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1.
Aflatoxins are the only mycotoxins with legal limits for spices in the European Union. A further limit for ochratoxin A is expected to be adopted soon. Thus, rapid simultaneous methods for quantifying the five mycotoxins are sought. Liquid extraction, immunoaffinity column cleanup, and HPLC-FD with a wavelength program were optimized for the analysis of the five mycotoxins in paprika, a complex fatty matrix. The limits of detection ranged from 0.3 to 0.6 microg/kg. Repeatability (RSDr) ranged from 7.9 to 13.4%, and recoveries were between 61.4 and 77.8%, in both cases at the lower spike level. Aflatoxins, when found, were far below the two legal limits of 5 microg/kg for aflatoxin B 1 and 10 microg/kg for total aflatoxins. Ochratoxin A was more frequently found, with a mean of 11.8 microg/kg, and in a more varied range (SD = 18.9 microg/kg). When an automation of the precolumn derivatization step was attempted, the procedure proved to be unfeasible, but experience derived from this trial and from the general employment of this reaction enables some comments on the possibilities and limitations of this procedure and on research for an alternative one to be made.  相似文献   

2.
Aflatoxins are a group of very carcinogenic mycotoxins that can be found on a wide range of food commodities including nuts, cereals, and spices. In this study, the first LC-MS/MS stable isotope dilution assay (SIDA) for the determination of aflatoxins in foods was developed. The development of this method was enabled by easily accessible isotope-labeled (deuterated) aflatoxins B2 and G2, which were synthesized by catalytic deuteration of aflatoxin B1 and G1, purified, and well-characterized by NMR and MS. All four aflatoxins of interest (B1, B2, G1, and G2) were quantified in food samples by using these two labeled internal standards. The response factors (RF) of the linear calibrations were revealed to be matrix independent for labeled aflatoxin B2/aflatoxin B2 and labeled aflatoxin G2/aflatoxin G2. For labeled aflatoxin B 2/aflatoxin B 1 and labeled aflatoxin B2/aflatoxin G1 matrix-matched calibration was performed for the model matrices almonds and wheat flour, showing significant differences of the RFs. Limits of detection (LOD) were determined by applying a statistical approach in the presence of the two model matrices, yielding 0.31 microg/kg (aflatoxin B1), 0.09 microg/kg (aflatoxin B2), 0.38 microg/kg (aflatoxin G1), and 0.32 microg/kg (aflatoxin G2) for almonds (similar LODs were obtained for wheat flour). Recovery rates were between 90 and 105% for all analytes. Coefficients of variation (CV) of 12% (aflatoxin B1), 3.6% (aflatoxin B2), 14% (aflatoxin G1), and 4.8% (aflatoxin G2) were obtained from interassay studies. For further validation, a NIST standard reference food sample was analyzed for aflatoxins B1 and B2. The method was successfully applied to determine trace levels of aflatoxins in diverse food matrices such as peanuts, nuts, grains, and spices. Aflatoxin contents in these samples ranged from about 0.5 to 6 microg/kg.  相似文献   

3.
A collaborative study of a liquid chromatographic method for the determination of aflatoxins B1, B2, G1, and G2 was conducted in laboratories located in the United States, Canada, South Africa, and Switzerland. Twenty-one artificially contaminated raw peanuts, peanut butter, and corn samples containing varying amounts of aflatoxins B1, B2, G1, and G2 were distributed to participating laboratories. The test portion was extracted with methanol-0.1N HCl (4 + 1), filtered, defatted with hexane, and then partitioned with methylene chloride. The concentrated extract was passed through a silica gel column. Aflatoxins B1 and G1 were derivatized with trifluoroacetic acid, and the individual aflatoxins were determined by reverse-phase liquid chromatography with fluorescence detection. Statistical analysis of the data was performed to determine or confirm outliers, and to compute repeatability and reproducibility of the method. For corn, relative standard deviations for repeatability (RSDr) for aflatoxin B1 ranged from 27.2 to 8.3% for contamination levels from 5 through 50 ng/g. For raw peanuts and peanut butter, RSDr values for aflatoxin B1 were 35.0 to 41.2% and 11.2 to 19.1%, respectively, for contamination levels from 5 through 25 ng/g. RSDr values for aflatoxins B2, G1, and G2 were similar. Relative standard deviations for reproducibility (RSDr) for aflatoxin B1 ranged from 15.8 to 38.4%, 24.4 to 33.4%, and 43.9 to 54.0% for corn, peanut butter, and raw peanuts, respectively. The method has been adopted official first action for the determination of aflatoxins B1, B2, G1, and G2 in peanut butter and corn at concentrations greater than or equal to 13 ng total aflatoxins/g.  相似文献   

