Aflatoxins contamination of maize in Serbia: the impact of weather conditions in 2015

ABSTRACT

In recent years climate changes recorded in temperate regions of Europe have led to aflatoxin (AF) contamination of maize. Thus, the aim of this study was to investigate the influence of weather conditions on levels of aflatoxin B₁ (AFB1), aflatoxin B₂ (AFB2), aflatoxin G₁ (AFG1) and aflatoxin G₂ (AFG2) in 180 maize samples collected from the main maize-growing regions (Western Ba?ka, North Banat, South Banat and Central Serbia) in Serbia after harvest in 2015. The concentrations of AFs were determined by a validated HPLC method with post-column derivatisation and fluorescence detection (HPLC-FLD). The presence of AFB1, AFB2, AFG1 and AFG2 was detected in 57.2%, 13.9%, 5.6% and 2.8% of maize samples in the concentration ranges of 1.3–88.8 µg kg^–1, 0.60–2.8 µg kg^–1, 1.8–28.5 µg kg^–1 and 2.1–7.5 µg kg^–1 respectively. The recorded smaller amount of precipitation and especially higher air temperatures during the summer of 2015 were favourable for AF production, which resulted in 32.2% and 21.1% of samples being unsuitable for human consumption, since AFB1 and the sum of AFs concentrations were above 5.0 and 10.0 µg kg^–1 respectively. Furthermore, the findings in this study indicate that the microclimate conditions in the investigated regions had a great influence on the contamination frequency of maize with AFs. The highest percentage of samples unsuitable for human consumption, considering both AFB1 and total AFs content were 72.5% and 51.5% respectively from Central Serbia, whilst the lowest percentages of 15.6% and 6.2% respectively were found in Western Ba?ka. These findings confirmed that maize should be continuously monitored in order to protect human and animal health from the harmful effects caused by AFs contamination.     

Survey of 11 mycotoxins in wheat flour in Hebei province,China

A survey of 11 mycotoxins in 348 wheat flour samples marketed in Hebei province of China were analysed by liquid chromatography-tandem mass spectrometry, was carried out. The selected mycotoxins consisted of four aflatoxins (AFs: AFB₁, AFB₂, AFG₁ and AFG₂) and seven Fusarium toxins, i.e. deoxynivalenol (DON), nivalenol, 3-acetyldeoxynivalenol and 15-acetyldeoxynivalenol, zearalenone, Fusarenon-X and deoxynivalenol-3-glucoside. Results indicated that most of the wheat samples analysed were contaminated with mycotoxins. Wheat was most susceptible to DON (91.4% contamination), with a mean level of 240 μg kg^?1. On average the probable daily intake (PDI, expressed as µg kg^?1 body weight day^?1) of mycotoxins was within the provisional maximum tolerable daily intake (PMTDI, 2.0 µg kg^?1 of body weight day^?1) as set by the Joint FAO/WHO Expert Committee on Food Additives. Nevertheless, exposure assessment revealed that the maximum PDI of mycotoxins was 4.06 µg kg^?1 body weight day^?1, which was twice the PMTDI value. Thus, consistent monitoring is recommended, as to keep the contamination level under control.     

Aflatoxins,ochratoxins and zearalenone in sorghum and sorghum products in Sudan

This survey examined 60 samples of sorghum and 30 samples of sorghum products from three states (Khartoum, Kordofan and Gadarif) of Sudan for aflatoxin B1, B2, G1 and G2 (AFB1, AFB2, AFG1, AFG2), ochratoxin A and B (OTA, OTB) and zearalenone (ZEN), using high performance liquid chromatography with fluorescence detection. The limits of detection and limits of quantification were in the range 0.01–0.6 µg kg^–1 and 0.03–2.0 µg kg^?1, respectively. The frequency of contaminated samples with AFB1 from Khartoum, Gadarif and Kordofan state was 38.1%, 22.2% and 23.8%, respectively. Only two samples of sorghum from Khartoum state were contaminated with OTA (3.3%). Concentrations of OTA and OTB were low and may not cause problems. No sample of sorghum or sorghum products was contaminated with ZEN. Some sorghum samples contained AFB1 concentrations above the European Union regulatory limits. The highest contaminated samples were found in Khartoum state.     

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.     

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.     

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.     

