Occurrence of aflatoxins B 1 , B 2 , G 1 and G 2 in some Brazilian pet foods

The presence of cereals and grains in the formulation of pet foods suggests the need to control aflatoxin contamination in these foods. The objective of the study was to analyse domestic pet food to determine the occurrence of aflatoxins as well as their risk to animal health. One hundred food samples (45 for dogs, 25 for cats, 30 for birds) were collected at random from pet shops in Alfenas city, south-east Brazil. Thin-layer chromatography was used for separation, identification and quantification of the compounds after validation of the method. Aflatoxins were detected in 12.0% of the samples. Levels of aflatoxins (B 1 + B 2 + G 1 + G 2 ) above the maximum limit established in Brazil (50 µg kg -1 ) for animal food were detected in five of the 12 positive samples (41.7%). The concentration of total aflatoxins was 15-374 µg kg -1 (mean 131 µg kg -1 ). All samples containing peanuts were positive for aflatoxin B 1 . Aflatoxins are carcinogenic and their consumption might be a risk for domestic animal health. The high prevalence of aflatoxin B 1 in foods prepared for birds, species highly susceptible to aflatoxins, shows the need for the re-evaluation of the use of peanuts (present in seven of the eight samples positives for aflatoxin) and/or the addition of fungicides to the food.     

Co-occurrence of aflatoxins B 1 , B 2 , G 1 , G 2 , zearalenone and fumonisin B 1 in Brazilian corn

Two hundred and fourteen unprocessed corn samples (1997-98 harvest), collected at wholesale markets in different regions in Brazil, were surveyed for the occurrence of mycotoxins. The samples were analysed for aflatoxins B 1 , B 2 , G 1 , G 2 , zearalenone and fumoni1sin B 1 using in-house validated methods. The occurrence of aflatoxin B 1 , zearalenone and fumonisin B 1 was found in 38.3, 30.4 and 99.1% of the samples, respectively. Aflatoxin B 1 , zearalenone and fumonisin B 1 contamination levels varied from 0.2 to 129, 36.8 to 719, and 200 to 6100 μg/kg, respectively. The cooccurrence of the two carcinogenic mycotoxins aflatoxin B 1 and fumonisin B 1 was observed in 100% of the aflatoxin-contaminated samples (82 samples). Cooccurrences of aflatoxin B 1 : zearalenone: fumonisin B 1 and aflatoxin B 1 : aflatoxin B 2 : fumonisin B 1 were found in 18 and 43 samples, respectively.     

Occurrence of aflatoxins B1, B2, G1, and G2 in peanuts and their products marketed in the region of Campinas, Brazil in 1995 and 1996.

Samples of peanuts and their products marketed in the region of Campinas, Brazil in 1995 and 1996 were analysed in terms of aflatoxin B1, B2, G1, and G2 by thin-layer chromatography, aiming at the protection of consumer health. Of the 80 samples analysed, 35 of peanuts and 45 of peanut products, 41 (51%) presented contamination with aflatoxin, 11 of them being peanuts (27%) and 30 being products (67%), in a range of 43 to 1099 micrograms/kg for B1 + B2 + G1 + G2 with a 90th percentile of 415 (B1 + G1) and of 615 (B1 + B2 + G1 + G2). The results also demonstrated that aflatoxin B1 (AFB1) reached the highest incidence and the highest upper limits compared with all the other aflatoxins.     

Occurrence of aflatoxins (B1, B2, G1, G2) in herbal tea consumed in Turkey

Aflatoxin contents in 12 types of herbal teas were determined by high performance liquid chromatography (HPLC) with fluorescence detector using immunoaffinity column clean-up. Forty eight samples were collected from four local herbal shops in Manisa, Turkey. Of the 48 samples analyzed, 43 were aflatoxin positive. The highest concentration of aflatoxin (34.18 µg/kg) was determined in a sample of camomile tea. The occurrence of AFB1, B2, G1 and G2 was found in samples at levels of 54, 29, 71 and 46 %, respectively. Aflatoxin B1, B2, G1 and G2 contamination levels varied from 0 to 14.2, 0 to 12.4, 0 to 13.5 and 0 to 28.7 µg/kg, respectively. Aflatoxin was not detected in five samples consisting of linseed, lime and fennel tea.     

Investigation of various extractants for the analysis of aflatoxin B1 in different food and feed matrices

Various extractants were investigated concerning their suitability for aflatoxin B₁ determinations in different matrices including spices, infant formula and animal feed employing an immunoaffinity clean-up procedure. It was shown that the use of aqueous acetonitrile extractants was limited due to the fact that dry sample material can absorb significant amounts of water from theextractant. This canresultin recoveries thataretoo high and therefore in incorrect values for the aflatoxin concentration if aliquots are taken for further analysis. Acorrection of theresultsby recovery calculationusing spiked blank materialis unsuitable, since material from the same group of food (e.g. paprika powder) or feed can vary significantly in the recovery values. Therefore it is recommended that aqueous methanol extractants are used, since no significant interaction with matrix constituents was observed. In addition,aqueous acetone extractants are a useful alternative with some limitations.     

