液相色谱-飞行时间质谱同时测定粮食中13种真菌毒素

建立了液相色谱-飞行时间质谱(LC-TOF MS)联用技术同时检测小麦和玉米中镰刀菌、曲霉菌和青霉菌产生的13种真菌毒素的分析方法。样品经乙腈-水-乙酸(84∶15∶1,体积比)混合溶剂提取,My-cosep 226多功能净化柱和强阴离子交换柱净化后,采用LC-TOF MS检测。在电喷雾正离子模式下,以保留时间和化合物精确分子离子质量对真菌毒素进行识别,以10 ppm为提取离子窗口进行定量。结果表明,13种真菌毒素在一定的线性范围内线性关系良好,相关系数均大于0.99,质量精确度均小于5 ppm,回收率为70%～113%,相对标准偏差为0.2%～14.5%。该方法可用于粮食中多种真菌毒素的同时测定。     

Development of an economic ultrafast liquid chromatography with tandem mass spectrometry method for trace analysis of multiclass mycotoxins in Polygonum multiflorum

Rapid, economic, and highly effective determination of multiple mycotoxins in complex matrices has given huge challenges for the analytical method. In this study, an economic analytical strategy based on sensitive and rapid ultrafast liquid chromatography coupled to hybrid triple quadrupole/linear ion trap mass spectrometry technique was developed for the determination of seven mycotoxins of different chemical classes (aflatoxin B₁, B₂, G₁, and G₂, ochratoxin A, T‐2 toxin, and HT‐2 toxin) in Polygonum multiflorum. Target mycotoxins were completely extracted using a modified quick, easy, cheap effective, rugged, and safe method without additional clean‐up steps. The types of extraction solvents and adsorbents for the extraction procedure were optimized to achieve high recoveries and reduce coextractives in the final extracts. Due to significant matrix effects for all analytes (≤68.9% and ≥110.0%), matrix‐matched calibration curves were introduced for reliable quantification, exploring excellent linearity for the seven mycotoxins with coefficients of determination >0.9992. The method allowed high sensitivity with limit of detection in the range of 0.031–2.5 μg/kg and limit of quantitation in the range of 0.078–6.25 μg/kg, as well as satisfactory precision with relative standard deviations lower than 8%. Recovery rates were between 74.3 and 119.8% with relative standard deviations below 7.43%. The proposed method was successfully applied for 24 batches of P. multiflorum samples, and six samples were found to be positive with aflatoxin B₁, B₂, G₁, or ochratoxin A. The method with significant advantages, including minimum analytical time, low time and solvent consumption, and high sensitivity, would be a preferred candidate for economic analysis of multiclass mycotoxins in complex matrices.     

Fast determination of 14 mycotoxins in chestnut by dispersive solid‐phase extraction coupled with ultra high performance liquid chromatography‐tandem mass spectrometry

A dispersive solid‐phase extraction coupled with ultra high performance liquid chromatography with tandem mass spectrometry method was developed and validated for the simultaneous determination of T‐2 toxin, penicillic acid, fumonisins B₁, B₂, and B₃, aflatoxins B₁, B₂, G₁, and G₂, ochratoxin A, deoxynivalenol, 3‐acetyldeoxynivalenol, 15‐acetyldeoxynivalenol, and zearalenone in chestnut samples. The method was used to analyze 136 samples obtained from Shandong province in China. The mycotoxins were extracted using a dispersive solid‐phase extraction method and cleaned using an improved quick, easy, cheap, effective, rugged, and safe approach. The mycotoxins were then detected using a triple‐quadrupole mass spectrometer. The limits of detection and quantification ranged from 0.02 to 1 and 0.1 to 2 μg/kg, respectively. The recovery rates ranged from 74.2 to 109.5%, with relative standard deviations below 15%. A total of 71 samples were contaminated with seven mycotoxins at concentrations ranging from 1.2 to 105.5 μg/kg, with a number of samples exceeding the maximum limits set in the European regulations for mycotoxins in unprocessed chestnuts.     

Identification and characterization of new Fusarium masked mycotoxins,T2 and HT2 glycosyl derivatives,in naturally contaminated wheat and oats by liquid chromatography–high‐resolution mass spectrometry

The presence of glucoside derivatives of T‐2 and HT‐2 toxins (type A trichothecene mycotoxins) in naturally contaminated wheat and oats is reported for the first time. The use of advanced high‐resolution mass spectrometry based on Orbitrap technology allowed to obtain molecular structure details by measuring exact masses of main characteristic fragments, with mass accuracy lower than 2.8 ppm (absolute value). A monoglucoside derivative of T‐2 toxin and two monoglucoside derivatives of HT‐2 toxin were identified and characterized. The analysis of their fragmentation patterns provided evidence for glucosylation at C‐3 position for T‐2 toxin and at C‐3 or C‐4 position for HT‐2 toxin. A screening for the presence of these new masked forms of mycotoxins was carried out on a set of naturally contaminated wheat and oats samples. On the basis of peak area ratio between glucoside derivatives and free T‐2 and HT‐2 toxins, the presence of glucoside derivatives was more likely in wheat than in oats samples. The present work confirms the widespread occurrence of trichothecene glucosides in cereal grains naturally contaminated with the relevant unconjugated toxins, thus suggesting the importance of developing suitable analytical methods for their detection. Besides toxicity studies, tracking down these new masked forms of trichothecenes along the food/feed chain would enable to collect information on their relevance in human/animal exposure to mycotoxin risk. Copyright © 2012 John Wiley & Sons, Ltd.     

