HPLC测定玉米中玉米赤霉烯酮和赭曲霉毒素A

建立玉米中玉米赤霉烯酮(Zearalenone,ZEN)和赭曲霉毒素A(Ochratoxin A,OTA)的超高效液相色谱检测方法。样品用乙腈/水(90∶10,体积比)振荡提取,过滤后过真菌毒素净化柱,取上清液氮气吹干,用流动相溶解残渣,过0.22μm有机滤膜于进样瓶,采用高效液相色谱检测法(High Performance Liquid Chromatography,HPLC)分析。在ZORBAX SB-C18反相柱上分离后,ZEN以甲醇/水(60∶40,体积比)作为流动相,OTA以乙腈/水/乙酸(43∶56∶1,体积比)为流动相,荧光检测器检测。ZEN的线性范围为0.5μg/mL~6.0μg/mL,相关系数大于0.99;OTA的线性范围为0.05μg/mL~0.8μg/mL,相关系数大于0.99。3个不同水平的加标平均回收率为86.4%~114.1%,相对标准偏差不大于10%。ZEN和OTA的检测限分别为5.0μg/kg和1.0μg/kg。应用该方法对收集的不同玉米样品进行测定,发现ZEN和OTA污染较严重,超标率均为71.4%。该方法具有操作简单、灵敏度高、重现性好等特点,能够用于玉米样品中真菌毒素的测定。     

SPE-HPLC-MS/MS测定粮食中玉米赤霉烯酮和脱氧雪腐镰刀菌烯醇

建立了以乙腈-水(84∶16,v/v)为提取溶剂、正己烷萃取和HLB固相萃取(SPE)净化、水和乙腈C18反相色谱梯度洗脱、电喷雾负离子多反应监测模式(MRM)同时测定粮食中玉米赤霉烯酮(ZEN)和脱氧雪腐镰刀菌烯醇(DON)2种物质含量的HPLC-MS/MS检测方法。结果表明,ZEN和DON检出限(以信噪比S/N=3计)依次为0.1μg/kg和2μg/kg,定量限(以信噪比S/N=10计)依次为0.3μg/kg和6μg/kg;两者线性范围分别为0.20~25μg/L(相关系数r=1.000 0)和4.0~500μg/L(相关系数r=0.999 9);不同样品2种目标物3个水平的加标回收率在88.7%~102.3%之间,相对标准偏差(RSD)在2.4%~7.3%之间。应用该方法对所检小麦样品进行分析,结果均发现严重污染样品的2种目标毒素。     

超导体包被免疫荧光试纸法同时测定玉米中脱氧雪腐镰刀菌烯醇和玉米赤霉烯酮适用性研究

为了能同时快速测定玉米中脱氧雪腐镰刀菌烯醇(deoxynivalenol,DON)和玉米赤霉烯酮(Zearalenone,ZEN)两种毒素,采用基于超导体包被的免疫荧光快速定量检测技术,检测了多个玉米样品,结合加标回收率、稳定性、检出限、精密度等指标,并与液相色谱法的检测结果进行比较分析,评估方法的适用性。结果表明,超导体包被的免疫荧光试纸法检测DON含量在100～2 500μg/kg,检测ZEN含量在5～200μg/kg的范围内线性良好。DON在500～2 000μg/kg添加水平范围内,回收率为103.72%～108.17%,ZEN在50～150μg/kg添加水平范围内,回收率为97.81%～111.27%。该方法于液相色谱法检测结果对比,DON相对偏差在3.72%～8.17%,ZEN相对偏差在2.19%～11.27%,均低于液相色谱法允许偏差23%和15%,超导体包被的免疫荧光试纸法是一种有效、实用、快速、定量的分析方法,能满足同时检测玉米中DON和ZEN的要求。     

