Determination of mycotoxins in cereals by liquid chromatography tandem mass spectrometry

A liquid chromatography tandem mass spectrometry (LC–MS/MS) method is described for simultaneous determination of aflatoxins (AFB₁, AFB₂, AFG₁ and AFG₂), ochratoxin A (OTA), zearalenone (ZEA), deoxynivalenol (DON), fumonisins (FB₁ and FB₂), T2 and HT2-toxin in cereals. One-step extraction using solvent mixtures of acetonitrile:water:acetic acid (79:20:1) without any clean-up was employed for extraction of these mycotoxins from cereals. The mean recoveries of mycotoxins in spiked cereals ranged from 76.8% to 108.4%. Limits of detection (LOD) and quantification (LOQ) ranged 0.01–20 and 0.02–40 ng/g, respectively. The developed method has been applied for the determination of mycotoxins in 100 cereal samples collected from Malaysian markets. A total of 77 cereal samples (77%) contaminated with at least one of these mycotoxins. Occurrence of mycotoxins in commercial cereal samples were 70%, 40%, 25%, 36%, 19%, 13%, 16, and 16% for aflatoxins, OTA, ZEA, DON, FB₁, FB₂, T2 and HT2-toxin, respectively. The results demonstrated that the procedure was suitable for the determination of mycotoxins in cereals and could be implemented for the routine analysis.     

Determination of aflatoxins B1 and B2 by adsorptive cathodic stripping voltammetry in groundnut

In this study, adsorptive stripping voltammetry was proposed for determination of aflatoxins B1 (AFB1) and B2 (AFB2) using hanging mercury drop electrode (HMDE) as the working electrode. Both aflatoxins were found to adsorb and undergo irreversible reduction reaction at the working mercury electrode. The experimental conditions were optimised by one-at-a time and experimental design to obtain the best characterised peak in terms of peak height with analytical validation of the method for each aflatoxin. The calibration curves for aflatoxins AFB1 and AFB2 were linear in the ranges of 0.4–40 ng ml⁻¹ and 0.2–70 ng ml⁻¹ with the limit of detections (LOD) 0.15 and 0.10 ng ml⁻¹, respectively. The proposed method was applied for the analysis of aflatoxins in groundnut samples and the results were compared with those obtained by the HPLC technique.     

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.     

Determination of aflatoxins in eggs,milk, meat and meat products using HPLC fluorescent and UV detectors

Raw and pasteurised sheep’s, cow’s and goat’s milk, eggs, and beef samples from different local markets in Jordan were collected during a period of 5 months (January through May 2007) and examined for aflatoxins B1(AFB1), B2(AFB2), G1(AFG1), G2(AFG2), M1(AFM1) and M2(AFM2). The samples were analysed with high performance liquid chromatography (HPLC) using UV and Fluorescent detectors. The analysed samples of milk collected in January were found to contain 0.56 μg L⁻¹ AFM1 and 0.1 μg L⁻¹ AFM2 whilst, the concentration of AFM1 and AFM2 was < 0.05 μg L⁻¹ for milk samples collected between March and May. The AFB1, AFB2, AFG1 and AFG2 contents in the analysed food products ranged from 1.10 to 8.32 μg L⁻¹ and 0.15 to 6.36 μg L⁻¹ in imported and fresh meat samples collected during March, respectively. The mean recovery for the HPLC method was 92% to 109% and the quantification levels were 50 ng L⁻¹ for AFM1 and AFM2. The AFM1 was found in 10% of the tested samples with concentrations between 0.08 and 1.1 μg kg⁻¹ and AFM2 was only found in 1.82% of the tested samples with a level of 0.1 μg kg⁻¹. The AFM1 levels in the examined foods were higher than the maximum level of AFM1 in liquid milk set by the European Community and Codex Alimentarius of 50 ng L⁻¹.     

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

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

A novel quantum dot-based fluoroimmunoassay method for detection of Enrofloxacin residue in chicken muscle tissue

A rapid indirect competitive fluorescence-linked immunosorbent assay (cFLISA) based on quantum dots (QDs) as the fluorescent marker has been developed for the detection of Enrofloxacin (ENR) in chicken muscle tissue. The end-point fluorescent detection system was carried out using QDs conjugated with goat anti-mouse secondary antibody. The cFLISA method allowed for ENR determination in a liner working range of 1–100 ng mL⁻¹ with the 50% inhibition value (IC₅₀) of 8.3 ng mL⁻¹ and the limit of detection (LOD) of 2.5 ng mL⁻¹. The recoveries for chicken muscle samples spiked with ENR at levels of 50–200 μg kg⁻¹ ranged from 81% to 94% with coefficients of variation (CV) of 10–13%. In real chicken tissue sample analysis, the results of cFLISA were similar to those obtained from an indirect competitive enzyme-linked immunosorbent assay (cELISA) to a high performance liquid chromatography method (HPLC), which indicated that cFLISA is suitable as screening method for the monitoring of veterinary drug residues.     

