Determination of Fumonisin B1 in Corn by High Performance Liquid Chromatography With Fluorescence Detection

Abstract

A method is described for the determination of fumonisin B1 in corn. The method involves sample extraction with methanol:water (75:25) and partial purification using a solid phase extraction. Fumonisin B1 is reacted with naphthalene-2,3-dicarboxaldehyde (NDA) to produce a highly fluorescent derivative, 1-cyano-2-alkyl-benz[f]isoindole (CBI) and then separated from the sample matrix on a reverse phase C-18 column with a mobile phase of acetonitrile:water:acetic acid (55:45:1). The NDA-derivative is quantitated by fluorescence detection at 410 nm excitation and a 440 nm, long past emission filter. Recoveries of fumonisin B1 added to corn at levels of 0.25–20.0 μg/g averaged 88.1% with a coefficient of variation of 10.3%. Confirmation of fumonisin B1 in corn samples was accomplished by fast atom bombardment (FAB) spectroscopy.     

A reversed phase high performance liquid chromatography method for the determination of fumonisins B1 and B2 in food and feed using monolithic column and positive confirmation by liquid chromatography/tandem mass spectrometry

The development of a reversed phase high performance liquid chromatography fluorescence method for the determination of the mycotoxins fumonisin B(1) and fumonisin B(2) by using silica-based monolithic column is described. The samples were first extracted using acetonitrile:water (50:50, v/v) and purified by using a C(18) solid phase extraction-based clean-up column. Then, pre-column derivatization for the analyte using ortho-phthaldialdehyde in the presence of 2-mercaptoethanol was carried out. The developed method involved optimization of mobile phase composition using methanol and phosphate buffer, injection volume, temperature and flow rate. The liquid chromatographic separation was performed using a reversed phase Chromolith(?) RP-18e column (100 mm × 4.6 mm) at 30 °C and eluted with a mobile phase of a mixture of methanol and phosphate buffer pH 3.35 (78:22, v/v) at a flow rate of 1.0 mL min(-1). The fumonisins separation was achieved in about 4 min, compared to approximately 20 min by using a C(18) particle-packed column. The fluorescence excitation and emission were at 335 nm and 440 nm, respectively. The limits of detections were 0.01-0.04 μg g(-1) fumonisin B(1) and fumonisin B(2), respectively. Good recoveries were found for spiked samples (0.1, 0.5, 1.5 μg g(-1) fumonisins B(1) and B(2)), ranging from 84.0 to 106.0% for fumonisin B(1) and from 81.0 to 103.0% for fumonisin B(2). Fifty-three samples were analyzed including 39 food and feeds and 14 inoculated corn and rice. Results show that 12.8% of the food and feed samples were contaminated with fumonisin B(1) (range, 0.01-0.51 μg g(-1)) and fumonisin B(2) (0.05 μg g(-1)). The total fumonisins in these samples however, do not exceed the legal limits established by the European Union of 0.8 μg g(-1). Of the 14 inoculated samples, 57.1% contained fumonisin B(1) (0.16-41.0 μg g(-1)) and fumonisin B(2) (range, 0.22-50.0 μg g(-1)). Positive confirmation of selected samples was carried out using liquid chromatography-tandem mass spectrometry, using triple quadrupole analyzer and operated in the multiple reaction monitoring mode.     

Robotic automated clean-up for detection of fumonisins B1 and B2 in corn and corn-based feed by high-performance liquid chromatography

