Occurrence Of Ochratoxin A Research Articles

Determination of ochratoxin a in wine by high-performance thin-layer chromatography using charged coupled device

A method was developed and validated in-house for determination of ochratoxin A (OTA) in wine using charge coupled device (CCD) to acquire the fluorescence images of mycotoxin from high-performance thin-layer chromatography (HPTLC) under UV lamp. The method showed a mean recovery of 90.4%. The quantification and detection limist were 0.1 µg L-1 and 0.016 µg L-1 per spot, respectively. The results of validation confirmed the efficiency of the method, which is sensitive enough to be used to quantify OTA in wine. The occurrence of OTA in Brazilian wines was evaluated. OTA was found in one sample at a level of 4.5 µg L-1, which is higher than the limit of 2 µg L-1 considered acceptable by the Scientific Commission of the European Communities. This study demonstrated the applicability of HPTLC using CCD as a tool to determine OTA in wine.

Aspergillus section Nigri species isolated from different wine-grape growing regions in Argentina

Vineyards located in eight grape growing regions of Argentina during the harvest season 2006/07 were evaluated. The aims of this study were to determine the incidence of Aspergillus section Nigri, their ability to produce ochratoxin A (OTA) and to evaluate the OTA natural occurrence in grapes. Bunches of grapes at maturation stage were collected, and grapes (50 per sample) were plated on Petri dishes containing dichloran-glycerol 18% agar (DG18) and dichloran-rose bengal-chloramphenicol agar (DRBC) media. After an incubation period of 7 days at 25 °C, the mycoflora belonging to Aspergillus section Nigri was identified. OTA occurrence and the toxicogenic ability of the strains were analyzed by HPLC. A. niger aggregate strains were dominant showing the highest infection percentage (81%), followed by A. carbonarius (11%) and Aspergillus uniseriate (8%). A. carbonarius strains presented the highest percentage of OTA-producer strains (82%) and the highest toxin levels (mean 202 ng/g). A positive correlation between the isolation percentage of A. carbonarius in grapes and temperature was found. The warmest regions showed the highest A. carbonarius incidence. OTA was detected at low levels in grapes during the survey. OTA levels in grapes and rain at harvest time correlated positively.

Occurrence of total aflatoxins, ochratoxin A and deoxynivalenol in foodstuffs available on the Lebanese market and their impact on dietary exposure of children and teenagers in Beirut

Mycotoxins are fungal metabolites commonly present in food and constituting a potential threat to food safety. Total aflatoxins (AFT), ochratoxin A (OTA) and deoxynivalenol (DON) are among the most widespread mycotoxins. We assessed the occurrence of AFT, OTA and DON in some foodstuffs available on the Lebanese market and evaluated the potential risk to the health of children and teenagers in Beirut from dietary exposure to these mycotoxins. Analytical data on the contamination of raw and processed cereals and cereal-based products, pulses and nuts were collected. The mean levels of AFT, OTA and DON were calculated for each food items. Levels of dietary exposure were obtained by combining food consumption data with the mean mycotoxin levels, adopting a deterministic approach. Intakes were calculated for average and high consumers (75th and 95th percentile) among children and teenagers, and compared with the provisional tolerable daily intake (PTDI) or provisional tolerable weekly intake (PTWI) of the respective mycotoxin. The results showed that mycotoxin levels in the food samples were generally below national/European maximum limits. However, high levels of AFT, OTA and DON were found in some samples of nuts, biscuits and bread. The calculated intake for AFT exceeded its respective PTDI in all groups by a factor ranging from 3 to 7. The intakes of OTA and DON were found to be below the threshold of toxicological concern established for these mycotoxins by international expert groups, although the intake of DON in children at the highest percentile (P95) was close to its PTDI. Our study highlights the need to reconsider the maximum permissible levels of total aflatoxins in some foodstuffs and to establish regulatory guidelines regarding OTA and DON. The results also emphasise the need for routine monitoring of the levels of mycotoxin contamination in foodstuffs that are highly consumed by Lebanese children and teenagers, i.e. cereals and cereal-based products.