4.
A previously published method for ochratoxin A was evaluated and proved appropriate for simultaneous determination of aflatoxins, ochratoxin A, sterigmatocystin, and zearalenone, with considerable savings in time and reagent costs. The detection limits were 2, 5, 15, and 55 micrograms/kg, respectively. The recoveries and coefficients of variation obtained with artificially contaminated samples were 91-101% and 0-16% for aflatoxin B1, 98-117% and 0-17% for sterigmatocystin, and 96-107% and 0-17% for zearalenone, respectively. The coefficients of variation for naturally contaminated samples (aflatoxins in rice and ochratoxin A in beans) ranged from 0 to 8%. The method was used to survey 296 samples that included 10 cultivars of dried beans, 8 types of corn products, 3 types of cassava flour, and both polished and parboiled rice between May 1985 and June 1986 in Campinas, Brazil. Only aflatoxin B1 (9 samples, 20-52 micrograms/kg), aflatoxin G1 (4 samples, 18-31 micrograms/kg), and ochratoxin A (5 samples, 32-160 micrograms/kg) were found. The average contamination percentage was 4.7%; beans showed the highest (6.6%) and rice showed the lowest (3.3%) incidence rates. Zearalenone and sterigmatocystin were not detected. Positive samples were confirmed by chemical derivatization, corroborated by development in 3 solvent systems.  相似文献   

5.
beta-Cyclodextrin enhances the fluorescence of aflatoxins B1 and G1 in aqueous systems. This effect was utilized in developing a unique reverse-phase liquid chromatographic (LC) method for determination of aflatoxins B1, B2, G1, and G2 (B1 detection limit 1 ppb), without preparing derivatives of B1 and G1. The aflatoxins are dissolved in methanol or the mobile phase for injection onto the LC system. Using a mobile phase of methanol-beta-cyclodextrin (1 + 1), the aflatoxins are resolved on a C18 column. Fluorescence of the aflatoxins is enhanced by post-column introduction of an aqueous concentrated beta-cyclodextrin solution. All 4 aflatoxins elute within 10 min in the order G2, G1, B2, B1. Fluorescence responses for B1 and G1 standards were linear over the concentration range 0.5-10 ng, yielding correlation coefficients (r) of 0.9989 and 1.000, respectively. The average peak response ratio for G1:B1 for the mobile phase-enhancement solution described was 0.765 with a coefficient of variation (CV) of 0.98%. CVs were 6.2, 9.0, and 7.5% for multiple assays of aflatoxin B1 in 3 samples of naturally contaminated corn. For samples of corn spiked to a total B1 content of 8.3 ng/g, average B1 recovery was 90% (CV 11.7%).  相似文献   

6.
A simple, systematic analytical method for multiple mycotoxins was developed for detecting 14 mycotoxins; aflatoxins B1, B2, G1, and G2, sterigmatocystin, T-2 toxin, diacetoxyscirpenol, neosolaniol, fusarenon X, zearalenone, ochratoxin A, citrinin, luteoskyrin, and rugulosin. These mycotoxins were extracted with 20% H2SO4-4% KCl-acetonitrile (2 + 20 + 178), defatted with isooctane, and transferred to chloroform. The chloroform extract was cleaned up by silica gel column chromatography; the first 10 toxins were eluted with chloroform-methanol (97 + 3) and the remaining 4 toxins with benzene-acetone-acetic acid (75 + 20 + 5). Each fraction was analyzed by thin layer chromatography for the final determination. The method has been applied to polished rice, rough rice, corn, wheat, and peanuts as an analytical screening procedure. The detection limits in these commodities ranged from 10.00 to 800.0 microgram/kg, depending on the mycotoxin, but all limits were superior to those obtained for the individual mycotoxins by using other methods.  相似文献   

7.
This study was designed to develop a sensitive liquid chromatography tandem mass spectrometry (LC-MS/MS) method for the simultaneous detection and quantification of 25 mycotoxins in cassava flour, peanut cake and maize samples with particular focus on the optimization of the sample preparation protocol and method validation. All 25 mycotoxins were extracted in a single step with a mixture of methanol/ethyl acetate/water (70:20:10, v/v/v). The method limits of quantification (LOQ) varied from 0.3 μg/kg to 106 μg/kg. Good precision and linearity were observed for most of the mycotoxins. The method was applied for the analysis of naturally contaminated peanut cake, cassava flour and maize samples from the Republic of Benin. All samples analyzed (fifteen peanut cakes, four maize flour and four cassava flour samples) tested positive for one or more mycotoxins. Aflatoxins (total aflatoxins; 10-346 μg/kg) and ochratoxin A (相似文献   