Aflatoxins and ochratoxin A in stored barley grain in Spain and impact of PCR-based strategies to assess the occurrence of aflatoxigenic and ochratoxigenic Aspergillus spp

Contamination of barley by moulds and mycotoxins results in quality and nutritional losses and represents a significant hazard to the food chain. The presence of aflatoxin B1 (AFB1), B2 (AFB2), G1 (AFG1) and G2 (AFG2) and ochratoxin A (OTA) in stored barley in Spain has been studied. Species-specific PCR assays were used for detection of Aspergillus flavus, A. parasiticus, A. ochraceus, A. steynii, A. westerdijkiae, A. carbonarius and A. niger aggregate in mycotoxin-positive barley samples at different incubation times (0, 1 and 2 days). Classical enumeration techniques (CFU/g) in different culture media for evaluation of Aspergillus in sections Flavi, Circumdati and Nigri were also used. One hundred and five barley kernel samples were collected in Spanish grain stores from 2008 to 2010, and analyzed using a previously optimized method involving accelerated solvent extraction, cleanup by immunoaffinity column, liquid chromatographic separation, post-column derivatization with iodine and fluorescence detection. Twenty-nine samples were contaminated with at least one of the studied mycotoxins. AFB1, AFB2, AFG1, AFG2, and OTA were detected in 12.4%, 2.9%, 4.8%, 2.9%, and 20% of the samples, respectively. Aflatoxins and OTA co-occurred in 4.8% of the samples. Maximum mycotoxin levels (ng/g) were 0.61 (AFB1), 0.06 (AFB2), 0.26 (AFG1), 0.05 (AFG2), and 2.0 (OTA). The results of PCR assays indicated the presence of all the studied species, except A. westerdijkiae. The PCR assays showed high levels of natural contamination of barley with the studied species of Aspergillus which do not correspond to the expected number of CFU/g in the cultures. These results suggest that a high number of non-viable spores or hyphae may exist in the samples. This is the first study carried out on the levels of aflatoxins and OTA in barley grain in Spain. Likewise, this is the first report on the presence of aflatoxigenic and ochratoxigenic Aspergillus spp. in barley grain naturally contaminated with those mycotoxins using a species-specific PCR approach.     

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.     

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

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

Dispersive liquid–liquid microextraction combined with sweeping micellar electrokinetic chromatography for the determination of some neonicotinoid insecticides in cucumber samples

A rapid, simple and sensitive method has been developed for the analysis of some neonicotinoid insecticides in cucumber samples by using dispersive liquid–liquid microextraction (DLLME) coupled with sweeping in micellar electrokinetic chromatography (MEKC). Under optimised conditions, the enrichment factors were achieved in the range from 4000 to 10,000. The linearity of the method was in the range from 2.7 to 200 ng g⁻¹ for thiacloprid, acetamiprid and imidacloprid, and in the range from 4.0 to 200 ng g⁻¹ for imidaclothiz in cucumber samples, with the determination coefficients (r²) ranging from 0.9924 to 0.9968. The limits of detection (LODs, S/N = 3) ranged from 0.8 to 1.2 ng g⁻¹. The relative standard deviations (RSDs) at the concentration levels of 10.0 and 50.0 ng g⁻¹ each of the neonicotinoid insecticides in cucumber samples varied from 3.8% to 6.3%. The developed method has been successfully applied to the analysis of the neonicotinoid insecticides in cucumbers with a satisfactory result.     

Immunoassay based on monoclonal antibody for aflatoxin detection in poultry feed

An indirect competitive enzyme-linked immunosorbent assay (ic-ELISA) based on an anti-aflatoxin B₁ monoclonal antibody was standardised and validated for aflatoxin screening in poultry feed samples and its performance was compared to high-performance liquid chromatography (HPLC). The ic-ELISA showed good linearity (r² = 0.994) and detection limits of 1.25 ng g⁻¹ for broiler feed and 1.41 ng g⁻¹ for laying hen feed. Mean aflatoxin recovery rates by ic-ELISA were 102% (laying hen feed) and 98% (broiler feed). Aflatoxins were detected in 88.2% of the 34 broiler feed samples by ic-ELISA and HPLC at means of 10.48 ng g⁻¹ and 8.41 ng g⁻¹, respectively, while 92% of laying hen feed samples (n = 36) showed aflatoxin contamination at means of 20.83 and 19.75 ng g⁻¹. The standardised ic-ELISA showed reliability and a high correlation with HPLC of 0.97 (broiler feed) and 0.98 (laying hen feed) indicating its potential for aflatoxin screening in poultry feed samples.     