A high-performance liquid chromatography (HPLC) system with fluorescence detection and an automated on-line solid-phase extraction procedure for fumonisins B1 and B2 in corn and corn-based products is described. Different amounts of strong anion-exchange, C18 and end-capped C18 (C(18 ec)) silicas were tested for sample clean-up. Various HPLC parameters were analyzed. The best methodology was found to be extraction with acetonitrile-water and clean up on C(18 ec) disposable extraction cartridges. The system has the advantage of running in an unattended mode of operation and allows processing of 40 samples without system refuel, performing clean-up, o-phthaldialdehyde derivatization, injection and fumonisin detection by fluorescence detection linked to a computer integrator for automated data processing. Recoveries were performed with corn and corn-based feed samples (n=3) spiked with 0.1, 0.5, 1.0, 5.0 and 10 microg/g. Average recoveries for corn and corn-based feed were, respectively, 92.6 and 88.3% with relative standard deviations (RSDs) of 5.04 and 6.22%, for fumonisin B1 and 91.2 and 89.0% with RSDs of 5.84 and 7.88% for fumonisin B2. Detection limits (S/N=3) for corn and corn-based feed were approximately 0.03 microg/g for fumonisin B1 and 0.05 microg/g for fumonisin B2     

Characterization of the mycotoxin fumonisin B1: comparison of thermospray, fast-atom bombardment and electrospray mass spectrometry.

The utility of thermospray mass spectrometry (TSMS), fast-atom bombardment mass spectrometry (FABMS), and electrospray mass spectrometry (ESMS) for the analysis of Fumonisin B1 is investigated. In addition, the analysis of two different standards of Fumonisin B1 as well as an inoculated corn culture extract that contained Fumonisin B1 is reported. The results of these efforts show that ESMS, as well as FABMS and a combination of FAB and tandem mass spectrometry (FABMS/MS), provide useful data for the characterization of Fumonisin B1. The detection limit was 50 pg for Fumonisin B1 when analyzed by full scan FABMS, and 5 pg when analyzed by single-reaction monitoring FABMS/MS.     

Determination of fumonisins B1 and B2 in corn and corn flakes by liquid chromatography with immunoaffinity column cleanup: collaborative study.

A liquid chromatographic (LC) method for the determination of fumonisins B1 (FB1) and B2 (FB2) in corn and corn flakes was collaboratively studied by 23 laboratories, which analyzed 5 blind duplicate pairs of each matrix to establish the accuracy, repeatability, and reproducibility characteristics of the method. Fumonisin levels in the corn ranged from <0.05 (blank) to 1.41 microg/g for FB1 and from <0.05 to 0.56 microg/g for FB2, whereas in the corn flakes they ranged from <0.05 to 1.05 microg/g for FB1 and from <0.05 to 0.46 microg/g for FB2. The method involved double extraction with acetonitrile-methanol-water (25 + 25 + 50), cleanup through an immunoaffinity column, and LC determination of the fumonisins after derivatization with o-phthaldialdehyde. Relative standard deviations for the within-laboratory repeatability (RSDr) of the corn analyses ranged from 19 to 24% for FB1 and from 19 to 27% for FB2; for the corn flakes analyses, RSDr ranged from 9 to 21 % for FB1 and from 8 to 22% for FB2. Relative standard deviations for the between-laboratories reproducibility (RSDR) of the corn analyses ranged from 22 to 28% for FB1 and from 22 to 30% for the FB2; for corn flakes analyses, RSDR ranged from 27 to 32% for FB1 and from 26 to 35% for FB2. Mean recoveries of FB1 and FB2 from corn spiked with FB1 at 0.80 microg/g and with FB2 at 0.40 microg/g were 76 and 72%, respectively; for corn flakes spiked at the same levels recoveries were 110 and 97% for FB1 and FB2, respectively. HORRAT ratios for the analyses of corn ranged from 1.44 to 1.53 for FB1 and from 0.96 to 1.48 for FB2, whereas for corn flakes they ranged from 1.60 to 1.82 for FB1 and from 1.39 to 1.68 for FB2.     