8.
A multimycotoxin thin layer chromatographic method is described for the analysis of corn. Aflatoxins are extracted from the samples with acetonitrile-water, and sodium bicarbonate is added to separate the acidic ochratoxin from zearalenone and aflatoxin B1. After chloroform extraction, 1N NaOH is added to separate zearalenone and aflatoxin B1. The separated mycotoxins are spotted on TLC plates, which are then examined under ultraviolet light. The following recoveries (%) were obtained for corn samples: aflatoxin B1 71, ochratoxin A 87, and zearalenone 85. The limits of detection for the respective mycotoxins were 2, 40, and 200 ppb.  相似文献   

9.
This paper reports the results of an extensive survey on the occurrence of filamentous fungi isolated from wine-grapes in Lebanon and to test their ability to produce ochratoxin A (OTA) and aflatoxin B1 (AFB1) on CYA culture medium, in order to assess their potential for producing these mycotoxins on grapes. From the 470 grapes samples taken during season 2004, 550 fungi strains were isolated with 490 belonging to Aspergillus spp. and 60 belonging to Penicillium spp. All these isolated fungi starins were tested for their ability to produce OTA and AFB1. Aspergillus carbonarius shows that it is the only species able to produce OTA with a production percentage reaching 100% and a maximum concentration of 52.8 microg/g of Czapek yeast extract agar (CYA). In its turn, Aspergillus flavus was considered as the only AFB1-producing species with production percentage of 45.3% and a maximum concentration reaching 40 microg/g CYA. A total of 47 handmade musts produced from the collected grapes were also analyzed in order to correlate the presence of OTA in must and the occurrence of filamentous fungi on grapes; 57.4% were contaminated with OTA at low level with concentrations ranging between 0.011 and 0.221 microg OTA L(-1). The analysis of these must samples was not performed with regard to AFB1. Seventy samples of finish red wine were also assayed for OTA content. The results showed that 42 of the tested samples (60%) were found to be positive for OTA with low levels (0.012-0.126 microg OTA L(-1)).  相似文献   

10.
A method is described for simple and rapid determination of aflatoxins in corn, buckwheat, peanuts, and cheese. Aflatoxins were extracted with chloroform-water and were purified by a Florisil column chromatographic procedure. Column eluates were concentrated and spotted on a high performance thin layer chromatographic (HPTLC) plate, which was then developed in chloroform-acetone (9 + 1) and/or ether-methanol-water (94 + 4.5 + 1.5) or chloroform-isopropanol-acetone (85 + 5 + 10). Each aflatoxin was quantitated by densitometry. The minimum detectable aflatoxin concentrations (micrograms/kg) in various test materials were 0.2, B1; 0.1, B2; 0.2, G1; 0.1, G2; and 0.1, M1. Recoveries of the aflatoxins added to corn, peanut, and cheese samples at 10-30 micrograms/kg were greater than 69% (aflatoxin G2) and averaged 91%, B1; 89%, B2; 91%, G1; 78%, G2; and 92%, M1. The simple method described was compared with the AOAC CB method, AOAC BF method, and AOAC milk and cheese method. These methods were applied to corn, peanut, and cheese composites spiked with known amounts of aflatoxins, and to naturally contaminated buckwheat and cheese. Recoveries were much lower for the BF method compared with our simple method and the CB method.  相似文献   

11.
A laboratory model, set to simulate the in vivo conditions of the porcine gastrointestinal tract, was used to study the small intestinal absorption of several mycotoxins and the effectiveness of Standard Q/FIS (a carbon/aluminosilicate-based product) in reducing mycotoxin absorption when added to multitoxin-contaminated diets. Mycotoxins were quickly absorbed in the proximal part of the small intestine at levels of 105 and 89% for fumonisins B1 and B2, respectively, 87% for ochratoxin A, 74% for deoxynivalenol, 44% for aflatoxin B1, and 25% for zearalenone. Addition of Standard Q/FIS to the diet (up to 2%, w/w) significantly reduced mycotoxin absorption, in a dose-dependent manner, up to 88% for aflatoxin B1, 44% for zearalenone, and 29% for the fumonisins and ochratoxin. Standard Q/FIS was ineffective in reducing deoxynivalenol uptake. These findings suggest that Standard Q/FIS can be used as a multitoxin adsorbent material to prevent the individual and combined adverse effects of mycotoxins in animals.  相似文献   