Presence of mycotoxins in sorghum and intake estimation in Tunisia

Sorghum samples (n = 60) from Tunisian markets were analysed for the occurrence of 22 of both traditional and emerging mycotoxins. Samples were extracted with a QuEChERS-like method and mycotoxins were detected by LC-MS/MS. This method was validated and adequate analytical parameters were obtained. All samples had contamination with mycotoxins and several samples had higher contamination levels than European Union legislative limits (MLs). The most frequently found mycotoxins were ENB (100%), OTA (98%), ENA₁ (63%), ENB₁ (56%), BEA (48%), AFB1 (38%) and STG (33%). Mean contaminations were 30.7, 1.93, 33.2, 51.0, 15.4, 1.49 and 20.5 µg kg^–1, respectively. While two samples were contaminated with FB2 and FB3 at mean values of 16.2 and 45.9 µg kg^–1, respectively, one sample was contaminated with AFB2 and ZEA at levels of 0.82 and 45.0 µg kg^–1, respectively. The results were used to estimate the daily intake of mycotoxins through sorghum consumption with regard to normal consumers (low-risk population) and high consumers such as babies (high-risk consumers) who are facing an alarming situation.     

Multiresidue determination of tetracycline antibiotics in propolis by using HPLC-UV detection with ultrasonic-assisted extraction and two-step solid phase extraction

An analytical method in propolis was developed and validated for the determination of four tetracyclines (TCs) by high performance liquid chromatography (HPLC) for the first time. After extraction by ultrasound, the extracting solution was subjected to Oasis HLB and weak cation-exchange cartridge to remove water-soluble and fat-soluble flavonoids, aromatic acids, terpenoid compounds, wax, and pollen debris. The calibration curves of fortified samples showed acceptable linear response (R² > 0.99) through a range of 100–5000 ng g⁻¹ in 20 replicates of six concentrations and the analysis of variance (ANOVA) was performed to validate the regression data. The limit of quantification of four TCs were 100 and 150 ng g⁻¹, respectively. The recoveries of the four TCs for propolis samples spiked with 100–500 ng g⁻¹ were in the range of 61.9–88.5% and the RSDs were between 4.80% and 13.2%. Traces of tetracycline were found in two out of 30 analysed real samples.     

Determination of copper with 5,5-dimethylcyclohexane-1,2,3-trione 1,2-dioxime 3-thiosemicarbazone in olive oils by adsorptive stripping square wave voltammetry

In the present paper a method for the determination of Cu in olive oil samples by adsorptive stripping square wave voltammetry (Ad-SSWV) is presented. It has been proven that Cu reacts with 5,5-dimethylcyclohexane-1,2,3-trione 1,2-dioxime 3-thiosemicarbazone, DCDT, in strongly acid media giving rise to a complex. In Ad-SSWV the complex Cu–DCDT experiments an adsorptive reductive process which promotes the appearance of a peak at −0.570 V. The extraction process of Cu from olive oil is carried out with hot concentrated HCl. Calibration graph has been constructed from 0 to 35 ng mL⁻¹ and the detection limit was 0.49 ng mL⁻¹. The method has been applied to commercial olive oils samples and the amounts of Cu found are very similar to those obtained when samples are analysed by AAS.     

Analysis of fumonisins B1, B2 and their hydrolysed metabolites in pig liver by LC–MS/MS

A liquid chromatography–tandem mass spectrometry method for the quantitative determination of fumonisin FB1, FB2 and their respective hydrolysed metabolites HFB1 and HFB2 in pig liver has been developed and validated. The method was based on an easy extraction procedure followed by SPE purification. Chromatographic separation of the analytes was performed on a C18 column using 0.3% of formic acid in water and acetonitrile as elution solvent. The mass spectrometer operated in the positive electrospray ionisation mode (ESI+) using multiple reaction monitoring (MRM). An intralaboratory validation was carried out with fortified samples for determining precision, trueness, specificity, limit of detection (LOD) and limit of quantification (LOQ). The method showed good performance characteristics and proved to be sensitive, selective and reliable. The LOD was 0.05 ng g⁻¹ and the LOQ was 10 ng g⁻¹ for all the analytes. The developed method has been applied to seven pig liver samples in order to test its applicability to evaluate exposure of food animals to fumonisins.     