北京地区部分市售食用植物油中玉米赤霉烯酮和脱氧雪腐镰刀菌烯醇的污染状况分析

目的分析北京市售植物油中玉米赤霉烯酮和脱氧雪腐镰刀菌烯醇的污染状况。方法实验采用同位素稀释-超高效液相色谱-串联质谱法,在多反应监测模式下对所采集玉米油、花生油、调和油等食用油样中的玉米赤霉烯酮(ZEN)、脱氧雪腐镰刀菌烯醇(DON)进行了检测,并对其污染现状进行分析。结果 ZEN和DON在3~500 ng/mL浓度范围内线性良好,相关系数为0.999,加标回收率为95.05%~107.10%,ZEN和DON方法的检出限分别为0.03μg/kg和0.9μg/kg;ZEN和DON方法的定量限分别为0.1μg/kg和3μg/kg。对北京市售30个油样检测结果表明,DON为未检出,ZEN检出率为100%,最高含量为333μg/kg,最低含量为1.95μg/kg,平均含量为67.7μg/kg,低于欧盟规定的限量标准400μg/kg。结论通过分析食用油中真菌毒素的含量状况,初步了解了北京市售植物油中玉米赤霉烯酮和脱氧雪腐镰刀碱烯醇的污染状况。     

基于超导体包被的免疫荧光快检试纸法同时测定玉米中3种毒素的适用性研究

应用基于超导体包被的免疫荧光快速定量检测技术，建立了一种AFB1-ZEN-DON三合一联检卡法，可同时快速测定玉米中的黄曲霉毒素B1(AFB1)、玉米赤霉烯酮（ZEN）、呕吐毒素（DON）三种常见真菌毒素。对质控样准确度、加标回收率、检测限、定量限进行测定，检测多个玉米样品并与液相色谱法的检测结果进行对比，评估方法的适用性。结果表明，AFB1-ZEN-DON三合一联检卡检测AFB1含量在0.5～30μg/kg,ZEN含量在5～200μg/kg,DON含量在100～2 500μg/kg的范围内线性良好。AFB1在5～20μg/kg加标量范围时，回收率为97.62%～99.37%,ZEN在50～150μg/kg加标量范围时，回收率为98.03%～104.07%, DON在500～2000μg/kg加标量范围时，回收率为102.49%～108.53%。与液相色谱法相比，使用该方法检测AFB1相对偏差在–2.84%～11.91%,ZEN相对偏差在–3.43%～18.12%,DON相对偏差在1.83%～12.91%，均在液相色谱法允许偏差20%、15%和23%之内。该方法检测AFB1检测限为0....     

玉米及其制品和小麦及其制品中白僵菌素和恩镰孢菌素的高效液相色谱-串联质谱检测方法的建立

目的建立了玉米及其制品和小麦及其制品中白僵菌素(beauvericin,BEA)、恩镰孢菌素A(enniatin A,ENA)、恩镰孢菌素A_1(enniatin A_1,ENA_1)、恩镰孢菌素B(enniatin B,ENB)和恩镰孢菌素B_1(enniatin B_1,ENB_1)的高效液相色谱-串联质谱检测方法。方法样品经乙腈-水(85∶15,V/V)提取、室温静置、固相萃取柱净化后,以2 mmol/L乙酸铵水-乙腈作为流动相,采用电喷雾电离正离子多反应监测模式进行检测。结果该方法对玉米及其制品和小麦及其制品中BEA和4种恩镰孢菌素(ENNs)的检出限范围分别为0.01~0.12和0.02~0.21μg/kg,定量限范围分别为0.02~0.44和0.05~0.68μg/kg,平均加标回收率范围分别为91.6%~149.7%和100.5%~128.2%,基质抑制或增强效应分别为88.0%~101.5%和55.8%~106.5%,且BEA和4种ENNs在所测基质中线性关系和精密度良好,相关系数均0.99,相对标准偏差均15%。结论所建立的方法操作简便、灵敏度高、准确性好,可用于玉米及其制品和小麦及其制品中BEA和4种ENNs的测定。     