Development of a liquid chromatography/tandem mass spectrometry method for the quantification of fumonisin B1, B2 and B3 in cornflakes

A liquid chromatography/tandem mass spectrometry (LC/MS/MS) method for the determination of fumonisin B1 (FB1), B2 (FB2) and B3 (FB3) in cornflakes is described. During method development, special attention was paid to the selection of a suitable internal standard (IS) in order to offer a good alternative for deuterated FB1. In this respect, the C12-sphinganine analogue (2S,3R)-2-aminododecane-1,3-diol was chosen because of its structural similarity to the fumonisin backbone and its chromatographic elution between the target analytes. For the extraction of the fumonisins from the cornflakes matrix, MeOH/H2O (adjusted to pH 4 with 0.1 M HCl; 70:30, v/v), ACN/MeOH/H(2)O (25:25:50, v/v/v) and acidified ACN/MeOH/H2O (25:25:50, v/v/v; pH 4) were evaluated. Preference was given to acidified MeOH/H2O (70:30, v/v) with mean recoveries (n=12) for FB1, FB2 and FB3 of, respectively, 84+/-10, 78+/-7 and 85+/-9%. Cleanup was performed using immunoaffinity columns (FumoniTest, VICAM). The chromatography was performed under isocratic conditions at a flow of 0.3 mL min-1 with a mobile phase consisting of ACN/H2O (60:40, v/v) containing 0.3% formic acid. The mass spectrometer was operated in the positive electrospray ionization (ESI+) mode using multiple reaction monitoring (MRM). An intralaboratory validation was conducted with fortified samples determining limits of detection (LOD), limits of quantification (LOQ), precision, trueness, specificity and measurement uncertainty. The LOD concentrations for FB1, FB2 and FB3 were 20, 7.5 and 12.5 microg/kg. The LOQs were 40 microg/kg for FB1, 15 microg/kg for FB2 and 25 microg/kg for FB3. The coefficients of variation (CVs) under repeatability conditions varied from 11 to 13% for FB1, from 9 to 14% for FB2 and from 7 to 10% for FB3. Under within-laboratory reproducibility conditions, the CVs ranged from 12 to 17% for FB1, from 9 to 16% for FB2 and from 7 to 13% for FB3. The percent bias for FB1 varied from -12 to -10%, while for FB2 and FB3 bias ranged, respectively, from -4 to -2% and from -12 to -5%. The expanded measurement uncertainties for FB1, FB2 and FB3 were, respectively, 19, 18 and 22%.     

柱后衍生-高效液相色谱法测定玉米中伏马菌素B1和B2

建立了邻苯二甲醛(OPA)柱后衍生-高效液相色谱测定玉米中伏马菌素B1和B2(FB1和FB2)的方法。采用ZORBAX SB-C18色谱柱,以0.1 mol/L磷酸二氢钠溶液(pH 3.3)-甲醇为流动相,梯度洗脱。流动相流速为0.8 mL/min,柱温40 ℃;衍生剂的流速为0.4 mL/min,衍生温度为室温。实验对衍生剂缓冲液的pH、衍生剂的浓度和流速、激发和发射波长等重要条件进行了优化。结果表明,衍生剂的pH在10.5、OPA的质量浓度为2 g/L、流速为0.4 mL/min、激发波长335 nm、发射波长440 nm时测定效果良好,FB1、FB2在0.2～20 mg/L范围内线性关系良好,相关系数大于0.999; FB1和FB2的检出限均为0.02 mg/kg;在0.1～ 4.0 mg/kg范围内,3个添加水平的平均回收率为82.5%～89.8%。该方法精确、简单、快速,适合玉米中FB1和FB2的测定。     

Determination of fumonisins B1 and B2 in corn by liquid chromatography/mass spectrometry with immunoaffinity column cleanup: single-laboratory method validation

