Five-year survey of ochratoxin A in processed sultanas from Turkey

The results of surveillance for ochratoxin A (OTA) in 1885 samples of sultanas taken during five crop years between 1999 and 2003 are reported. The analytical method was based on extraction with methanol + sodium bicarbonate and clean-up by immunoaffinity column chromatography followed by high-performance liquid chromatography with fluorescence detection. The limit of detection for OTA was 0.3 µg kg^-1. The results show that 9.3% of the samples contained no detectable levels of OTA, whereas 0.6% had concentrations exceeding 10 µg kg^-1; the remaining 90.3% had levels within the range 0.3-10 µg kg^-1. The overall mean OTA concentration in the total number of 1885 samples taken was 1.36 ± 2.91 µg kg^-1; the overall median was calculated as 0.90 µg kg^-1.     

Ochratoxin A in dried vine fruits on the Canadian retail market

Between 1998 and 2000, 151 samples of raisins and sultanas and two samples of currants were collected from retail outlets across Canada and analysed for ochratoxin A. Samples were extracted with methanol-sodium bicarbonate, and the extracts were cleaned-up by immunoaffinity column chromatography. Ochratoxin A was quantified by liquid chromatography with fluorescence detection. The minimum quantifiable level was 0.1 ng g^-1. Ochratoxin A was present, above the minimum quantifiable level, in 67 (79%) of 85 samples of raisins, in 39 (59%) of 66 samples of sultanas, and in both samples of currants. The overall mean level of ochratoxin A was 1.8 ng g^-1 in both the raisins and sultanas, and 2.8 ng g^-1 in the currants.     

Ochratoxin A in dried vine fruit: method development and survey

A method is described for the determination of concentrations of the mycotoxin ochratoxin A in dried vine fruits (currants, raisins and sultanas) using acidic methanolic extraction,immunoaffinity chromatography clean-up and HPLC determination. The limit of detection was estimated as 0.2mug/kg, and recoveries of 63-77% were achieved at 5mug/kg. HPLC-mass spectrometric confirmation of the identity of ochratoxin was obtained. Ochratoxin A and aflatoxins were determined in 60 samples of retail dried vine fruits purchased in the United Kingdom. Ochratoxin A was found in excess of 0.2mug/kg in 19 of 20 currant, 17 of 20 sultana and 17 of 20 raisin samples examined, an overall incidence of 88% . The maximum level found was 53.6mug/kg. No aflatoxin was found in any sample analysed, using a method with a detection limit of 0.2mug/kg for each of aflatoxin B1, B2, G1 and G2.     

Fumonisin contamination and fumonisin producing black Aspergilli in dried vine fruits of different origin

Aspergillus niger isolates are able to produce fumonisins in high quantities on agar media with a low water activity. Several agricultural products fit this criterion, including dried vine fruits, dates and figs. Data on the occurrence and role of this species in fumonisin contamination of agricultural products with high sugar content are needed to clarify the importance of A. niger in human health. The mycobiota and fumonisin contamination of various dried vine fruit samples collected from different countries were examined to clarify the role of black Aspergilli in fumonisin contamination of such products. All except two of the examined samples were contaminated with black Aspergilli. Species assignment of the isolates was carried out using sequence analysis of part of the calmodulin gene. The range of fumonisin isomers present in the raisin samples, and produced by A. niger isolates collected from dried vine fruits was also examined using reversed-phase high-performance liquid chromatography/electrospray ionization-ion trap mass spectrometry (RP-HPLC/ESI-ITMS). Among the 30 A. niger/A. awamori isolates identified, 20 were found to be able to produce fumonisins (average contamination: 5.16 mg/kg; range: 0.017-19.6 mg/kg). The average fumonisin content of the 7 dried vine fruit samples which were found to be contaminated by potential fumonisin producing black Aspergilli was 7.22 mg/kg (range: 4.55-35.49 mg/kg). The isolates produced several fumonisin isomers also present in the dried vine fruit samples, including fumonisins B_1-4, 3-epi-FB₃, 3-epi-FB₄, iso-FB₁, and two iso-FB_2,3 forms. Fumonisin B₁ was detected for the first time in A. niger cultures. Most of these isomers have previously only been identified in Fusarium species. Our data indicate that A. niger and A. awamori are responsible for fumonisin contamination of dried vine fruits worldwide. The observed levels of contamination are alarming and pose a new threat for food safety.     