啤酒酿造原料中黄曲霉毒素B_1、玉米赤霉烯酮和赭曲霉毒素A同时检测分析方法研究

研究建立同时检测啤酒及酿造原料中黄曲霉毒素B1、玉米赤霉烯酮和赭曲霉毒素A的免疫亲和柱净化-柱后化学衍生-高效液相色谱方法。样品经过甲醇-水(80∶20,v/v)提取,通过免疫亲和柱进行富集和净化,采用Thermo BDS HYPERSIL C18色谱柱,以乙腈-2%乙酸(40∶60,v/v)为流动相,等度洗脱,柱后以0.5%碘溶液衍生、改变波长荧光检测。结果表明,黄曲霉毒素B1、玉米赤霉烯酮和赭曲霉毒素A检出限分别为0.05μg/kg(AFB1)、3.19μg/kg(ZEA)和0.22μg/kg(OTA),标准曲线的线性范围分别为0.2~10.0μg/L、10.0~1000.0μg/L和5.0~50.0μg/L;在大麦样品中加标回收率为90.0%~109.5%,相对标准偏差为2.18%~4.94%。被检37个样品的真菌毒素含量检测结果表明,正常贮存下的啤酒原料均未检测出真菌毒素,但在霉变大麦样品中可以检测出少量的真菌毒素。     

赭曲霉毒素A单克隆抗体免疫亲和柱的制备

制备了赭曲霉毒素A(OTA)单克隆抗体免疫亲和柱,并用间接竞争ELISA和HPLC法评价了免疫亲和柱的性能,其柱容量(结合OTA的能力)约为200ng,加标回收率为90.38%～100.1%,可反复使用3次。 建立了免疫亲和柱-HPLC联用分析谷物中OTA的方法,最低检出限为0.2μg/kg,线性范围为0.6～400μg/kg,OTA加标量为1～10μg/kg时谷物样品中的回收率为78.7%～87.1%,变异系数小于6.5%。用此法检测了大米、小麦、玉米和玉米饲料等15份市售样品,检出率为46.7%,其中OTA的最高含量为0.785μg/kg。     

赭曲霉毒素A时间分辨荧光侧流层析试纸条的研究

目的 构建一种基于时间分辨荧光纳米微球的赭曲霉毒素A (ochratoxin A, OTA)侧流层析试纸条。方法 基于免疫层析原理, 以时间分辨荧光纳米微球为信号探针, 降低非特异性荧光的干扰, 提高检测灵敏度, 并通过优化样品提取液和样品稀释液, 进一步提高现场检测OTA的灵敏度和准确性。结果 OTA在1~50 μg/kg范围内, T线和C线荧光强度的比值与OTA浓度的对数值具有良好的线性关系, 相关系数r2为0.9981~0.9998。不同基质中OTA的检出限(limit of detection, LOD)和定量限(limit of quantitation, LOQ)分别为0.401 μg/kg~0.614 μg/kg和0.970 μg/kg~1.617 μg/kg, 加标回收率为89.53%~118.37%, 相对标准偏差(relative standard deviations, RSDs)小于12% (n=3), 且与呕吐毒素、伏马菌素B1、黄曲霉毒素B1、玉米赤霉烯酮和T-2毒素的交叉反应率均小于5%, 特异性良好。基于荧光定量快速检测技术平台OTA侧流层析试纸条可在8 min内快速准确地定量检测出待测样本中OTA的含量。结论 本研究所制备的时间分辨荧光侧流层析试纸条可实现玉米、小麦和饲料中OTA的快速定量检测, 并具有成本低、灵敏度高、操作简便、准确高、重复性好、特异性好的优点, 可满足国内外OTA检测的技术要求, 为真菌毒素快检技术的发展提供技术支撑。     

安徽河南粮食中脱氧雪腐镰刀菌烯醇和玉米赤霉烯酮的污染调查

目的：调查安徽、河南两省粮食中镰刀菌毒素污染情况。方法：以玉米、小麦为材料,分别采用酶联免疫吸附法(ELISA)和高效液相色谱法(HPLC)检测了脱氧雪腐镰刀菌烯醇(DON)和玉米赤霉烯酮(ZEN)的含量,采用SPSS软件对检测结果进行统计学分析。结果：安徽河南两省玉米、小麦中的DON和ZEN含量平均值分别为424.0μg/kg和187.2μg/kg,检出率分别为76.7%和75.3%。其他省份的玉米、小麦中DON和ZEN含量的平均值为52.2μg/kg和24.1μg/kg,检出率分别为60%和70%。结论：和其他省份相比,安徽、河南两省的粮食受镰刀菌毒素污染程度更严重。根据现有的国家限量标准,ZEN的超标率比DON更高。     