12.
A liquid chromatographic (LC) technique has been developed that uses the Mycosep multifunctional cleanup (MFC) column. MFC columns provide a rapid 1-step extract purification. They are designed to retain particular groups of compounds that may create interferences in analytical methods. At the same time, MFC columns allow compounds of interest to pass through. In the method presented, test samples are extracted in a blender with acetonitrile-water (9 + 1). A portion of the extract is forced through an MFC column designed especially for analysis of numerous mycotoxins. Analytical interferences are retained, while aflatoxins pass through the column. Aflatoxins B1 and G1 are converted to their hemiacetals by heating a mixture of purified extract and water-trifluoroacetic acid-acetic acid (7 + 2 + 1) at 65 degrees C for 8.5 min. An aliquot of this mixture is analyzed by isocratic LC with acetonitrile-water mobile phase and fluorescence detection. A detection limit of less than 0.5 ng/g for aflatoxin B1 was obtained. Average recoveries greater than 95% total aflatoxins (B1, B2, G1, and G2) and coefficients of variation of less than 3% were obtained. The method was successfully applied to the following commodities: corn, almonds, pista-chios, walnuts, peanuts, Brazil nuts, milo, rice, cottonseed, corn meal, corn gluten meal, fig paste, and mixed feeds.  相似文献   

13.
A chemical cleanup procedure for low-level quantitative determination of aflatoxins in major economically important agricultural commodities using HPLC has been developed. Aflatoxins were extracted from a ground sample with MeOH/H2O (80:20, v/v), and after a cleanup step on a minicolumn packed with Florisil, aflatoxins were quantified by HPLC equipped with a C18 column, a photochemical reactor, and a fluorescence detector. Water/MeOH (63:37, v/v) served as the mobile phase. Recoveries of aflatoxins B1, B2, G1, and G2 from peanuts spiked at 5, 1.7, 5, and 1.7 ng/g were 89.5+/-2.2, 94.7+/-2.5, 90.4+/-1.0, and 98.2+/-1.1, respectively (mean+/-SD, %, n=3). Similar recoveries, precision, and accuracy were achieved for corn, brown and white rice, cottonseed, almonds, Brazil nuts, pistachios, walnuts, and hazelnuts. The quantitation limits for aflatoxins in peanuts were 50 pg/g for aflatoxin B1 and 17 pg/g for aflatoxin B2. The minimal cost of the minicolumn allows for substantial savings compared with available commercial aflatoxin cleanup devices.  相似文献   

14.
A monoclonal antibody (Mab) against ochratoxin A (OTA) was produced from the hybridoma cell line C7G25, which was established by the fusion of Sp2/0-Ag14 myeloma cells with spleen cells isolated from a BALB/c mouse immunized with the OTA-bovine serum albumin conjugate. This Mab belongs to the IgG(2a) heavy-chain subclass with a kappa-type light chain. The level of 50% inhibition concentration was 1.20 ng/mL in a competitive direct enzyme-linked immunosorbent assay (cdELISA), and the detection limit was 0.12 ng/mL. This antibody is specific for OTA but also shows cross-reactivity with ochratoxin B (31.7%) in a cdELISA. On the basis of the sandwich format using the produced Mab against OTA, a rapid immunochromatographic assay was developed to efficiently detect OTA. This method was able to detect up to 500 ng/mL of OTA in <10 min.  相似文献   

15.
A method for the accurate one-dimensional thin layer chromatographic (TLC) determination of aflatoxins B1, B2, G1, and G2 in mixed feeds is presented. The aflatoxins are extracted from the sample with chloroform and purified by solvent partitioning. Each aflatoxin is separated from pulp interference by thin layer chromatography on aluminum-backed silica plates. The separated aflatoxins are detected by fluorescence densitometry. Average recoveries for samples spiked from 10 to 100 ppb B1 and G1 and from 3 to 30 ppb B2 and G2 are 82, 84, 95, and 94% for B1, B2, G1, and G2, respectively. The above recovery data, when analyzed for overall method repeatability, produced relative standard deviations of 6.8, 4.3, 6.9, and 7.6% for B1, B2, G1, and G2, respectively. Minimum detection level is less than 1 ppb for each aflatoxin. B1 is confirmed by trifluoroacetic acid derivative formation on a silica TLC plate.  相似文献   