Mycological survey and ochratoxin A natural contamination of swine feedstuffs in Rio de Janeiro State, Brazil

Mycotoxin contamination of animal feeds represents a hazard to human and animal health due to potential transmission to meat and milk. Barley by-products are alternative feeding supplies for animal production. The aims of this assay were to study the mycobiota of feedstuffs and finished swine feed, to determine the ability of Aspergillus and Penicillium isolates to produce ochratoxin A (OTA) and to evaluate OTA occurrence in these substrates. Corn, brewers' grains and finished swine feed samples were collected from different factories. Fungal counts were higher than 2.8 × 10 4 CFU g −1. Fusarium, Aspergillus and Penicillium genera were isolated at high levels. A 23.7% of the isolates produced 9–116 μg kg −1 of OTA in vitro. Corn samples (44%) were contaminated with 42–224 μg kg −1 of OTA. Finished feed (31%) and brewers' grains samples (13%) were contaminated with 36–120 μg kg −1 and 28–139 μg kg −1 of OTA, respectively. This is the first scientific report on contamination by OTA-producer molds and OTA in swine feedstuffs from Brazil. The presence of OTA in raw materials and finished feed requires periodic monitoring to prevent mycotoxicoses in animal production, reduce economic losses and minimize hazards to human health.

The involvement of mycotoxins in the development of endemic nephropathy

Endemic nephropathy is a human kidney disease that still escapes scientific explanation. It is accompanied by a high incidence of urothelial tumors in rural populations in endemic areas, which suggests that a natural nephrotoxic and carcinogenic compound may be involved in the etiology. The most imputed causative agent of endemic nephropathy is the mycotoxin ochratoxin A (OTA), because of its confirmed nephrotoxic and carcinogenic action. This paper presents a review of studies of OTA in food collected in the endemic areas and in blood and urine of their residents. Data on the co-occurrence of OTA and other nephrotoxic and carcinogenic mycotoxins such as citrinin and fumonisin B(1) in food are also presented. Unfortunately, there is no study on the co-occurrence of OTA and other mycotoxins in humans and there is only one study on fumonisin B(1) exposure in endemic areas. The paper also presents experimental data on cultured cells and laboratory animals treated with combinations of OTA and other nephrotoxic mycotoxins, because most such combinations show a synergistic effect. The occurrence of OTA- and aristolochic acid-DNA adducts is also presented.

Natural occurrence of ochratoxin A in Tunisian cereals

Ochratoxin A (OTA) is a mycotoxin produced by several fungal species from Aspergillus and Penicillium genera. It is widespread in food and feed and its occurrence has been reported in cereals, cereal-derived products, dried fruits and spices. This mycotoxin was implicated in several human and animal pathologies such as the Balkan Endemic Nephropathy (BEN) and the Tunisian Chronic Interstitial Nephropathy (CIN) of unknown cause. In Tunisia, a clear correlation has been established between the consumption of OTA contaminated food and the induction of specific pathologies. Thereby, OTA was detected in human blood and tissues. The aim of our study was to investigate the presence of OTA in widely consumed cereals commercialized in Tunisia. The analytical methods used in our study involved the extraction of OTA by acidified toluene, immunoaffinity (IAC) clean-up and HPLC quantification with fluorescence detection. Levels and percentages of OTA contamination in different types of cereals, 110 wheat, 103 barley, 113 sorghum and 96 rice samples, were evaluated with incidences of 38%, 40%, 38% and 28%, respectively. The average of contamination by OTA found were 55, 96, 44 and 117 μg/kg, respectively, for wheat, barley, rice and sorghum. Our results showed that contamination percentages and levels in the period from 2004 to 2005 were higher then usual norms (5.0 μg OTA/kg) established by the European commission in 2002. The present report is the first one ever carried out on the natural occurrence of OTA in cereals, largely consumed by the Tunisian population.