Evolution of fungal population and mycotoxins in sorghum silage

Silage, one of the most important feed sources for cattle, is vulnerable to contamination by spoilage moulds and mycotoxins because ensilage materials are excellent substrates for fungal growth. The aim of this study was to identify the mycobiota of sorghum silages, to determine the presence of aflatoxins and fumonisins, and to correlate these results with physical parameters of the silage. A total of 275 samples of sorghum were collected from dairy farms in the south-west region of Uruguay were silage practices are developed. The presence of fungi was observed in all of the sorghum samples with values varying from 0.2 × 10⁴ to 4085 × 10⁴ UFC g^?1. Significant difference were detected in the total number of fungi during the storage period; at six months there is a high risk of fungal spoilage. The most frequent genera isolated from sorghum samples were Penicillium (70%), Aspergillus (65%), Absidia (40%), Fusarium (35%), Paecilomyces (35%) and Alternaria, Cladosporium, Gliocadium and Mucor (30%). The toxigenic species most frequently found were Penicillium citrinum, Aspergillus flavus and Fusarium nygamai. Only two samples were contaminated by AFB1 with levels of 1 and 14 µg kg^–1. Fumonisin was detected in 40% of freshly harvest samples with levels ranged from 533 µg kg^–1 to 933 µg kg^–1. The use of silo bags seems to be an effective tool to store sorghum. However, the presence of toxigenic fungi show that regular screening for mycotoxins levels in silages must be performed to avoid the exposure of animals to contaminated feed and the introduction of these compounds into the food chain.     

Development and validation of an UHPLC-MS/MS method for the determination of mycotoxins in grass silages

An ultra-high-performance liquid chromatography-tandem mass spectrometry (UHPLC-MS/MS) multi-mycotoxin analytical method was developed to simultaneously identify and quantify 20 mycotoxins in grass silages, inclusive of mycotoxins that are currently regulated in European Union feeds. Extraction of mycotoxins from dried grass silages was performed using of a modified QuEChERS extraction employing an acidified aqueous extraction (0.1 N HCl) with no further clean-up. Following chromatographic separation, analytes were detected using a fast polarity-switching MS/MS method that allowed both positive and negative ions to be analysed from a single injection, thus the reducing time and cost of analysis. The limits of detection and quantification ranged between 3 µg kg^–1 DM (aflatoxin B₁, beauvericin and enniatin A and A₁) and 200 µg kg^–1 DM (deoxynivalenol), and between 10 µg kg^–1 DM (aflatoxin B₁, beauvericin and enniatin A₁) and 500 µg kg^–1 DM (deoxynivalenol), respectively. Inter-assay accuracy and precision ranged between 90% and 107% and between 3.9% and 15.0% CV, respectively. The accuracy of the method was assessed through the application to a range of incurred samples in an inter-laboratory study.     

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.     

Occurrence of enniatins and beauvericin in 60 Chinese medicinal herbs

A total of 60 Chinese medicinal herbs were examined for contamination of the emerging Fusarium mycotoxins enniatins (ENNs) A, A1, B, B1 and beauvericin (BEA). The herbs under study are commonly used in China as both medicines and food. The dried samples of herbs were randomly collected from traditional Chinese medicine stores in Zhejiang province, China. Sample preparation was achieved by methanol extraction, followed by a simple membrane filtration step; no tedious clean-ups were involved. ENNs A, A1, B, B1 and BEA were analysed by the recently developed stable isotope dilution assays, using liquid chromatography-tandem mass spectrometry (LC-MS/MS). With limits of detection ranging between 0.8 and 1.2 µg kg^–1 for the analytes under study, 25% of all analysed samples were contaminated with at least one of the ENNs and BEA. BEA was the most frequently detected toxin with a 20% incidence in all samples. The percentages of ENN-positive samples were lower: each single ENN was detected in 6.7–11.7% of all samples. Considering the total amounts of the five mycotoxins in single samples, values between 2.5 and 751 µg kg^–1 were found. The mean total amount in positive samples was 126 µg kg^–1. Regarding ginger, the frequent occurrence of ENNs and BEA in dried ginger could be confirmed in samples from Germany. However, in fresh ginger root the toxins were not detectable. This is the first report on the presence of ENNs and BEA in Chinese medicinal herbs.     

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.     