Simultaneous determination of multi-mycotoxins in palm kernel cake (PKC) using liquid chromatography-tandem mass spectrometry (LC-MS/MS)

Palm kernel cake (PKC) is a useful source of protein and energy for livestock. Recently, it has been used as an ingredient in poultry feed. Mycotoxin contamination of PKC due to inappropriate handling during production and storage has increased public concern about economic losses and health risks for poultry and humans. This concern has accentuated the need for the evaluation of mycotoxins in PKC. Furthermore, a method for quantifying mycotoxins in PKC has so far not been established. The aims of this study were therefore (1) to develop a method for the simultaneous determination of mycotoxins in PKC and (2) to validate and verify the method. A liquid chromatography-tandem mass spectrometry (LC-MS/MS) method using an electrospray ionisation interface (ESI) in both positive- and negative-ion modes was developed for the simultaneous determination of aflatoxins (AFB₁, AFB₂, AFG₁ and AFG₂), ochratoxin A (OTA), zearalenone (ZEA), deoxynivalenol (DON), fumonisins (FB₁ and FB₂), T-2 and HT-2 toxin in PKC. An optimum method using a 0.2 ml min^–1 flow rate, 0.2% formic acid in aqueous phase, 10% organic phase at the beginning and 90% organic phase at the end of the gradient was achieved. The extraction of mycotoxins was performed using a solvent mixture of acetonitrile–water–formic acid (79:20:1, v/v) without further clean-up. The mean recoveries of mycotoxins in spiked PKC samples ranged from 81% to 112%. Limits of detection (LODs) and limits of quantification (LOQs) for mycotoxin standards and PKC samples ranged from 0.02 to 17.5 μg kg^?1 and from 0.06 to 58.0 μg kg^?1, respectively. Finally, the newly developed method was successfully applied to PKC samples. The results illustrated the fact that the method is efficient and accurate for the simultaneous multi-mycotoxin determination in PKC, which can be ideal for routine analysis.     

Natural levels of ethyl formate in stored grains determined using an improved method of analysis

Ethyl formate was readily determined with a gas chromatograph (GC) equipped with a flame ionisation detector. Natural levels of ethyl formate in Australian wheat, barley, oats, and canola were analysed by GC, after extraction with ammonium nitrate solution. Background levels of ethyl formate were present in newly harvested and stored grain. The levels of ethyl formate (0.1-0.6 mg kg⁻¹) in grains varied with commodity, temperature, moisture and period of storage. The values ranged from 0.1 to 0.2 mg kg⁻¹ for newly harvested wheat, barley and oats, and 0.3-0.4 mg kg⁻¹ for newly harvested canola. Ethyl formate was present in grains at harvest, increased during the first 7 months of storage, and then began to decline, particularly at grain temperatures higher than 25°C and moisture contents higher than 12.5% (for wheat, barley and oats) and 6.5% (for canola). The natural levels of ethyl formate should be considered when establishing maximum residue limits.     

Modified paramagnetic beads in a microfluidic system for the determination of zearalenone in feedstuffs samples

In this work, we have developed and characterised a novel microfluidic immunoassay methodology for rapid and sensitive quantification of ZEA in feedstuffs samples. The detection of ZEA was carried out using a competitive direct immunoassay method based on the use of anti-ZEA monoclonal antibodies immobilized on magnetic microspheres 3-aminopropyl-modified manipulated for an external remobilize magnets. The ZEA in feedstuffs sample is allowed to compete with ZEA-horseradish peroxidase (HPR) conjugated for the immobilized anti-ZEA antibody. The HPR, in the presence of hydrogen peroxide (H₂O₂) catalyses the oxidation of 4-tert-butylcatechol (4-TBC) whose back electrochemical reduction was detected on gold electrode at 0.0 V. The calculated detection limits for electrochemical detection and ELISA procedure were 0.41 and 2.56 μg kg⁻¹ respectively, the intra and inter-assay coefficients of variation were below 6.5% and the total assay time was 30 min. The microfluidic immunosensor showed higher sensitivity and lower detection limits than the standard ELISA method, which shows potential for detecting ZEA in foods and feeds diagnosis.     

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

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

Development and validation of a simple analytical method for the determination of 2,4,6-trichloroanisole in wine by GC–MS

A novel method for the residue analysis of wine spoilage compound 2,4,6-trichloroanisole is reported. Wine (60 ml) was extracted with 2 ml toluene in presence of 24 g MgSO₄ and 6 g NaCl. Cleanup of the toluene phase by dispersive solid phase extraction with mixture of 100 mg CaCl₂, 25 mg primary secondary amine and 50 mg MgSO₄ was effective in minimising co-extractives and matrix effects. Time-of-flight and tandem mass spectrometric parameters were optimised to achieve linearity over 0.25–500 ng ml⁻¹ and method detection limit 0.0083 ng ml⁻¹ which is well below the odour threshold of 0.04 ng ml⁻¹. Recoveries at 0.04, 0.2 and 0.8 ng ml⁻¹ were within 80–110% (±8%). The method was reproducible when tested for Argentinean wines with intra-laboratory Horwitz ratios being <0.20 in white and red wines at both the laboratories of India and Argentina. The method could be successfully applied for incurred wine samples.