Ochratoxin A and citrinin producing species of the genus Penicillium from feedstuffs

In Spain, low ochratoxin A (OTA) levels have been detected in several pork products but there is no information published about the fungi involved in this OTA contamination. It is well known that P. verrucosum is much more frequently found on cereals in countries where they occasionally have OTA problems as in North European countries compared with South Europe where levels of OTA generally seem to be lower or not detected. Much less information is available about citrinin (CIT) and CIT producing species in cereals and their by products. The aim of this study was to determine, identify and characterize the occurrence of potential OTA and CIT producing Penicillium spp. from mixed feeds and raw materials purchased in the Spanish market and used as feedstuffs. A total of 155 Penicillium spp. isolates belonging to 34 species were analyzed in order to know if they are able to produce OTA and/or CIT. From these isolates, 11 P. verrucosum which were characterized by RAPD analyses, produced OTA. Fourteen isolates were CIT producers, 10 isolates of P. verrucosum and 4 of P. citrinum. Although the occurrence and abundance of OTA and CIT Penicillium producing species have been low in our study, our results confirm the potential risk of OTA and CIT production in feeds if stored improperly. Our results also confirm the occurrence of P. verrucosum in South European countries and that it is the only OTA producing Penicillium species in these substrates.

Maternal dietary habits and mycotoxin occurrence in human mature milk

During 2006, 82 samples of human mature milk were collected at Italian hospitals and checked for aflatoxin M1 (AFM1) and ochratoxin A (OTA) by immunoaffinity column extraction and HPLC. AFM1 was detected in four (5%) of milk samples (ranging from < 7 ng/L to 140 ng/L; mean level: 55.35 ng/L); OTA was detected in 61 (74%) of milk samples (ranging from < 5 ng/L to 405 ng/L; mean level: 30.43 ng/L. OTA levels were significantly higher (p less, not double equals 0.05) in milk of habitual consumers of bread, bakery products and cured pork meat. No other statistically significant differences were observed although habitual consumers of pasta (p = 0.059), cookies (p = 0.061) and juices (p = 0.063) had mean contamination values of OTA higher than the moderate consumer. The very few AFB1 positive samples did not allow statistical comparisons. The present study confirms that the occurrence of OTA in human milk is related to maternal dietary habits. The findings support the possibility of dietary recommendations to woman, during pregnancy and lactation, aimed to tentatively reduce the OTA contamination of human milk.

Mycobiota of Czech wine grapes and occurrence of ochratoxin A and Alternaria mycotoxins in fresh grape juice, must and wine.

The aim of this study was to monitor the mycobiota of wine grapes, occurrence of ochratoxigenic microfungi in wine grapes and occurrence of ochratoxin A and Alternaria mycotoxins in fresh grape juice, must and wine from domestic crops in the year 2004. Thirteen samples of wine grapes (white /nine samples/ and red /four samples/) were collected during harvesting in the Znojmo wine region, SE Moravia. One sample of a wine grape variety was represented by three subsamples of wine grapes, which were sampled in left, middle and right part of the vineyard. Five wine grape berries per bunch were randomly selected, plated onto Dichloran Rose Bengal Chloramphenicol (DRBC) agar, and incubated for 5–7 days at 25 °C. Alternaria alternata, Cladosporium herbarum, C. cladosporioides, Penicillium expansum, P. aurantiogriseum, P. spinulosum and Rhizopus nigricans were isolated from the samples. Ochratoxigenic microfungi, e. g. Aspergillus carbonarius, and other species of section Nigri, A. ochraceus, Penicillium verrucosum and P. nordicum, were not found in the samples. The HPTLC method for quantification of ochratoxin A (OTA) and Alternaria mycotoxins (alternariol (AOH), alternariol monomethyl ether (AME), altenuene (ALT), and tenuazonic acid (TeA)) in fresh grape juice (13 samples), must (13 samples) and wine (13 samples) was used. The limit of quantification (LoQ) was 8 ng/l for OTA, 1.5 μg/l for AOH, 1.5 μg/l for AME, 1.5 μg/l for ALT and 7.5 μg/l for TeA. Occurrence of OTA and Alternaria mycotoxins in fresh grape juice, must and wine was not proved.