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.     

柱前衍生-高效液相色谱法检测食品中的黄曲霉毒素B_1、B_2、G_1、G_2

目的:改进柱前衍生-高效液相色谱法测定食品中的黄曲霉毒素B1、B2、G1、G2。方法:分别用甲醇-水(8∶2,v∶v)或二氯甲烷提取食品中的黄曲霉毒素。提取液经免疫亲和柱净化后,采用三氟乙酸(或甲酸)进行衍生,并利用高效液相色谱仪进行测定。结果:黄曲霉毒素B1、B2、G1、G2的检出限分别为0.2、0.2、0.2、0.2μg/kg;在低、中、高加标浓度下的回收率分别为81.0%~94.1%、75.6%~92.0%、75.0%~92.4%、77.6%~91.3%。结论:改进后的柱前衍生-高效液相色谱法克服了样品基质的干扰,测定结果更准确。     

Aflatoxins in Foods and Feedstuffs. Part 3. Fluorodensitometric method for determination of aflatoxins M1 and M2 in presence of aflatoxins B1, B2, G1 and G21

The method of aflotoxins M₁ and M₂ determination in presence of aflatoxins B₁, B₂, G₁ and G₂ is presented. The fluorescence properties of aflatoxin M₁ and M₂ are discribed. A simple method of aflatoxin M extraction is proposed. Results of different tests for confirmation of aflatoxins M are discussed. During control of powdered milk from three factories aflatoxins M were not detected in final products.     

Kinetic modelling of aflatoxins B1 conversion and validation in corn,rice, and peanut during thermal treatments

A kinetic model of the aflatoxins B₁ conversion was plotted and validated successfully in the aflatoxin B₁-contaminated rice during thermal treatments. Specifically, the kinetic Model A and Model B of the aflatoxin B₁ conversion were plotted based on the differential-scanning calorimetry and thermogravimetric analysis, respectively, with the pure aflatoxin B₁. In succession, Model A and Model B were validated in the aflatoxin B₁-contaminated corn, rice, and peanut during thermal treatments. Model A successfully simulated the conversion of aflatoxin B₁ in the aflatoxin B₁-contaminated rice with the correlation coefficients of 0.859 and average-absolute deviation of 6.61. The activation energy and conversion order of the aflatoxin B₁ conversion were 89.0 kJ/mol and 0.12, respectively. Moreover, the relationships between the conversion degree and time vs temperature were plotted based on Model A. These plots would help to predict the final content of aflatoxin B₁ after thermal treatments, and give an instruction to develop a food processing.     

Occurrence of aflatoxins M(1) and M(2) in milk commercialized in Ribeir?o Preto-SP, Brazil.

Aflatoxins are toxic metabolites found in foods and feeds. When ruminants eat foodstuffs containing aflatoxins B(1) and B(2), these toxins are metabolized and excreted as aflatoxin M(1) and M(2) in milk. The aim was to determine the incidence of these aflatoxins in commercial milk collected from supermarkets in Ribeir?o Preto-SP, Brazil, and consisting of 60 ultrahigh temperature (UHT) milk samples and 79 pasteurized milk samples. The milk samples were analysed according to method 986.16 of AOAC International. None of the milk samples analysed were contaminated with aflatoxin M(2), and aflatoxin M(1) was detected in 29 (20.9%) of samples in the range 50-240 ng l(-1). The results show that despite a high occurrence of aflatoxin M(1) in commercial pasteurized and UHT milk sold in Ribeir?o Preto in 1999 and 2000, the contamination level of these toxins could not be considered a serious public health problem according to MERCOSUR Technical Regulations. However, levels in 20.9% of the milk samples exceeded the concentration of 50 ng l(-1) permitted by the European Union. Although it is not necessary to continue monitoring the incidence and levels of aflatoxins M(1) and M(2) in milk samples, surveillance could be appropriate.     

Occurrence of aflatoxin B1, citrinin and ochratoxin A in rice in five provinces of the central region of Vietnam

The possible coexistence of three mycotoxins in rice, including aflatoxin B1 (AFB1), citrinin (CIT) and ochratoxin A (OTA), was investigated. The samples of rice were collected in large markets in five provinces of the central region of Vietnam. These toxins were extracted, purified and finally quantified by HPLC with fluorimetry detection. Contamination of AFB1 was found to be the most, followed by OTA, while contamination of CIT was insignificant. The coexistence of CIT with AFB1/OTA in rice was found in high percentage. Some samples overpassed the authorized limit by Europe in OTA and/or AFB1.     