免疫亲和柱净化-HPLC法同时测定玉米制品中的赭曲霉毒素A和玉米赤霉烯酮

建立了同时测定玉米制品中赭曲霉毒素A和玉米赤霉烯酮的免疫亲和柱净化高效液相色谱方法。试样经乙腈-水溶液(体积比80?20)提取,复合免疫亲和柱富集和净化后,采用Syncronis C18色谱柱(250 mm×4.6 mm,5μm)固定相,以0.5%冰乙酸-乙腈-甲醇(体积比10?45?45)为流动相,等度洗脱,荧光检测器检测(Ex:333 nm,Em:470 nm)。结果表明:赭曲霉毒素A和玉米赤霉烯酮检出限分别为5.65×10-2μg/kg和4.53×10-1-μg/kg;标准曲线的线性范围分别为1~50 ng/mL(r=0.9999)和10~500 ng/mL(r=0.9999);3个不同水平的加标平均回收率为95.81%~98.31%,RSD值为1.24%~1.65%。20批试样中赭曲霉毒素A和玉米赤霉烯酮测定值分别为0.59~0.69μg/kg和3.84~5.22μg/kg,均无过量残留。该方法具有操作简便、灵敏度高、重现性佳等优点,适用于同时检测玉米制品中赭曲霉毒素A和玉米赤霉烯酮。     

Development and application of analytical methods for the determination of mycotoxins in organic and conventional wheat.

The aim of this study was to develop a multicomponent analytical method for the determination of deoxynivalenol (DON), ochratoxin A (OTA) and zearalenone (ZEN), nivalenol (NIV), 3-acetyl-DON (3-acDON), 15-acetyl-DON (15-acDON), zearalenol (ZOL) and citrinin (CIT) in wheat. It also aimed to survey the presence and amounts of DON, OTA and ZEN in Belgian conventionally and organically produced wheat grain and in wholemeal wheat flours. After solvent extraction, an anion-exchange column (SAX) was used to fix the acidic mycotoxins (OTA, CIT), whilst the neutral mycotoxins flowing through the SAX column were further purified by filtration on a MycoSep cartridge. OTA and CIT were then analysed by high-performance liquid chromatography (HPLC) using an isocratic flow and fluorescence detection, while the neutral mycotoxins were separated by a linear gradient and detected by double-mode (ultraviolet light fluorescence) detection. The average DON, ZEN and OTA recovery rates from spiked blank wheat flour were 92, 83 and 73% (RSDR = 12, 10 and 9%), respectively. Moreover, this method offered the respective detection limits of 50, 1.5 and 0.05 microg kg-1 and good agreement with reference methods and inter-laboratory comparison exercises. Organic and conventional wheat samples harvested in 2002 and 2003 in Belgium were analysed for DON, OTA and ZEN, while wholemeal wheat flour samples were taken from Belgian retail shops and analysed for OTA and DON. Conventional wheat tended to be more frequently contaminated with DON and ZEN than organic samples, the difference being more significant for ZEN in samples harvested in 2002. The mean OTA, DON and ZEA concentrations were 0.067, 675 and 75 microg kg-1 in conventional samples against 0.063, 285 and 19 microg kg-1 in organically produced wheat in 2002, respectively. Wheat samples collected in 2003 were less affected by DON and ZEN than the 2002 harvest. Organic wholemeal wheat flours were more frequently contaminated by OTA than conventional samples (p < 0.10). The opposite pattern was shown for DON, organic samples being more frequently contaminated than conventional flours (p < 0.10).     