Survey: Ochratoxin A in European special wines

The occurrence of Ochratoxin A (OTA) was examined in 121 special wines made using different winemaking techniques and from many European origins. The wine groups with the highest OTA content and occurrence, above 90%, were those were the must was fortified before fermentation (mean: 4.48μg/l) and those made from grapes dried by means of sun exposure (mean: 2.77μg/l). Fortified wines with long aging in wooden casks were about 50% contaminated, with OTA levels below 1.00μg/l. Wines affected by noble rot, late harvest wines and ice wines did not contain OTA. Overall, 19.8% of the wines studied contained OTA levels above the maximum permissible limit for the European Union (2μg/kg) in wine (excluding liqueur wines).

An overview of ochratoxin A in beer and wine

Ochratoxin A (OTA) is a mycotoxin produced mainly by several fungal species of the genera Aspergillus and Penicillium. This mycotoxin has been shown to be nephrotoxic, hepatotoxic, teratogenic and carcinogenic to animals and has been classified as a possible carcinogen to humans. OTA occurs in a variety of foods, including beer and wine. Reports on OTA occurrence in beer indicate that this is a worldwide problem due to the widespread consumption of this beverage. At present, the European Union (EU) has not set a maximum allowable limit (MAL) for this mycotoxin in beer, although there is a limit in barley and malt. Studies carried out in different countries agree in the high proportion of samples contaminated with OTA although levels are, usually, below 0.2 ng/ml. OTA occurrence has been related to the contamination of malt barley with ochratoxigenic species, particularly Penicillium verrucosum. OTA produced in grains is carried to wort and, although fermentation decreases the concentration, the toxin is not eliminated. Reducing the fungal contamination of malt barley is the most promising strategy for reducing OTA in beer. With regard to wine, surveys on the presence of OTA have been conducted worldwide. The proportion of wines in which OTA is detected is very high (above 50%) in some countries (especially in the Mediterranean basin) although only a few wines contained concentrations exceeding the MAL laid down by the EU (2.0 ng/ml). A gradient of concentration is usually recognized; OTA levels decrease in the order red, rosé, and white wine but also with increasing latitude of the producing countries. OTA presence in wines is due to the black aspergilli, mainly A. carbonarius, which can grow on grapes in the vineyards and produce the toxin. At grape crushing, the juice can be contaminated with the toxin which is carried over into wine, where it persists due to its stability. Pre- and post-harvest treatments are being investigated to diminish contamination of wines as much as possible.

Occurrence of ochratoxin A in bread consumed in Morocco

One hundred samples of commercial bread purchased from January to October (2006) from retail bakeshops in five different cities (Rabat, Témara, Salé, Casablanca and Meknès) in Morocco were surveyed for the presence of ochratoxin A (OTA) using liquid chromatography coupled to fluorescence detection. The identification of OTA in positive bread samples was confirmed by methyl ester derivatization. Analytical results showed that forty eight (48%) samples were positive with OTA greater than the limit of quantification (LOQ = 0.051 ng/g). Levels of OTA in positive samples ranged between 0.14 and 149 ng/g. The average contamination of bread samples with OTA was 13 ± 1.5 ng/g. The highest frequency of positive samples (61.5%) and the most contaminated sample (149 ng/g) were found in bread commercialized in the Casablanca area. Twenty six of the positive samples exceeded the maximum level of 3 ng/g set by European regulations for OTA in cereals and derivatives. Based in the results presented in this study, the estimated daily intake of OTA from bread was 126 ng/kg bw/day. The present paper is the first ever drafted on the natural occurrence of OTA in bread consumed in Morocco. Data on the daily intake of OTA by the Moroccan population are also estimated for the first time.