Validation of a confirmatory analytical method for the determination of aflatoxins B1, B2, G1 and G2 in foods and feed materials by HPLC with on-line photochemical derivatization and fluorescence detection

A sensitive and selective analytical method for the simultaneous separation and quantitative determination of aflatoxins B₁, B₂, G₁ and G₂ in foodstuffs and materials for feed has been validated. The method is based on high performance liquid chromatography with on-line post-column photochemical derivatization and fluorescence detection. The chromatographic separation of aflatoxins was accomplished using a C₁₈ column eluted with an isocratic mobile phase consisting of water, methanol and acetonitrile. The sample preparation required a simple extraction of aflatoxins with MeOH/H₂O (80:20, v/v) and a purification step by immunoaffinity column cleanup. The total analysis time, including sample preparation and chromatographic separation, did not exceed 40 min with a run time of 10 min. The on-line photochemical derivatization ensures better results in terms of simplicity, sensitivity and reproducibility with respect to chemical derivatization techniques, and provides an increase of the peak resolution and an extent of automation in comparison with the electrochemical ones. The procedure for the determination of aflatoxins in food samples and cereals for animal consumption was extensively validated following Regulation (EC) No. 882/2004. Detection limits in wheat bran samples of 0.08 µg kg^?1 for AFB₁, 0.02 µg kg^?1 for AFB₂, 0.16 µg kg^?1 for AFG₁ and 0.04 µg kg^?1 for AFG₂ were attained. The method allows high recovery with mean values ranging from 72 to 94% and it satisfies the necessary requirements for sensitivity, linearity, selectivity, precision and ruggedness, demonstrating the conformity of the method with provisions of Regulation (EC) No. 401/2006.     

高效液相色谱-串联质谱法同时测定花生制品中的黄曲霉毒素B1、B2、G1、G2

采用高效液相色谱-串联质谱建立了高效液相色谱-串联质谱法同时测定花生制品中4种黄曲霉毒素(B1、B2、G1、G2)的方法.结果表明,在ESI正离子模式下,高效液相色谱-串联质谱法的最低检出限为0.2μg/kg,定量限为0.6 μg/kg;标准工作液在0.5～50.0μg/kg的范围内线性良好,相关系数达到0.9990.     

Aflatoxin M(1) and ochratoxin A in a human milk bank in the city of S?o Paulo, Brazil.

Because infants are more susceptible to the adverse effects of mycotoxins, this work was carried out to determine aflatoxin M(1) (AFM(1)) and ochratoxin A (OA) in milk from the Human Milk Bank of the Southern Regional Hospital, S?o Paulo, Brazil. Analytical methods were first established and evaluated. The methods involved the extraction of AFM(1) with methanol and OA with 1% aqueous sodium bicarbonate solution and methanol, clean-up with immunoaffinity columns having antibodies specific for each mycotoxin and quantification by high performance liquid chromatography (HPLC) with fluorescence detection. The method established for AFM(1) had mean recovery percentages of 94, 77 and 82% and coefficients of variation of 17.5, 3.4 and 4.2% at 0.01, 0.03 and 0.05 ng ml(-1), respectively. For the OA method, the corresponding values were 84, 84 and 75% for recovery and 14.1, 3.7 and 4.0% for the coefficient of variation. The limit of quantification for both methods was 0.01 ng ml(-1). Of a total of 50 samples analysed, only one was contaminated with AFM1, at 0.024 ng ml(-1), and two with OA, at 0.011 and 0.024 ng ml(-1). Although the incidence observed was low, it is recommended that the study be extended to other milk banks of the city of S?o Paulo.     

Aflatoxins in Foods and Feedstuffs. Part 2. Determination of the aflatoxin B1, B2, G1 and G2 contents in Grain crops1

The simple method for determination of small amounts of aflatoxins (about 5–10 μg/kg) was described. The method was adopted for wheat, barley, rye and oats. Difficulties of aflatoxins determination in cereals are discussed, mainly observed during purification of extracts and resolution by TLC. Different tests were compared for confirmation of aflatoxins in cereals. Results of cereal crops control for contamination with aflatoxins are presented.     