A single-laboratory method validation was conducted to establish the effectiveness of an immunoaffinity column cleanup procedure followed by liquid chromatography/mass spectrometry (LCIMS) for the determination of fumonisins B1 and B2 (FBI + FB2) in corn. The test portion is extracted with acetonitrile-methanol-water (25 + 25 + 50). The extract is filtered, diluted with phosphate-buffered saline solution, and applied to an immunoaffinity column. FB1 + FB2 are removed with methanol and directly determined by reversed-phase LC with MS detection using selected-ion monitoring of 2 characteristic ions in each case. Test portions of blank corn samples were spiked with a mixture of FB1 + FB2 to give total levels of 200 and 500 ng/g, respectively. Recoveries of both FB1 and FB2 from spiked samples averaged 90.4-101%. Based on results for spiked raw corn (triplicates at 2 levels), the relative standard deviation for repeatability ranged from 2.8 to 7.1%. The accuracy of the method was demonstrated by analysis of Food Analysis Performance Assessment Scheme (FAPAS) test material. The method was also applied to a small survey of processed corn products such as corn chips, cornflakes, and popcorn.     

Identification and determination of fumonisin FB1 and FB2 in corn and corn products by high-performance liquid chromatography-electrospray-ionization tandem mass spectrometry (HPLC-ESI-MS-MS)

Summary A new method has been developed for qualitative and quantitative analysis of fumonisin FB₁ and FB₂ in corn and corn products. After extractive sample preparation followed by solid phase extraction using a strong anionexchanger, highest selectivity and sensitivity was achieved by high-performance liquid chromatography-electrospray-ionization, tandem mass spectrometry (HPLC-ESI-MS-MS) analysis combined with selected reaction monitoring (SRM). With deuterated fumonisin FB₁-d₆ as internal standard the routine quantitation limit for FB₁ was set at 400 pg (0.8 ng g^–1 sample) with a signal-to-noise ratio of 101. Almost all corn grit and corn meal samples under study were found to be contaminated with fumonisin FB₁ and FB₂ in concentrations ranging from 1 to 200 ng g^–1. Lower fumonisin levels were found in cornflakes, popcorn and baby food, while no fumonisins were detected in canned sweet corn.     

A simple aptamer molecular beacon assay for rapid detection of aflatoxin B1

A simple aptamer molecular beacon assay for rapid detection of aflatoxin B1 (AFB1) was achieved. AFB1-binding induced formation of a hairpin structure and closeness of fluorophore label and quencher probe, causing fluorescence decrease.     

Determination of type B fumonisin mycotoxins in maize and maize-based products by liquid chromatography/tandem mass spectrometry using a QqQlinear ion trap mass spectrometer

A sensitive liquid chromatography/tandem mass spectrometry (LC/MS/MS) method for determining the type B fumonisin mycotoxins in corn-based foodstuffs is described. Fumonisins FB1 and FB2 were extracted from a 1 g sample by homogenization with acetonitrile/water (75:25, v/v, 50 mmol/L formic acid, 25 mL final volume) and the extract was defatted on C18 phase. Volumes of 5 mL of crude extracts were cleaned up on Carbograph-4 cartridges. The final solution was analyzed by HPLC with electrospray ionization mass spectrometry in positive ion mode using multiple reaction monitoring with a QqQ linear ion trap mass spectrometer. Recoveries for spiked corn-based foodstuffs ranged from 91-105% (RSD% < or =8%), and method detection limits were < or =2 ng/g for FB1 and < or =1 ng/g for FB2. Two different spiking levels were tested (5000 and 100 ng/g for FB1, 1000 and 20 ng/g for FB2). Quantitation was achieved by an external calibration procedure using matrix-matched standards, with diclofenac added post-cleanup as internal standard for the LC/MS/MS analyses. Calibration curves showed linearity in the concentration range 0.005-5 ng/microL of final extract (0.992 < or = R2< or =0.995). Two other fumonisins, FB3 and FB4, were identified in naturally contaminated samples of corn meal using an information-dependent acquisition protocol that looped three experiments, including neutral loss scan, enhanced resolution scan, and enhanced product ion scan. FB3 and FB4 quantitation was estimated as peak area ratios relative to the FB2 response in view of the lack of both standards. This work also includes an application of the present LC/MS/MS method to some maize and maize-based product samples (corn meal, cornflakes and popcorn) collected from Italian stores. FB1 and FB2 contamination levels exceeding the European Union recommendation were found in 8 out of 15 corn meal samples.     