16.
Aflatoxins produced by Aspergillus flavus are commonly found in human and animal foods including grains, cereals, peanut products, sorghum, and soy seeds. Exposure to aflatoxins has been associated with carcinogenicity. This paper reports a simple, portable, and rapid fluorescence polarization (FP) assay for aflatoxin determination in grains. This immunoassay is field portable, homogeneous, and without any washing and cleaning steps. The assay is based upon the competition between free aflatoxin and an aflatoxin-fluorescein tracer for an aflatoxin-specific monoclonal antibody in solution. A series of naturally contaminated corn, sorghum, peanut butter, and peanut paste samples were analyzed by FP and compared with HPLC results. Similarly, spiked popcorn samples were analyzed by FP. FP results of naturally contaminated samples correlated well with HPLC (r (2) = 0.97). FP analysis of spiked popcorn samples (with a mixture of B(1)/B(2)/G(1)/G(2), 7/1/3/1, w/w) gave a good correlation with spiked values (r (2) = 0.99). However, FP consistently underestimated the aflatoxin contents. This was perhaps due to low cross-reactivity of the antibody used toward B(2), G(1), and G(2) aflatoxins. These results combined with the portability and simplicity of the assay suggest that the assay can be used for screening total aflatoxin in grains.  相似文献   

17.
A simple, rapid, and solvent-efficient method for determining aflatoxins in corn and peanut butter is described. Aflatoxins B1, B2, G1, and G2 were extracted from 50 g sample with 200 mL methanol-water (85 + 15). A portion of the extract was diluted with 10% NaCl solution to a final concentration of 50% methanol, and then defatted with hexane. The aflatoxins were partitioned into chloroform. The chloroform solution was evaporated, and the residue was placed on a 0.5 g disposable silica gel column. The column was washed with 3 mL each of hexane, ethyl ether, and methylene chloride. Aflatoxins were eluted with 6 mL chloroform-acetone (9 + 1). The solvent was removed by evaporation on a steam bath, and the aflatoxins were determined using thin layer chromatography (TLC) with silica gel plates and a chloroform-acetone (9 + 1) developing solvent. Overall average recovery of aflatoxin B1 from corn was 82%, and the limit of determination was 2 ng/g. For mass spectrometric (MS) confirmation, aflatoxin B1 in the extract from 3 g sample (20 ng/g) was purified by TLC and applied by direct on-column injection at 40 degrees C into a 6 m fused silica capillary gas chromatographic column. The column was connected directly to the ion source. After injection, the temperature was rapidly raised to 250 degrees C, and the purified extract was analyzed by negative ion chemical ionization MS.  相似文献   

18.
A screening method has been developed for simultaneous determination of aflatoxin B1 and ochratoxin A in black olives. The technique includes extraction of both mycotoxins with aqueous methanol, cleanup using lead acetate, defatting with hexane, partitioning in chloroform, and thin layer chromatography. Detection limits are 5-7 micrograms aflatoxin B1 and 20 micrograms ochratoxin A/kg.  相似文献   

19.
A liquid chromatographic (LC) method was developed for the determination of aflatoxins in feedstuffs containing citrus pulp. The feed-stuff sample is extracted with chloroform, followed by Sep-Pak Florisil cartridge cleanup and Sep-Pak C18 cartridge cleanup. The final eluate (water-acetone, 85 + 15, v/v) is submitted to reverse-phase liquid chromatography with water-methanol-acetonitrile (130 + 70 + 40, v/v/v) as mobile phase and postcolumn derivatization with iodine. Citrus components are removed from the extract efficiently. The limit of detection for aflatoxin B1 is less than 1 microgram/kg. Other aflatoxins can also be detected and measured. Recoveries of aflatoxins B1, B2, G1, and G2 for dairy rations spiked at 13, 5, 10, and 4 micrograms/kg were 87, 86, 81, and 82%, respectively. Corresponding coefficients of variation were 3.1, 3.6, 5.2, and 3.8%, respectively.  相似文献   

20.
A high pressure liquid chromatographic method has been developed for determining aflatoxins B1, B2, G1, and G2 in peanut butter. The method is based on extraction with acidified aqueous methanol, partition of the aflatoxin into methylene chloride, and purification of the extract on a 2 g silica gel column. The extracted aflatoxins are resolved on a microparticulate (10 micrometer) porous silica gel column in ca 10 min with a water-washed chloroform-cyclohexane-acetonitrile solvent that contains 2% isopropanol. The fluorescence detection system determines aflatoxins B1, B2, G1, and G2 at low levels, i.e., 0.25 ppb B1, 0.5 ppb G1, and 0.2 ppb B2 and G2. Multiple assays of 5 samples of naturally contaminated peanut butters containing total aflatoxins (B1 + B2 + G1 + G2) at levels of 1, 2, 3, 9, and 17 ppb gave intralaboratory coefficients of variation of 7, 4, 4, 11, and 3%, respectively. Samples spiked at levels of 5, 9, and 17 ppb total aflatoxins showed recoveries of 79, 81, and 81%, respectively.  相似文献   

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