Ochratoxin A in brazilian instant coffee

The aim of this study was to determine the ochratoxin A (OTA) contamination of instant coffee samples collected in the market of the city of São Paulo, Brazil from August to December, 2004. The EN 14133/2003 method, originally developed to quantify OTA in wine, grape juice and beer samples, was evaluated and approved for analyzing OTA in instant coffee samples. OTA was isolated in an immunoaffinity column and quantified by HPLC with fluorescence detection. The established detection and quantification limits were 0.16 and 0.52 ng/g, respectively. The recoveries from spiked samples were 92.6 ± 1.7, 83.7 ± 0.8, and 91.0 ± 1.2 % at levels of 3.0, 5.0, and 8.0 ng/g, respectively. Of a total of 82 samples analised, 81 (98.8%) contained OTA at levels ranging from 0.17 to 6.29 ng/g. The high frequency of OTA occurrence in the instant coffee samples demonstrates the importance of an effective control of this product by governmental authorities and industries. The rapid methodology for OTA analysis in instant coffee used in this study was defined and validated, permitting it´s use for quality control of this product.

Occurrence of ochratoxin A in Turkish wines

A total of 95 wine samples including 34 white, 10 rosé and 51 red wines originating from four different Turkish areas were analysed for ochratoxin A (OTA). An analytical method based on immunoaffinity column (IAC) for clean-up and high performance liquid chromatography with fluorescence detection (HPLC-FD) was used to determine OTA in wines. The limit of detection (LOD) was estimated as 0.006 ng ml − 1 for white wine and 0.010 ng ml − 1 for rosé and red wines. The limit of quantification (LOQ) was estimated as 0.020 ng ml − 1 in white wine and 0.030 ng ml − 1 in rosé and red wines. Recovery experiments were carried out with spiked samples in the range 0.1–1 ng ml − 1 of OTA. The average OTA recoveries from spiked white wine samples varied from 79.43% to 85.07%; while the mean recoveries for rosé and red wine samples were in the range of 77.48–83.96% and 76.61–83.55%, respectively. OTA was detected in 82 (86%) wine samples at levels of < 0.006–0.815 ng ml − 1 , which were below the maximum allowable limit established by the European Community. The mean OTA concentration in red wines was slightly higher than in white and rosé wines. Furthermore, our data indicate that the geographic region of origin has strong influence on OTA level for white, rosé and red wines: wines originating from Thrace ( n = 44, mean = 0.158 ng ml − 1 ) and Aegean ( n = 28, mean = 0.060 ng ml − 1 ) regions of Turkey were more contaminated with OTA compared with wines originating from central ( n = 15, mean = 0.027 ng ml −1) and east Anatolia ( n = 8, mean = 0.027 ng ml − 1 ) areas. This study showed that the occurrence of OTA in Turkish wines is high, but at levels that probably leads to a non-significant human exposure to OTA by consumption of wines.

Incidence of ochratoxin A in rice and dried fruits from Rabat and Salé area, Morocco