High-performance liquid chromatography-tandem mass spectrometry method for simultaneous detection of ochratoxin A and relative metabolites in Aspergillus species and dried vine fruits

A simple, sensitive and reliable quantification and identification method was developed for simultaneous analysis of ochratoxin A (OTA) and its related metabolites ochratoxin alpha (OTα), ochratoxin B (OTB) and mellein. The method was assessed by spiking analytes into blank culture media and dried vine fruits. Performance was tested in terms of accuracy, selectivity and repeatability. The method involves an ultrasonic extraction step for culture samples using methanol aqueous solution (7:3, v/v); the mycotoxin is quantified by high-performance liquid chromatography coupled with electrospray ionisation and triple quadrupole mass spectrometry (LC-ESI-MS/MS). The recoveries were 74.5–108.8%, with relative standard deviations (RSDs) of 0.4–8.4% for fungal culture. The limits of detection (LODs) were in the range of 0.03–0.87 μg l^–1, and the limits of quantification (LOQs) ranged from 0.07 to 2.90 μg l^–1. In addition, the extraction solutions and clean-up columns were optimised specifically for dried vine fruit samples to improve the performance of the method. Methanol–1% sodium bicarbonate extraction solution (6:4, v/v) was determined to be the most efficient. Solid-phase extraction (SPE) was performed as a clean-up step prior to HPLC-MS/MS analysis to reduce matrix effects. Recoveries ranged from 80.1% to 110.8%. RSDs ranged from 0.1% to 3.6%. LODs and LOQs ranged from 0.06 to 0.40 μg kg^–1 and from 0.19 to 1.20 μg kg^–1, respectively. The analytical method was established and used to identify and quantify OTA and related compounds from Aspergillus carbonarius and Aspergillus ochraceus in cultures and dried vine fruits. The results showed that A. carbonarius produced OTα, OTB and OTA, whereas A. ochraceus produced OTB, OTA and mellein after 7 days of cultivation. Of 30 commercial samples analysed, 10 were contaminated with ochratoxins; OTB, OTα and mellein were also detected in different samples.     

Ochratoxin A concentrations in Greek domestic wines and dried vine fruits.

A survey for the presence of ochratoxin A (OTA) was conducted from 1995 to 1999 on 268 locally produced commercial wines, and on 81 samples of domestic dried vine fruits (currants and sultanas) collected between 1998 and 2000 from sites of primary storage and processing. The OTA concentration in red dry wines (n = 104, median = 0.09 microgram l(-1)) was not significantly different from that for white (n = 118, median = 0.06 microgram l(-1)) and rosé (n = 20, median = 0.08 microgram l(-1)) wines. Eighteen samples of dessert wines (sweet, semi-sweet, semi-dry) and eight samples of retsina wine showed larger OTA concentrations with medians of 0.33 and 0.27 microgram l(-1), respectively. Our data indicate that the geographic region of origin influences OTA contamination for the red dry wines. In fact, a trend of increasing OTA contamination was observed for red wines from northern to southern Greece. Regarding the OTA levels in dried vine fruits, sultanas (n = 27, median = 0.6 microgram kg(-1)) were less contaminated than currants (n = 54, median = 1.3 microgram kg(-1)). Also, sultanas produced in 2000 and currants produced in 1999 showed the lowest incidence of OTA contamination, with medians of 0.3 and 0.9 microgram kg(-1), respectively.     