Evaluation of Fumonitest Immunoaffinity Columns

Abstract

A method for the determination of fumonisins B1 and B2 in corn was developed. The method involves sample extraction with methanol:water (80:20) and the use of a commercially available Fumonitest column for sample cleanup. The capacity, selectivity, column-to-column and lot-to-lot reproducibility of the Fumonitest columns were evaluated. The total capacity of the column was found to be 1.2 μg fumonisin. Both fumonisins B1 and B2 had an equal affinity toward the Fumonitest column, with the sample matrix demonstrating little effect on the column performance. The maximum sample size was 0.5 g for samples containing total fumonisins of less than 2 ppm. After elution from the immunoaffinity column, fumonisins B1 and B2 were reacted with naphthalene-2, 3-dicarboxaldehyde (NDA) to produce a highly fluorescent derivative, 1-cyano-2-alkyl-benz[f]isoindole (CBI). The derivatives were then separated from the sample matrix on a reverse phase C-18 column with a mobile phase consisting of acetonitrile:water:acetic acid (55:45:1) Average recoveries of fumonisins B1 and B2 from corn samples spiked at a level of 1000 ng (500ng B1 + 500ng B2)/g were 85.4 and 87.1%, respectively. The detection limit for B1 and B2 was estimated to be 10 and 4 ppb, respectively. The coefficient of variations for fumonisins B1 and B2 were determined to be 10.2% and 10.6%, respectively.     

Effect of temperature and solvent composition on extraction of fumonisins B1 and B2 from corn products

Fumonisins B1 and B2 were extracted from naturally contaminated corn products by using different extraction solvent compositions (methanol-water, acetonitrile-methanol-water, ethanol-water, and 100% water) and a range of temperatures from ambient to 150 degrees C. Ground samples of several corn products and 1 rice sample were mixed with an adsorbent material (Hydromatrix), and the fumonisins were extracted in 2 sequential 5 min static extractions at various temperatures. The combined extracts were cleaned up and analyzed by reversed-phase liquid chromatography with fluorescence detection after o-phthaldialdehyde-mercaptoethanol derivatization. The results showed a clear influence of temperature and solvent composition on recovery of fumonisins from some matrixes. With acetonitrile-methanol-water (1 + 1 + 2) the quantity of fumonisins extracted from naturally contaminated taco shells almost tripled in going from 23 degrees to 80 degrees C, and increased by another 30% when ethanol-water (3 + 7) was used as extraction solvent at 80 degrees C. Similar results were obtained with nacho chips. These effects were less pronounced with cornmeal, and small differences due to temperature and solvent composition were observed for corn flakes and rice. The ethanol-water extraction solvent combinations were specifically evaluated in an effort to use the cheapest, least toxic, and most environmentally friendly solvents for organic residue analysis. At 80 degrees C, ethanol-water combinations performed equally or better than methanol-water (8 + 2) or acetonitrile-methanol-water (1 + 1 + 2), combinations which are commonly used for fumonisin extractions. Even 100% water was successful for extracting fumonisins from the products, except for rice. However, increased amounts of water created technical problems and required an increased amount of Hydromatrix in the samples prior to extraction.     

Analysis of fumonisins B(1), B(2) and B(3) in corn-based baby food by pressurized liquid extraction and liquid chromatography/tandem mass spectrometry