One hundred samples of dried fruits (20 dried raisins, 20 walnuts, 20 peanuts, 20 dried figs and 20 pistachios) and 20 samples of rice purchased from retail shops in the Rabat and Salé area in Morocco were analysed for ochratoxin A (OTA) by immunoaffinity clean-up (IAC) and liquid chromatography (LC) with fluorescence detection. The limit of quantification (LOQ) (S/N = 10:1) of OTA was 0.02 ng g−1 in rice, 0.03 ng g−1 in pistachio, peanut and walnut, and 0.03 ng g−1 in dried raisins and dried figs. The incidences of occurrence of OTA in dried raisins, walnuts, peanuts, dried figs and rice were 30, 35, 25, 65 and 90%, respectively. Analytical results showed that pistachio samples contained no detectable OTA, but concentrations ranged from 0.02 ± 0.01 to 32.4 ± 2.10 ng g−1 in rice, from 0.10 ± 0.05 to 2.36 ± 0.75 in peanut, from 0.03 ± 0.01 to 1.42 ± 0.45 in dried figs, from 0.05 ± 0.02 to 4.95 ± 0.02 in dried raisins, and from 0.04 ± 0.01 to 0.23 ± 0.05 in walnuts. The results also showed that 15% of the total number of rice samples analysed exceeded the 2002 regulatory limit set by European Union regulations for cereals. This is the first report on the occurrence of OTA in dried fruits and rice available in Morocco.

Ochratoxin A and the occurrence of ochratoxin A‐producing black aspergilli in stored peanut seeds from Córdoba, Argentina

AbstractSince there is no available information about the natural occurrence of ochratoxin‐producing fungi and ochratoxin A (OTA) in peanut seeds in Argentina, the aims of this work were to isolate and identify Aspergillus section Nigri and to evaluate the natural occurrence of OTA in stored peanut seeds. Likewise, the capacity to produce OTA by Aspergillus section Nigri was studied. Forty‐seven samples of peanut seeds were obtained from a storage plant in the south of Córdoba Province, Argentina. Each sample of 100 seeds was surface‐disinfected and plated onto a dichloran 18% glycerol agar (DG18). OTA was detected by HPLC. The production of OTA in strains belonging to section Nigri was tested in YES medium (2% yeast extract, 15% sucrose). Aspergillus spp., the most frequent mould, occurred in 100% of samples, followed by Penicillium spp. (87%) and Eurotium spp. (6.4%). A. flavus was isolated in 100% of the samples, followed by A. niger var. niger, A. niger var. awamori and A. carbonarius at a lower frequency, 89%, 68% and 4%, respectively. OTA was found in 32% of the peanut seed samples, with mean levels ranging from 0.5 to 170 ng g−1. The mean recovery of the method used was 85 ± 2%. Forty‐three isolates (27%) of Aspergillus section Nigri, were found to be OTA producing strains. The highest percentage of ochratoxigenic strains (57%) was found within the A. carbonarius ssp. These results indicate a human exposure to OTA in Argentina through the ingestion of peanut seeds and peanut products. Copyright © 2006 Society of Chemical Industry

Natural Occurrence of Mycotoxins in Staple Cereals from Ethiopia

The occurrence of mycotoxins in barley, sorghum, teff (Eragrostis tef) and wheat from Ethiopia has been studied. Samples were analyzed for aflatoxin B(1) (AFB1), ochratoxin A (OTA), deoxynivalenol (DON), nivalenol (NIV) and zearalenone (ZEN) using high performance liquid chromatography (HPLC) and for fumonisins (FUM) using enzyme linked immunosorbent assay (ELISA). AFB1 and OTA were detected in samples of all the four crops. AFB1 was detected in 8.8% of the 352 samples analyzed at concentrations ranging from trace to 26 microg kg(-1). OTA occurred in 24.3% of 321 samples at a mean concentration of 54.1 microg kg(-1) and a maximum of 2106 microg kg(-1). DON occurred in barley, sorghum and wheat at 40-2340 microg kg(-1) with an overall incidence of 48.8% among the 84 mainly 'suspect' samples analyzed; NIV was co-analyzed with DON and was detected at 40 microg kg(-1) in a wheat sample and at 50, 380, and 490 microg kg(-1) in three sorghum samples. FUM and ZEN occurred only in sorghum samples with low frequencies at concentrations reaching 2117 and 32 microg kg(-1), respectively. The analytical results indicate higher mycotoxin contamination in sorghum, which could be related to the widespread storage of sorghum grain in underground pits leading to elevated seed moisture contents. This is the first report on the occurrence of OTA in teff.