Mycotoxins in botanicals and dried fruits: A review

Botanicals are used in many countries for medicinal and general health-promoting purposes. Numerous natural occurrences of mycotoxins in botanicals and dried fruits have been reported. Aflatoxins or ochratoxin A (OTA) have been found in botanicals such as ginseng, ginger, liquorice, turmeric, and kava-kava in the USA, Spain, Argentina, India, and some other countries, while fumonisins have been found in medicinal wild plants in South Africa and in herbal tea and medicinal plants in Turkey. Zearalenone was identified in ginseng root. Dried fruits can be contaminated with aflatoxins, OTA, kojic acid, and, occasionally, with patulin or zearalenone. One main area of concern is aflatoxins in dried figs; bright greenish yellow fluorescence under ultraviolet light is associated with aflatoxin contamination. OTA in dried vine fruits (raisins, sultanas, and currants) is another concern. There are also reports of aflatoxins in raisins and OTA in dried figs, apricots, dried plums (prunes), dates, and quince. Maximum permitted levels in the European Union include 4 µg kg^-1 for total aflatoxins in dried fruit intended for direct consumption and 10 µg kg^-1 for OTA in dried vine fruit. This review discusses the occurrence of mycotoxins in botanicals and dried fruits and analytical issues such as sampling, sample preparation, and methods for analysis. Fungal contamination of these products, the influence of sorting, storage, and processing, and prevention are also considered.     

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.     

Development and validation of a solid-phase extraction method coupled with HPLC-UV detection for the determination of biogenic amines in Chinese rice wine

ABSTRACT

A reliable and accurate method for the determination of seven biogenic amines (BAs) was developed and validated with Chinese rice wine samples. The BAs were derivatised with dansyl chloride, cleaned up using solid-phase extraction (SPE) and separated by high-performance liquid chromatography (HPLC) coupled with ultraviolet (UV) detection. The optimised derivatisation reaction, conducted at pH 9.6 and 60°C for 30 min, ensured baseline separation and peak symmetry for each BA. SPE clean-up using Oasis MCX cartridges yielded good recovery rates for all BAs and effectively reduced matrix effects. The developed method shows good linearity with determination coefficients of more than 0.9989 over a concentration range of 0.1–100 mg l^?1. The limits of detection (LODs) for the investigated BAs ranged from 2.07 to 5.56 µg l^?1. The intra- and inter-day relative standard deviations (RSDs) ranged from 0.86% to 3.81% and from 2.13% to 3.82%, respectively. Spiking experiments showed that the overall recovery rates ranged from 85% to 113%. Thus, the proposed method was demonstrated as being suitable for simultaneous detection, with accurate and precise quantification, of BAs in Chinese rice wine.     

Detection and quantification of Ochratoxin A in milk produced in organic farms

Dairy cows (as all other ruminants) possess physiological systems for mycotoxins’ detoxification. However, in organic farming practices the detoxification could be impaired because of possible higher contamination of feedstuff, changes in rumen pH and other factors. So the organic milk could be at risk in this respect. The results of an investigation on the presence of Ochratoxin A (OTA) in 63 samples of organic and 20 samples of conventional milk are reported in this paper. The quantification was carried out by means of the HPLC method described by Sørensen and Elbœk (2005). The method has been modified in the purification step so as to shorten the time of analysis and lower the cost of the assay.     

Ochratoxin A in beverages from Morocco: a preliminary survey

In a preliminary study, samples of Moroccan wines (n = 30), beers (n = 5) and fruit juices (n = 14) were assayed for ochratoxin A (OTA) by HPLC with fluorimetric detection, followed by confirmation by cleavage of the OTA molecule using carboxypeptidase with HPLC-fluorimetric determination of ochratoxin alpha (OT alpha). All the wine samples were contaminated, and the overall median OTA concentration was 0.65 microg/l (range 0.028-3.24 microg/l). One of the 14 samples of fruit juices was contaminated with a concentration of 1.16 microg/l, whereas none of the five beer samples was contaminated. This is the first report on the occurrence of OTA in various beverages from Morocco.     

Incidence of toxigenic fungi and ochratoxin A in dried fruits sold in Brazil

A total of 117 dried fruit samples (black sultanas, white sultanas, dates, dried plums, dried figs and apricots) from different origins were analysed both for toxigenic fungi and for the presence of ochratoxin A. Amongst the fungi found, Aspergillus niger was predominant, with 406 isolates, of which 15% were ochratoxin A producers. They were followed by A. ochraceus, with 15 isolates and 87% ochratoxigenics, and A. carbonarius, with only five isolates of which 60% were ochratoxin A producers. The average infection rates for A. niger in black sultanas, plums, figs, dates and white sultanas were 22.0, 8.0, 4.0, 1.5 and 0.5%, respectively. The apricot samples were not contaminated by any fungi or ochratoxin A. Black sultana and dried figs contained the highest contamination with ochratoxin A, with 33 and 26.3% of the samples containing more than 5 µg kg^-1 respectively, while all the white sultanas, dates and plums had no sample that exceeded this limit.     

