基于自建数据库的超高效液相色谱-四极杆-飞行时间质谱法测定谷物中7种真菌毒素

Determination of seven mycotoxins in cereals by ultra performance liquid chromatography⁃quadrupole⁃time of flight mass spectrometry based on the self⁃built database

建立了基于自建数据库的超高效液相色谱-四极杆-飞行时间质谱(UPLC-Q-TOF/MS)测定水稻、小麦中7种真菌毒素的方法。样品经02%甲酸水-乙腈(50∶50,v/v)提取,用QuEChERS盐包脱水盐析,用HSS T3色谱柱(100 mm×21 mm,18μm)分离。采用UPLC-Q-TOF/MS在全信息串联质谱(MS^(E))模式下对样品进行筛查,针对筛查结果为阳性的样品,采用ESI源,在正、负离子飞行时间多反应监测(TOF-MRM)模式下进一步检测,用基质匹配标准曲线进行校准定量。结果表明,7种真菌毒素在各自范围内具有良好的线性关系,相关系数(r)为09900~09998。7种真菌毒素的检出限为050~400μg/kg,定量限为100~800μg/kg。水稻和小麦基质在低、中、高3个加标水平下的平均回收率分别为881%~1239%和1020%~1234%;RSD分别为02%~136%和08%~148%。结果表明,在46批水稻中筛查出了黄曲霉素B_(1)(AFB_(1))和黄曲霉素B_(2)(AFB_(2)),筛出率各为22%,其中有1批样品AFB1超过限量;在24批小麦中筛查出了脱氧雪腐镰刀菌烯醇(DON)和玉米赤霉烯酮(ZEN),筛出率分别为375%和792%,均未超过限量。本方法采用MSE进行定性筛查,避免干扰物因质量数和保留时间都与目标物接近形成的假阳性现象,对筛查阳性的真菌毒素采用TOF-MRM模式进行定量。该法具有快捷、准确、灵敏度高的特点,适用于水稻、小麦样品中真菌毒素残留的分离和定量检测,可为水稻、小麦中真菌毒素污染监测和风险预警提供有力的技术支持。

A method for the determination of seven mycotoxins in rice and wheat by ultra per⁃formance liquid chromatography⁃quadrupole⁃time of flight mass spectrometry(UPLC⁃Q⁃TOF/MS)based on self⁃built database was established.Samples were extracted with 0.2%formic acid aqueous solution⁃acetonitrile(50∶50,v/v),dehydrated and salted according to the QuEChERS method(4 g of magnesium sulfate,1 g of sodium chloride,1 g of sodium citrate,0.5 g of citrate disodium salt),and separated on an HSS T3 column(100 mm×2.1 mm,1.8μm).UPLC⁃Q⁃TOF/MS with MS^(E) screening was performed,and the positive and negative ions of the screened mycotoxins were calibrated and quantified using matrix⁃matched standard curves with time of flight multiple reaction monitoring(TOF⁃MRM).The results showed that aflatoxin G_(1)(AFG_(1)),aflatoxin G_(2)(AFG_(2)),aflatoxin B1(AFB1),aflatoxin B_(2)(AFB_(2)),and ochratoxin A(OTA)exhibited moderate matrix effects in rice,while OTA and zearalenone(ZEN)exhibited moderate matrix effects in wheat.The seven mycotoxins showed good lineari⁃ties in their respective concentration ranges,with correlation coefficients(r)of 0.9900-0.9998.The limits of detection(LODs)for rice and wheat were 0.50-400 and 0.50-200μg/kg,respectively,and the limits of quantification(LOQs)for both cereals were 1.00-800μg/kg.In rice,the average recoveries at three spiked levels of low,medium,and high were 88.1%-123.9%,with relative standard deviations(RSDs)of 0.2%-13.6%.In wheat,the aver⁃age recoveries at three spiked levels of low,medium,and high were 102.0%-123.4%,with RSDs of 0.8%-14.8%.As a result,one batch of 46 batches of rice was screened out for AFB_(1) and AFB_(2),with a screening rate of 2.2%,of which the measured values were 10.8μg/kg and 1.2μg/kg,respectively.According to GB 2761⁃2017,the maximum allowable level of AFB1 in rice is 10μg/kg;thus,the exceeding rate for AFB1 is 2.2%.Deoxynivalenol(DON)was screened out in 9 out of 24 batches of wheat(screening rate,37.5%),while ZEN was screened out in 19 batches(screening rate,79.2%).According to GB 2761⁃2017,the maximum allowable levels of DON and ZEN in wheat are 1000 and 60μg/kg,respectively.The levels of DON and ZEN detected in the wheat samples did not exceed these limits.The proposed method uses MSE for qualitative screening to avoid the occurrence of false positives caused by interfering com⁃pounds with mass numbers and retention times similar to those of the analytes.TOF⁃MRM mode is then used to quantify the positively screened mycotoxins.The method is fast,accu⁃rate,sensitive,and suitable for the isolation and quantitative detection of mycotoxin residues in rice and wheat samples.The findings provide powerful technical support for mycotoxin contam⁃ination monitoring in rice and wheat and early risk⁃warning efforts.