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

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

Comparison of methods and optimisation of the analysis of fumonisins B1 and B2 in masa flour,an alkaline cooked corn product

A comparison study of different extraction and clean-up procedures for the liquid chromatographic analysis of fumonisins B₁ (FB₁) and B₂ (FB₂) in corn masa flour was performed. The procedures included extraction (heat or room temperature) with acidic conditions or EDTA-containing solvents, and clean-up by immunoaffinity or C18 solid-phase extraction columns. Thereafter an analytical method was optimised using extraction with an acidic mixture of methanol–acetonitrile–citrate/phosphate buffer, clean-up through the immunoaffinity column and determination of fumonisins by liquid chromatography with automated pre-column derivatisation with o-phthaldialdehyde reagent. Recovery experiments performed on yellow, white and blue masa flours at spiking levels of 400, 800 and 1200?µg?kg^?1 FB₁ and of 100, 200 and 300?µg?kg^?1 FB₂ gave overall mean recoveries of 99% (±6%) for FB₁ and 88% (±6%) for FB₂. Good recoveries (higher than 90% for both FB₁ and FB₂) were also obtained with corn tortilla chips. The limits of quantification of the method (signal-to-noise ratio of 10) were 25?µg?kg^?1 for FB₁ and 17?µg?kg^?1 for FB₂. The method was tested on different commercial corn masa flours as well as on white and yellow corn tortilla chips, showing fumonisin contamination levels (FB₁?+?FB₂) up to 1800?µg?kg^?1 (FB₁?+?FB₂) in masa flour and 960?µg?kg^?1 in tortilla chips. Over 30% of masa flours originating from Mexico exceeded the European Union maximum permitted level.     

A Rapid Single-Extraction Method for the Simultaneous Determination of Aflatoxins B1, B2, G1, G2, Fumonisin B1, and Zearalenone in Corn Meal by Ultra Performance Liquid Chromatography Tandem Mass Spectrometry

A single-extraction method to simultaneously determine aflatoxins (B1, B2, G1, G2), fumonisin B1, and zearalenone in corn meal by ultra performance liquid chromatography tandem mass spectrometry, using a triple quadrupole, was optimized, validated, and applied in an occurrence study. Different extraction solutions were tested, with better performance for methanol/acetonitrile/water (60:20:20, v/v/v). Linearity was observed from 0.25 to 1.50 ng/mL for aflatoxins, from 20 to 120 ng/mL for fumonisin, and from 7.00 to 42.00 ng/mL for zearalenone. Significant matrix effects were shown for all groups. Selectivity was demonstrated, as matrix or spectral interferences were not observed at the predicted retention time window of the target analytes. Average recoveries of 87.57, 93.18, 93.35, 94.20, 78.76, and 95.98% were obtained for aflatoxins (B1, B2, G1, and G2) fumonisin and zearalenone, respectively. A z-score of 0.19 was estimated in a corn certified reference material for fumonisin B1. Maximum relative standard deviation values under repeatability and intermediate precision conditions were determined to be 13.6 and 13.6% for aflatoxins, 3.7 and 6.3% for fumonisin, and 3.5 and 4.0% for zearalenone, respectively. In the occurrence study, 50 samples were analyzed and 44% had measurable levels of fumonisin. Zearalenone was detected in 18%. The proposed method showed considerable advantages, considering environmental impacts, efficiency, and reliability.     

Single-Laboratory Validation for the Determination of Aflatoxin B1, B2, G1, and G2 in Foods Based on Immunoaffinity Column and Liquid Chromatography with Postcolumn Derivatization and Fluorescence Detection

This study presents a method validation procedure for the determination of aflatoxin B₁, B₂, G₁, and G₂ in hazelnut, hazelnut paste, walnut, peanut, pistachio, corn, and wheat. The method consisting of clean-up with immunoaffinity column, high performance liquid chromatography with postcolumn derivatization and fluorescence detection was validated in accordance with Commission Regulation 2004/882/EC. The selectivity, linearity, decision limit, detection capability, detection and quantification limits, precision, recovery, ruggedness, and measurement uncertainty of the method were determined. The limit of detection and limit of quantification values (μg/kg) were: aflatoxin B₁, 0.02, 0.07; aflatoxin B₂, 0.01, 0.02; aflatoxin G₁, 0.02, 0.07; and aflatoxin G₂, 0.01, 0.03. The relative standard deviation values for the repeatability and within-laboratory reproducibility were below 4 and 5 %, respectively. The recovery values of the spiked samples ranged from 80 to 105 %. These results complied with minimum performance criteria established by regulation 2006/401/EC. Therefore, the procedure can be implemented for the routine analysis of aflatoxins in the studied matrices.     

Optimisation of the determination of thiamin, 2-(1-hydroxyethyl)thiamin,and riboflavin in food samples by use of HPLC

The aim of this study was first to optimise and validate a method using an enzyme-mixture to liberate protein- and phosphate-bound thiamin and riboflavin in food by the use of ultrasonication and HPLC, and second to include the quantitation of the vitamin B₁ active compound 2-(1-hydroxyethyl)thiamin (HET).