磁控双色上转换荧光适配体传感器同时检测玉米和燕麦粉中的赭曲霉毒素A与玉米赤霉烯酮

目的 建立上转换荧光法同时检测食品中赭曲霉毒素A（ochratoxin A,OTA）与玉米赤霉烯酮（zearalenone,ZEN）的含量 方法 利用溶剂热法合成了两种油酸封端的核壳型上转换纳米材料,通过表面改性法和戊二醛法制备了表面分别偶联OTA、ZEN适配体的核壳型上转换荧光探针。同时制备了表面原位生长四氧化三铁纳米颗粒的二硫化钼纳米片,作为淬灭剂。OTA、ZEN和适配体特异性结合后,通过磁分离后检测溶液的荧光强度值,从而实现OTA和ZEN浓度的检测。结果 该方法在最佳检测条件下,OTA与ZEN的浓度在0.05~500.00 ng/mL的线性检测范围内,与两种上转换荧光探针的荧光强度的对数值呈良好的线性关系,相关系数分别为0.9949和0.9972,对OTA的检测限为3.97×10-2 ng/mL,对ZEN的检测限为3.11×10-2 ng/mL,应用于玉米粉和燕麦粉中OTA和ZEN的检测,加标回收率为91.7%~109.4%。结论 该方法成功检测灵敏度较高,并具有较好的特异性,可用于食品中OTA和ZEN的高灵敏检测。     

高效液相色谱法同时检测粮谷中的黄曲霉毒素和赭曲霉毒素

建立粮谷类食品中黄曲霉毒素(B1、B2、G1 和G2)和赭曲霉毒素A 的同时检测方法。样品经过甲醇- 水(80:20,V/V)提取,液液萃取净化和富集后,三氟乙酸衍生,采用Agilent ZORBAX SB-C18 色谱柱(4.6mm ×250mm),以乙腈和体积分数2% 冰醋酸溶液为流动相梯度洗脱,在线变换波长荧光检测。根据3 倍信噪比的峰 响应值,确定黄曲霉毒素(B1、B2、G1 和G2)和赭曲霉毒素A 检出限分别为0.06、0.03、0.18、0.05μg/kg 和0.51μg/kg,上述5 种毒素分别在质量浓度0.05～100、0.125～25.00、0.05～100、0.125～25.00μg/L 和0.05～50.00μg/L 范围内呈线性相关,相关系数r 分别为0.9998、0.9998、0.9998、0.9996 和0.9998;在玉米、大米、小麦3 类样品中加标回收率平均为71.73%～115.37%,相对标准偏差为3.00%～9.88%,方法学验证结果表明,黄曲霉毒素和赭曲霉毒素A 5 种毒素同时检测结果与现行国标的单独检测方法检测结果无显著性差异(P ＞ 0.05)。     

Occurrence and Safety Evaluation of Ochratoxin A in Cereal‐based Baby Foods Collected from Iranian Retail Market

Contamination of agricultural commodities with ochratoxin A (OTA) is a worldwide concern in recent decades. Consumption of OTA‐contaminated baby foods exerts health implications especially in children as the most vulnerable subpopulations. In the current study, for the first time in Iran, 64 baby foods (rice, wheat, and multigrain) samples from five different brands available in the Iranian market were analyzed to determine OTA level, using a HPLC with fluorescence detector. Overall, OTA was observed in 41% of analyzed samples with a mean and maximum level of 0.42 ± 0.27 and 1.1 μg/kg, respectively. OTA levels in five of 64 samples (7.8 %) were higher than the permissible limit recommended by European Commission (permissible limit: 0.5 μg/kg) and OTA levels in two of 64 samples (3.1%) were higher than the standard set by Iranian standard organization (1 μg/kg). The highest OTA contamination was observed in rice‐based baby food cereals (1.1 μg/kg; 57% of the samples), followed by wheat‐based (23%) and multigrain (20%) samples. OTA intake in infants (≥9 months old) was more than established provisional tolerable weekly intake by the Joint FAO/WHO Expert Committee on Food Additives (JECFA) and the European Food Safety Authority (EFSA) (100 and 120 ng OTA per kg of body weight, respectively). OTA content in baby food and cereals, as well as other raw foodstuff should be investigated comprehensively to reduce the exposure rate of young children to OTA.     

Aflatoxins and ochratoxin A in flour: a survey of the Serbian retail market

This report presents data on the occurrence of aflatoxins (AF) and ochratoxin A in different types of flour marketed in Serbia. A total of 114 samples of wheat, buckwheat, rye, oat, barley, rice, millet and corn flour were collected in the period 2012–2016 and analysed using high performance liquid chromatography with fluorescence detection. Among flours other than corn, AFB1 was quantified only in rice, while ochratoxin A (OTA) was found in 29% of the samples. In corn flours the percentage of positive samples varied greatly over the years: AFB1 7.1–80.0%, OTA 30.0–40.6%, with a co-occurrence of 7.1–34.4%. Overall 5.2% of flours other than corn and 10.7% of corn flours exceeded the maximum levels (MLs) for AFB1 and/or OTA. The highest recorded levels were 8.80 μg kg^?1 of AFB1 (corn) and 23.04 μg kg^?1 of OTA (rye). Overall mean contamination levels of corn flours were 0.53 μg kg^?1 of AFB1 and 0.46 μg kg^?1 of OTA.     

