had OTA contents above 2 µg kg^–1. Cocoa sourced from pods in a bad state of health had a maximum OTA content of 39.2 µg kg^–1, while that obtained from healthy pods recorded 11.2 µg kg^–1. The production of OTA in cocoa beans increased according to the pod-opening delay and reached 39.2 µg kg^–1 after an opening delay of 7 days after harvest, while 6.1 and 11.2 µg kg^–1 were observed when pods were opened after 0 and 4 days. OTA production also seemed to depend considerably to the cocoa fermentation materials. When using plastic boxes for bean fermentation, the OTA production was enhanced and reached an average OTA content of about 4.9 µg kg^–1, while the raw cocoa treated in banana leaves and wooden boxes recorded 1.6 and 2.2 µg kg^–1 on average respectively. In parallel, the OTA production was not really influenced by either the mixing or the duration of the fermentation or the drying materials.