Dietary exposure to mycotoxins of the Hong Kong adult population from a Total Diet Study

Dietary exposure of Hong Kong adults to mycotoxins and their metabolites including aflatoxins (AFs), ochratoxin A (OTA), fumonisins (FNs), deoxynivalenol (DON), acetyldeoxynivalenols (AcDONs) and zearalenone (ZEA) was estimated using the Total Diet Study (TDS) approach to assess the associated health risk to the local people. Sixty commonly consumed food items, collected in four seasons, were sampled and prepared as consumed. These mycotoxins were primarily found at low levels. The highest mean levels (upper bound) were: AFs, 1.50 µg kg^–1 in legumes, nuts and seed; OTA, 0.22 µg kg^–1 in sugars and confectionery; FNs, 9.76 µg kg^–1 in cereals and their products; DON and AcDONs, 33.1 µg kg^–1 in cereals and their products; and ZEA, 53.8 µg kg^–1 in fats and oils. The estimated dietary exposures of Hong Kong adults to the mycotoxins analysed were well below the respective health-based guidance values, where available. For AFs, the upper-bound exposure for high consumers is 0.0049 µg kg bw^–1 day^–1, which was estimated to contribute to about 7.7 (< 1%) of liver cancer cases when compared with 1222 liver cancer cases per year in Hong Kong. The percentage contributions of the estimated 95th percentile dietary exposures (lower and upper bound) to the health-based guidance values of individual mycotoxins were: ochratoxin A, 3.6–9.2%; fumonisins, 0.04–8.5%; deoxynivalenol and acetyldeoxynivalenols, 21.7–28.2%; and zearalenone 3.3–34.5%. The findings indicate that dietary exposures to all the mycotoxins analysed in this study were unlikely to pose an unacceptable health risk to the Hong Kong population.     

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.     

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.     

Occurrence of ochratoxin A,fumonisin B2 and black aspergilli in raisins from Western Greece regions in relation to environmental and geographical factors

The presence of ochratoxin A (OTA), fumonisin B₂ (FB₂) and black aspergilli in raisins from Western Greece regions (Messinia, Corinthia, Achaia, Ilia and Zante Island) was investigated in relation to the different geographic and climatic conditions in the 2011 growing season. The biseriate species Aspergillus niger “aggregate” and A. carbonarius were mainly identified. The population of A. niger “aggregate” species occurred in all raisin samples at colony-forming units (CFU) concentrations significantly higher (mean 2.2 × 10⁵ CFU g^?1 homogenate) than those of A. carbonarius population (mean 4.9 × 10³ CFU g^?1 homogenate), which occurred in 80% of the raisin samples. OTA was found in 73% of the samples at levels ranging from 0.1 µg kg^?1 to 98.2 µg kg^?1, with the highest level occurring in a raisin sample from Ilia that also contained the highest level of A. carbonarius. The European Union legal limit for OTA was exceeded in 15% of the raisin samples. FB₂ was found in 29% of the raisin samples at levels ranging from 7.1 µg kg^?1 to 25.5 µg kg^?1, with 20% of the samples co-occurring with OTA. Principal-component analysis was applied to levels of mycotoxins, fungal contamination, geographical data and environmental conditions recorded in the harvesting (August) or drying (September) period. Principal-component analysis clearly indicated a good direct correlation of rainfall and relative humidity with OTA and A. carbonarius contamination. A lack of clustering was observed when A. niger and FB₂ contamination were considered. This is the first report on the co-occurrence of the mycotoxins OTA and FB₂ in dried vine fruits from Greece.     

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.     

Deoxynivalenol in wheat,maize, wheat flour and pasta: surveys in Hungary in 2008–2015

Among Fusarium mycotoxins, deoxynivalenol (DON) is the most common contaminant in case of cereals and cereal-based foods in Hungary. In this study, Hungarian wheat (n = 305), maize (n = 108), wheat flour (n = 179) and pasta (n = 226) samples were analysed (N = 818). The samples were collected during 2008–2015 in Hungary. Applied methods of analysis were enzyme-linked immunosorbent assay and liquid-chromatography coupled with a mass spectrometer. Results were compared and evaluated with Hungarian weather data. Among cereal samples, in 2011, wheat was contaminated with DON (overall average ± standard deviation was 2159 ± 2818 µg kg^?1), which was above the maximum limit (ML). In case of wheat flour and pasta, no average values above ML were found during 2008–2015, but higher DON contamination could be observed in 2011 as well (wheat flour: 537 ± 573 µg kg^?1; pasta: 511 ± 175 µg kg^?1).     

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.     

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).     

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.