固相萃取-HPLC法同时测定大米中甲萘威和阿特拉津

建立固相萃取-高效液相色谱二极管阵列检测器(DAD)法同时测定大米中甲萘威和阿特拉津的方法。样品经乙腈提取、净化、旋转蒸发和氮吹浓缩后,用甲醇定容至2 mL,通过XDB-C₁₈(250 mm×4.6 mm,5μm)色谱柱分离,流动相采用甲醇-水(60︰40,V/V),流速0.4 mL/min,柱温40℃,进样量5μL,在220 nm波长下测定。结果表明,在0~2.0μg/mL范围内,甲萘威和阿特拉津呈现良好线性关系,相关系数(R2)均为1.000,检出限分别为1.0μg/kg和3.0μg/kg。对0.2,0.4和0.8 mg/kg这3种不同浓度下甲萘威和阿特拉津准确度和精密度试验,回收率分别为88.3%~102.2%和87.5%~104.3%,相对标准偏差(RSD,n=6)分别为1.2%~3.3%和2.3%~4.7%。试验方法稳定性好、操作简单、灵敏度高,可满足于大米中甲萘威和阿特拉津测定。     

高效液相色谱法对玉米中玉米赤霉烯酮的测定

研究了高效液相色谱法测定玉米中玉米赤霉烯酮的方法.样品借鉴了GB/T 19540-2004中提取玉米赤霉烯酮的方法,通过Oasis HLB净化柱对提取液净化,以agilent extent C18色谱柱为分离柱,乙腈-水(V水:V乙腈=55:45)为流动相进行荧光检测(λex=235 nm,λem=460 nm).玉米赤霉烯酮的质量浓度在12～2 400μ/kg范围内呈良好线性,相关系数为0.9994,对添加高、中、低3个浓度玉米赤霉烯酮的玉米样品进行加标回收试验,平均回收率分别为96.736%、93.839%、86.240%,变异系数在1%～10%之间,最低检测限为10μ/kg.此方法对玉米中玉米赤霉烯酮的检测是可行的,且可给谷物中玉米赤霉烯酮检测方法优化提供参考.     

Detoxification of zearalenone and ochratoxin A by ozone and quality evaluation of ozonised corn

Zearalenone (ZEN) and ochratoxin A (OTA) are secondary toxic metabolites of fungi that can contaminate a wide range of food and feedstuff. In this study, the effects of ozone treatment on ZEN and OTA and the quality of ozonised corn are investigated. Ozone significantly affects ZEN and OTA solutions. ZEN was undetectable 5 s after being treated with 10 mg l^–1 ozone. However, OTA was resistant to ozonation with a degradation rate of 65.4% after 120 s of treatment. Moreover, ZEN and OTA solutions were difficult to degrade after being dried by a nitrogen stream. Results showed that ozone effectively degraded ZEN and OTA in corn. The degradation rates of ZEN and OTA in corn increased with ozone concentration and treatment time. The degradation of ZEN and OTA at different ozone concentrations appropriately conformed to first-order kinetics with an R² value > 0.8749. Furthermore, under the same conditions, corn with increased moisture content (MC) (19.6%) was more sensitive to ozone than corn with a low MC (14.1%). When treated with 100 mg l^–1 ozone for 180 min, ZEN and OTA in corn with 19.6% MC decreased by 90.7% and 70.7%, respectively. To evaluate the quality of ozonised corn, subsequent quality experiments were conducted using corn samples treated at different times with 100 mg l^–1 ozone. The MC of corn decreased after ozone treatment. The whiteness and yellowness of the corn increased and decreased with increasing time, respectively. The fatty acid value of the corn increased significantly (p ≤ 0.05) after 180 min of treatment. This study verified that ozone can effectively degrade ZEN and OTA in corn, but slightly affected corn quality.