A sensitive and reliable method using pressurized liquid extraction (PLE) and liquid chromatography (LC)/electrospray ionization (ESI) tandem mass spectrometry with a triple quadrupole (QqQ) analyzer has been developed for the analysis of fumonisin B(1) (FB(1)), fumonisin B(2) (FB(2)) and fumonisin B(3) (FB(3)) in corn-based baby foods. Influence of several extraction parameters that affect PLE efficiency such as temperature, pressure, solvent extraction, number of cycles and dispersant/clean-up agents were studied. The selected PLE operating method was: 3g of sample was packed into 11 ml stainless-steel cell and fumonisins were extracted with methanol at 40 degrees C, 34 atm in one cycle of 5 min at 60% flush. The analytes were ionized in ESI operating with positive ion mode and identified by selecting two monitoring transitions, permitting quantification and confirmation in a single injection. Recoveries ranged from 68% to 83% at fortification levels of 200 microg kg(-1) with relative standard deviation (RSD) from 4% to 12%. The limits of quantification were from 2 microg kg(-1) for FB(1) and FB(2), and 5 microg kg(-1) for FB(3), which are below the maximum residue level established by the European Union legislation in infant formulas. The proposed method was successfully applied to the analysis of twenty seven samples of baby food products collected from different markets, and one positive sample with a content of 15.9 microg kg(-1) for FB(1), 9.2 microg kg(-1)for FB(2) and 5.8 microg kg(-1) for FB(3) was obtained. Given the simplicity and potential of the proposed procedure, its application for safety control is recommended.     

膜基质免疫分析法同时检测玉米中伏马菌素B1和呕吐毒素

建立了基于聚偏氟乙烯膜(Polyvinylidene fluoride,PVDF)基质的直接竞争免疫分析法,同时检测玉米中的伏马菌素B1(Fumonisin B1,FB1)及呕吐毒素(Deoxynivalenol,DON)。PVDF膜用甲醇浸湿、激活,用移液器将FB1及DON全抗原点阵于相应的膜反应区,同时采用三聚氰氯法和高碘酸钠法分别制备抗FB1、抗DON的辣根过氧化酶(Horseradish peroxidase,HRP)标抗体(monoclonal antibody,McAb),以直接竞争免疫检测方法的模式实现同时检测玉米中的FB1及DON。该方法对于FB1和DON的检出限分别为2.5和50μg/L,样品前处理简单,检测时间15 min,可肉眼辨别结果,随机检测了30份市售玉米样品,并与市售ELISA试剂盒进行方法学比较,结果无明显差异。     

Rapid determination of fumonisins in corn-based products by liquid chromatography/tandem mass spectrometry

A simple, fast, and robust method was developed for the determination of fumonisin B1 (FB1), fumonisin B2 (FB2), and fumonisin B3 (FB3) in corn-based human food and animal feed (cornmeal). The method involves a single extraction step followed by centrifugation and filtration before analysis by ultra-performance liquid chromatographylelectrospray ionization (UPLC/ESI)-MS/MS. The LC/MS/MS method developed here represents the fastest and simplest procedure (<30 min) among both conventional HPLC methods and other LC/MS methods using SPE cleanup. The potential for high throughput analysis makes the method particularly beneficial for regulatory agencies and analytical laboratories with a high sample volume. A single-laboratory validation was conducted by testing three different spiking levels (200, 500, and 1000 ng/g for FB1 and FB2; 100, 250, and 500 ng/g for FB3) for accuracy and precision. Recoveries of FB1 ranged from 93 to 98% with RSD values of 3-8%. Recoveries of FB2 ranged from 104 to 108%, with RSD values of 2-6%. Recoveries of FB3 ranged from 94 to 108%, with RSD values of 2-5%.     

Combined synthetic/CD strategy for the stereochemical assignment of the tricarballylic acid side chains of fumonisin B(1)

The circular dichroism (CD) exciton chirality method was employed for the stereochemical assignment of the tricarballylic acid (TCA) side chains of fumonisin B(1) 1a (FB(1)). Using 2-naphthoate for chromophoric derivatization of the reduced TCA moieties, the absolute configuration was shown to be R. For additional confirmation, an optically active dihydroxy-tert-butanoate 2 related to the TCA group of fumonisin B(1) was synthesized to serve as a model compound.     