高效液相色谱法测定糕点中5种食品添加剂

建立糕点中常用的3种防腐剂和2种甜味剂的高效液相色谱(HPLC)分析方法。样品以中性氧化铝作为分散基体,研磨均匀后用甲醇—水(70∶30,体积比)进行超声波提取I,nertsil ODS-sp C18柱(4.6 mm×250 mm,5μm)分离。流动相A为含2.84 g/L硫酸钠+2.60 g/L溴化四丁基铵的水溶液,流动相B为甲醇(A∶B=40∶60,体积比),等度洗脱,二极管阵列检测器于波长220 nm和230 nm处检测,利用保留时间和光谱图定性,外标法定量。结果表明,5种物质在10 min内分离完全,在1μg/mL～200μg/mL范围内线性关系良好,相关系数均大于0.999 5,检出限(LOD)为0.016 mg/kg～0.039 mg/kg(信噪比S/N=3)。在20.0 mg/kg～120.0 mg/kg添加水平下,平均回收率为89.5%～101.6%,相对标准偏差RSD为0.98%～3.07%(n=4)。     

Ochratoxin A in Brazilian roasted and instant coffees

Thirty-four samples of roast and ground coffee, 14 samples of instant coffee and two samples of decaffeinated instant coffee were collected in markets and supermarkets in the city of Campinas, Brazil, and analysed for ochratoxin A using immunoaffinity columns for clean-up and HPLC with fluorescence detection for quantification. The limit of detection was 0.2 ng/g ochratoxin A. Twenty-three samples of ground and roast coffee were found to be contaminated with the toxin at levels ranging between 0.3 and 6.5 ng/g. The average concentration in all 34 samples was 0.9 ng/g. All samples of instant coffee contained ochratoxin A at levels ranging from 0.5 to 5.1 ng/g, with an average figure of 2.2 ng/g. Roast and ground coffee is the type of coffee most used by Brazilians for the preparation of the beverage. Considering that an average Brazilian adult takes five cups of coffee per day, which corresponds to 30 g of roast and ground coffee, the probable daily intake of ochratoxin A by a 70 kg adult would be 0.4 ng/kg bw, which is far below the current Provisional Tolerable Daily Intake of 14 ng/kg bw for ochratoxin A as set by the Codex Alimentarius. To study the transfer of ochratoxin A into coffee brew, the beverage was prepared by two methods: (a) the drip method and (b) the Brazilian country style method. No significant difference was observed between the two methods in terms of extraction of the toxin using five contaminated samples containing between 0.8 and 6.5 ng/g ochratoxin A. The drip method extracted 86 +/- 15% and the Brazilian country style 74 +/- 20% of the ochratoxin A initially present in the roast and ground coffee.     

Limited survey for the presence of aflatoxins in foods from local markets and supermarkets in Valencia, Spain

Aflatoxin B1 (AFB1), B2 (AFB2), G1 (AFG1) and G2 (AFG2) were extracted by matrix solid-phase dispersion with C18 silica and acetonitrile as the eluting solvent, analysed by liquid chromatography with fluorescence detection and confirmed by liquid chromatography with mass spectrometry using an electrospray interface in 58 samples grouped as cereals, dried fruits, herbs and spices, pulses, snacks, and nuts and nut products collected from local markets and supermarkets in Valencia, Spain. All samples analysed by the proposed method were previously studied with an enzyme-linked immunosorbent assay as a screening protocol for the fast detection of mycotoxins. The samples containing residues (3/58) were hazelnut (0.42 and 0.52 μg kg^-1 for AFB1 and AFG1, respectively), nut cocktail (0.29 and 0.47 μg kg^-1 for AFB1 and AFG1, respectively) and pinhol (0.30 μg kg^-1 for AFG1). Such values were below the legislated maximum residue levels for the European Union.     