Occurrence of ochratoxin A in wines in the Argentinean and Chilean markets

Wine contaminated with ochratoxin A (OTA) has been reported all over the world. Sixty-eight wine samples were analysed to assess OTA wine contamination in various regions of Argentina and Chile. In addition, some imported wines were analysed. Wine samples were collected at manufacturers' stock and retail markets in Argentina in 2003 and Chile in 2002. A high- performance liquid chromatographic method with fluorescence detection and two different immunoaffinity clean-up columns were employed, with recoveries higher than 90% (Argentina: LOD:0.008 µg/l, LOQ:0.015 µg/l; Chile: LOD:0.012 µg/l, LOQ: 0.04 µg/l). None of the analysed wines produced in Argentina or Chile were contaminated. The presence of OTA in wines would appear to be a lesser problem in Argentina and Chile than in other countries, but it still could contribute to OTA exposure of human populations and more studies of the occurrence of OTA in wine should be done.

Ochratoxin A in sultanas from Turkey I: Survey of unprocessed sultanas from vineyards and packing-houses

A method for the determination of ochratoxin A (OTA) in sultanas from Turkey using extraction with a sodium bicarbonate solution (2% NaHCO3) followed by immunoaffinity clean-up and liquid chromatography with fluorescence detection was used to assess the frequency of occurrence and level of OTA. In-house validation was carried out with spiked samples at levels of 0.15, 1.5, 5.0 and 10 µg kg−1 and average recoveries were 91, 93, 87 and 89%, respectively. The limits of detection and limit of quantification in Turkish sultanas were 0.026 and 0.09 µg kg−1, respectively. A survey for the presence of OTA was carried out on 264 unprocessed sultana samples during the production seasons between 1998 and 2000 collected annually from vineyards and from packing-houses. The analyses of unprocessed sultanas showed that 32.2% of the total number of samples contained no detectable OTA, whereas 9.8% of sultana samples had OTA concentrations above 10 µg kg−1, and the remaining 58% had levels within the range 0.026–10 µg kg−1. There were big differences in median concentrations between years. Considering the year of production, it appears that sultanas produced in 1998 and 2000 showed the lowest incidence of OTA contamination (median < 0.02 µg kg−1), whereas 2002 showed the highest incidence (median = 4.3 µg kg−1). The overall mean OTA concentration was calculated as 3.4 µg kg−1, and the overall median as 0.9 µg kg−1. Among the samples analysed, the highest detected level of OTA was 54 µg kg−1.

Occurrence of ochratoxin A in cocoa beans: Effect of shelling

The aim of this study was to investigate the influence of the shelling process on the presence of ochratoxin A (OTA) in cocoa samples. Twenty-two cocoa samples were analysed for the determination of OTA before (cocoa bean) and after undergoing manual shelling process (cocoa nib). In order to determine OTA contamination in cocoa samples, a validated high-performance liquid chromatography (HPLC) method with fluorescence detection was used for the quantitative analysis of ochratoxin A (OTA). In both types of samples, OTA was extracted with methanol–3% sodium hydrogen carbonate solution and then purified using immunoaffinity columns prior to HPLC analysis. Due to the fact that different recovery values were obtained for OTA from both types of samples, a revalidation of the method in the case of cocoa nibs was needed. Revalidation was based on the following criteria: Selectivity, limits of detection and quantification (0.03 and 0.1 µg kg−1, respectively), precision (within-day and between-day variability) and recovery 84.2% (RSD = 7.1%), and uncertainty (30%). Fourteen of the twenty-two cocoa bean samples (64%) suffered a loss of OTA of more than 95% due to shelling, six samples suffered a loss of OTA in the range 65–95%, and only one sample presented a reduction of less than 50%. The principal conclusion derived from this study is that OTA contamination in cocoa beans is concentrated in the shell; therefore, improvements of the industrial shelling process could prevent OTA occurrence in cocoa final products.