Use of ion-exchange chromatography coupled with TLC-laser scanning densitometry for the quantitation of fumonisin B(1)

A simple TLC-Laser scanning densitometric (TLC-LSD) method was developed for the quantitation of fumonisin B(1) (FB(1)) isolated from solid media cultures (corn) and liquid media cultures of toxigenic Fusarium moniliforme strains (F. moniliforme MRC 826, F. moniliforme 4223 and F. moniliforme 2927)). FB(1) was isolated from the cultures by solvent extraction (methanol:water, 3:1) and purified in a single step by ion-exchange chromatography using Dowex-1. FB(1) in the purified extracts was detected by TLC analysis using p-anisaldehyde as a post-chromatographic derivatizing agent. The major toxin identified was FB(1) (R(f) 0.51) along with traces of FB(2) (R(f) 0.57) and FB(3) (R(f) 0.60) based on their comparison with the reference standard fumonisins. The sensitivity of the TLC-LSD method for the quantitation of FB(1) was found to be 500 ng g(-1). The linear regression analysis performed for the quantitation of FB(1) by the TLC-LSD method showed a correlation coefficient (r) value of 0.9. Spiking studies revealed the recovery of standard FB(1) (5 and 10 mug g(-1)) loaded on to Dowex-1 in the range of 87-96%. The purity of FB(1) purified from the cultures was determined by the two-dimensional TLC analysis. Two-dimensional TLC-analysis of the purified FB(1) revealed the purity to be greater than 85%. The method developed may find wide application in the environmental monitoring of the FB(1) contaminations in the various agricultural commodities and screening fumonisin producing toxigenic strains of F. moniliforme.     

Evaluation of silica- and sepharose-based immunoaffinity sorbents for sample cleanup in determination of fumonisins B1 and B2 in corn products

Anti-fumonisin B1 polyclonal antibodies were isolated from the serum of rabbits, immobilized onto the surface of glutaraldehyde-activated silica or Sepharose CL-4B particles, and placed into empty small plastic solid-phase extraction cartridges. The immobilized antibodies were evaluated for their ability to retain fumonisin B1 and fumonisin B2. Cartridge capacity and elution conditions were determined, and the results were compared to those obtained with a commercially available cartridge. The cartridges, which were tested for their effectiveness to isolate the fumonisins from extracts of corn flour and nacho chips, detected fumonisins down to levels of about 20 ng/g. However, additional cleanup was required for detection at lower concentrations. With the use of a strong anion-exchange cartridge as a preliminary cleanup before immunoaffinity chromatography, the detection limit reached 2-5 ng/g in the products tested. The silica sorbent material exhibited strong interactions with the fumonisins, requiring acidified ethanol-water mixtures for elution and resulting in an additional degree of selectivity in isolating fumonisins from sample extracts. The silica-based immunoaffinity cartridges were successfully reused more than 10 times; the Sepharose-based cartridges were less robust. Liquid chromatography with fluorescence detection was used after prechromatographic derivatization with o-phthaldialdehyde-mercaptoethanol.     

Fast Determination of Fumonisin B1 and B2 in Corn Using a Modified QuEChERS Method and LC–MS–MS

A fast analytical method for the simultaneous determination of fumonisin B₁ (FB₁) and fumonisin B₂ (FB₂) in corn using a novel QuEChERS method and LC–MS–MS was developed and validated. Samples were extracted with methanol–water (3:1 v/v) by means of ultrasonic extraction. The extract was purified with a novel modified QuEChERS method. Firstly, FB₁ and FB₂ in the extract were retained with primary secondary amine (PSA). Then, FB₁ and FB₂ were released from PSA with 1.0 % formic acid in methanol. The final eluate was diluted with water, and analyzed by LC–MS–MS on a Waters Acquity BEH C₁₈ column with 0.1 % formic acid in water/methanol as mobile phase with gradient elution. Mean recoveries of 83.5–102.4 % with CVs of 3.6–10.5 % were obtained at fortification levels of 2, 50 and 1,000 μg kg⁻¹. The limit of quantification was 2.0 μg kg⁻¹.