Mode of action of the drying emulsion used in dried vine fruit production. II. The effect of emulsion pH

Alkaline pH is a critical component of the drying emulsion used in Australia to enhance the drying rate of Sultana grapes. Weight loss of grapes treated with 0.18M K₂CO₃ solution with or without 2% commercial drying oil increased approximately linearly from pH 4 to 14. Minimal weight loss after 24 hours of drying occurred between pH 2 and pH 6, in both the presence and absence of commercial drying oil. It was higher at pH 2 than at pH 4, while at pH values above 4 the weight loss became progressively greater, indicating an isoelectric point in the pH range 2 to 6. Alkali metal carbonate solutions of high pH, e.g. 11.5, were required for maximum drying rates, with effectiveness of the alkali metal ions increasing with decreasing hydrated ion radius, i.e. following the lyotropic series. Cation exchange capacity was also higher at high pH. The cuticle reacted comparably to a highly cross-linked cation exchanger of the weakly acidic type, and exhibited similarities to leaf and fruit cuticles of other species in its response to pH. The light colour of the dried sultanas was highly correlated with the rate of weight loss.     

Ochratoxin A in grape pekmez (grape molasses) consumed in Turkey

In this study, ochratoxin A (OTA) in 55 home-made, 20 commercial and 7 organic grape pekmez (grape molasses) produced in Turkey was investigated. OTA was detected in 73% of home-made pekmez samples, in 35% of commercial pekmez samples and in 71% of organic pekmez samples. Eleven per cent of the samples had OTA levels higher than 10 µg/l. The highest OTA level (31 µg/l) was detected in organic pekmez. The maximum OTA levels were 15 µg/l and 12 µg/l in home-made and commercial pekmez samples, respectively. Mean OTA levels were 3.5 µg/l, 1.4 µg/l and 9.2 µg/l in home-made, commercial and organic pekmez samples, respectively. Organic pekmez samples and home-made pekmez samples had higher OTA contamination than commercial pekmez samples. Results confirm OTA contamination in grape pekmez samples, indicating that the OTA level in grape pekmez could be a potential risk for consumers.     

进口葡萄酒中赭曲霉毒素A的含量分析

为了解进口葡萄酒中赭曲霉毒素A的含量和污染状况,本研究建立了无需净化、浓缩,直接进样,采用液相色谱-串联质谱联用法(LC-MS/MS)测定葡萄酒中赭曲霉毒素A的方法。赭曲霉毒素A空白基质标准溶液在1.0～20.0 ng/mL浓度范围内线性良好,方法检测限达到0.05 ng/mL,添加浓度为2.0 ng/mL质控样品的回收率为98.7%～113.2%,精密度<5.7%。应用该方法测定了84个进口葡萄酒样品中赭曲霉毒素A的含量,结果表明,赭曲霉毒素的检出率为100%,含量为0.14～1.10 ng/mL,平均含量为0.32 ng/mL。     

葡萄酒中赭曲霉素A的研究进展

赭曲霉素A（OTA）是一种真菌毒素,常见于谷物、谷物产品、水果、咖啡、葡萄干、葡萄酒和啤酒中。由于OTA在食物中稳定且不易分解,对人体构成了一种潜在的危害,国际癌症研究机构（IARC）已将其定义为group2B类致癌物。欧洲各国已经对赭曲霉素A进行限量,也制定了相应的官方检测方法。对葡萄酒中赭曲霉素A的性质、毒性、污染现状、产生菌、影响因素和测定方法进行了综述。     

Ochratoxin A in liquorice as affected by processing methods

A study of the effect of several processing methods on the concentration of ochratoxin A (OTA) in liquorice and derived products was carried out. The effect of the sorting, washing and peeling of fresh liquorice roots was investigated; as well as the production at a laboratory scale of liquorice extract and block liquorice from dry roots. Finally, the thermal stability of OTA was assessed. The OTA content was analysed by high-performance liquid chromatography-fluorescence and confirmed by methyl ester formation. The OTA level in liquorice extract was stable to heat treatment at 150°C for 60 min. The OTA concentration was unaffected by sorting or washing, but it was much reduced by peeling (a 53.1% reduction). A great reduction in the OTA level was found during the production of liquorice extract (78.6%) and block liquorice (91.8%).