固相萃取-液相色谱-质谱法测定益母草中7种真菌毒素

建立液相色谱-质谱法测定益母草中黄曲霉毒素G2、G1、B2、B1、T-2毒素、赭曲霉毒素A、展青霉素7种真菌毒素的方法。样品经过70%甲醇溶液超声提取,用水稀释后用固相萃取柱富集净化,以C18色谱柱分离,多反应监测(MRM)模式进行检测,7种真菌毒素得到快速分离。7种真菌毒素的质量浓度与色谱峰面积成良好的线性关系,线性相关系数在0.9972~0.9992之间,检出限为0.047~4.724μg/kg。平均加标回收率为72.8%~95.5%,测定结果的相对标准偏差为3.1%~5.7%(n=6)。该检测方法可同时高效准确检测益母草中7种真菌毒素的含量,可用于赭曲霉毒素A的定量风险评估,为益母草的安全评价提供了科学依据。     

Development of microextraction techniques in combination with GC–MS/MS for the determination of mycotoxins and metabolites in human urine

Simple and highly efficient sample preparation procedures, namely, dispersive liquid–liquid microextraction and salting‐out liquid–liquid extraction for the analysis of ten Fusarium mycotoxins and metabolites in human urine were compared. Various parameters affecting extraction efficiency were carefully evaluated. Under optimal extraction conditions, salting‐out liquid–liquid extraction showed a better accuracy (84–96%) and precision (<14%) than dispersive liquid–liquid microextraction. Hence, a multibiomarker method based on salting‐out liquid–liquid extraction followed by gas chromatography with tandem mass spectrometry was proposed. Satisfactory results in terms of validation were achieved. The method resulted in low limits of detection and quantitation within the range of 0.12–4 and 0.25–8 μg/L, respectively. The method accuracy and precision were evaluated at three spiking levels (8, 25 and 100 μg/L) and the recoveries were in a range from 70 to 120% with relative standard deviations lower than 15%. Matrix effect was evaluated and matrix‐matched calibrations were used for quantitation purpose. The developed method was applied in 12 human urine samples as a pilot study before and after sample treatment with β‐glucuronidase before the analysis to quantify the mycotoxin conjugates. Total deoxynivalenol (free + conjugated) was found in 83% of samples at an average concentration in positive samples of 31.6 μg/L.     

超高效液相色谱-四极杆/静电场轨道阱高分辨质谱法测定焙烤食品及其原料中11种真菌毒素

建立了超高效液相色谱-四极杆/静电场轨道阱高分辨质谱(UPLC-HRMS)测定焙烤食品及其原料中11种真菌毒素的检测分析方法。样品经20 mL含1%(体积分数)甲酸的乙腈-水(9∶1,v/v)溶液提取,经2.0 g无水硫酸、0.5 g氯化钠和300 mg C18盐析、净化后进行检测。采用CORTECS C18色谱柱(100 mm×2.1 mm,1.6μm),以含0.1%(体积分数)甲酸的2 mmol/L乙酸铵溶液和含0.1%(体积分数)甲酸的2 mmol/L乙酸铵甲醇溶液为流动相进行梯度洗脱。结果显示,11种真菌毒素在各自的质量浓度范围内线性关系良好(相关系数r2≥0.996 0),方法的定量限为0.15~20.00μg/kg,样品加标回收率为64.38%~122.61%,相对标准偏差为1.52%~12.99%(n=6)。该方法简单快速、灵敏度高、结果准确、可靠,利用该方法可有效测定焙烤食品及其原料中常见真菌毒素的含量。     

高效液相色谱-串联质谱法同时测定饲料中11种霉菌毒素

建立了高效液相色谱-串联质谱(HPLC-MS/MS)同时检测饲料中11种霉菌毒素的分析方法。样品经乙腈提取,MycoSep 228多功能净化柱填料和PRIME HLB固相萃取柱净化;采用Agilent Zorbax SB-C18色谱柱(150 mm×2.1 mm,3.5μm)分离,以甲醇和5 mmol/L乙酸铵水溶液(含0.1%(v/v)甲酸)为流动相梯度洗脱;采用ESI源正、负离子同时扫描,多反应监测(MRM)模式测定,内标法定量。结果表明,11种目标物在各自的线性范围内线性关系良好,相关系数均大于0.99,定量限为2.0~50.0μg/kg。11种霉菌毒素在3个加标水平(1、2、5倍定量限)下的平均回收率为79.3%~101.6%,相对标准偏差(RSD)为5.9%~13.2%(n=5)。该法简单快速,净化效果好,灵敏度高,适用于饲料中11种霉菌毒素的分析。