复合柱净化-超高效液相色谱-串联质谱法测定粮油中黄曲霉毒素和玉米赤霉烯酮类毒素

目的建立C_(18)-Al_2O_3复合固相萃取柱净化,同时检测粮食及植物油中黄曲霉毒素和玉米赤霉烯酮类毒素的超高效液相色谱-串联质谱方法。方法植物油样品用乙腈-水(90∶10,V∶V)提取,粮食样品先用乙腈-乙酸-水(84∶1∶16,V∶V)再用二氯甲烷-乙酸乙酯(90∶10,V∶V)进行二次提取,进一步采用C_(18)-Al_2O_3复合固相萃取柱净化,稳定同位素内标稀释,以乙腈和水为流动相,CORTECSTMC18色谱柱(2. 1 mm×100 mm,2. 7μm)进行分离,电喷雾离子化多反应监测模式检测。结果黄曲霉毒素和玉米赤霉烯酮类毒素的线性范围分别为0. 1～30. 0和1. 0～500. 0ng/ml,检出限分别为0. 03和0. 3μg/kg,定量限分别为0. 10和1. 0μg/kg。黄曲霉毒素和玉米赤霉烯酮类毒素的平均回收率分别为68. 3%～98. 6%和84. 5%～108. 0%,相对标准偏差分别为4. 6%～11. 5%和4. 2%～9. 0%(n=6)。采用本方法分析了国内外5种质控样品,测定值均在质控样品指定范围内。对大连市市售的100份粮油样品进行测定,结果表明散装花生油和玉米油中黄曲霉毒素B1和玉米赤霉烯酮检出率较高,最高含量分别为12. 80和370. 00μg/kg。结论本方法定量准确、操作简便、更环保、检测成本低,适于批量粮油样品中黄曲霉毒素和玉米赤霉烯酮类毒素的同时检测。     

12种植物性饲料原料中霉菌毒素污染状况的调查分析

对采集的12种植物性饲料原料中的59份样品进行玉米赤霉烯酮、黄曲霉毒素B1、B2、G1、G25种霉菌毒素的检测,结果表明,玉米赤霉烯酮共10种原料26份样品有检出;黄曲霉毒素B1共7种原料10份样品有检出;黄曲霉毒素B2共2种原料2份样品有检出;黄曲霉毒素G1、G2未检出。饲料原料受玉米赤霉烯酮、黄曲霉毒素B1污染较多,应加以重视。     

粮食加工品中玉米赤霉烯酮和呕吐毒素污染 情况调查

目的初步了解粮食加工品中玉米赤霉烯酮和呕吐毒素的污染情况。方法对北京地区和网购的200批次样品进行随机抽检,分别采用液相色谱法、免疫亲和层析净化高效液相色谱法对粮食加工品中的玉米赤霉烯酮和呕吐毒素进行了检测。结果初步调查发现玉米碾磨加工品中的玉米赤霉烯酮和小麦粉中的呕吐毒素检出率较高,分别达34%和74%;散装样品中玉米赤霉烯酮含量不合格率和检出率明显高于预包装样品。结论建议监管部门加强对散装玉米碾磨加工品和小麦粉等粮食加工品中毒素指标的监控力度和监测频次,同时针对我国特殊敏感人群加快设定合理限值。     

高效液相色谱串联质谱法测定花生及制品中的五种真菌霉素

建立了高效液相色谱-串联质谱(HPLC-MS/MS)测定花生及制品中黄曲霉毒素B1、黄曲霉毒素M1、脱氧雪腐镰刀菌烯醇、赭曲霉毒素A、玉米赤霉烯酮五种真菌霉素的快速分析方法。用甲醇-水(55:45,V/V)对样品进行提取,采用真菌毒素免疫亲和柱萃取,在ESI+模式下采用多反应监测(MRM)模式进行检测。目标物在C18色谱柱上实现了有效分离,在6 min内完成一个样品的分析,相关系数(r2,n=6)大于0.999,检测结果稳定、灵敏。黄曲霉毒素B1、黄曲霉毒素M1、脱氧雪腐镰刀菌烯醇线性范围0.5~50.0μg/L,检出限为0.05μg/kg(LOD,S/N=3),赭曲霉毒素A、玉米赤霉烯酮线性范围5.0~100.0μg/L,检出限为0.5μg/kg(LOD,S/N=3),方法回收率为86.8~102.7%,精密度RSD为0.36~4.79%。该方法快速、灵敏,适用于花生及制品中黄曲霉毒素B1、黄曲霉毒素M1、脱氧雪腐镰刀菌烯醇、赭曲霉毒素A、玉米赤霉烯酮五种真菌霉素的检测与确证。     

电子鼻融合BP神经网络预测玉米赤霉烯酮和黄曲霉毒素B1含量模型研究

为探究玉米赤霉烯酮和黄曲霉毒素B_1的无损快速定量测定方法,用电子鼻对7级不同霉变程度玉米样品进行检测,并用理化分析方法分别测定霉变玉米中的玉米赤霉烯酮与黄曲霉毒素B_1含量;在提取电子鼻响应信号的积分值作为特征参量的前提下,采用BP神经网络建立不同霉变程度下玉米样品中的玉米赤霉烯酮与黄曲霉毒素B_1含量的预测模型。同时,为了获得较为可靠的BP神经网络预测模型,在神经网络结构不变的条件下,对比分析了不同训练集、测试集构建的预测模型。结果发现在各预测模型的70组测试样本中,相对误差控制在5%以内的样本数量都在60个以上,最大相对误差控制在15%以内,从而证明了BP神经网络预测模型的有效性、可靠性。该研究为实施玉米霉变毒素的快速无损检测提供了一种途径。     

时间分辨荧光免疫层析法定量检测谷物中黄曲霉毒素B1和玉米赤霉烯酮残留

建立了一种快速定量检测谷物产品中黄曲霉毒素(Aflatoxin B1,AFB1)和玉米赤霉烯酮(Zearalenone,ZEN)的时间分辨荧光免疫层析方法。采用时间分辨荧光微球标记黄曲霉毒素B1抗体和玉米赤霉烯酮抗体,研究了如p H值、标记抗体浓度、荧光探针使用量、检测T线包被原浓度、质控C线羊抗鼠Ig G浓度、样品前处理方法等因素对时间分辨荧光免疫层析方法灵敏度的影响。结果表明:AFB1的检出限为0.80 ng/mL,线性范围(IC20~IC80)为0.81~5.67 ng/mL,半抑制浓度(IC50)为2.15 ng/mL。在ZEN检出限为4.58 ng/mL,线性范围(IC20~IC80)为4.76~85.60 ng/mL,半抑制浓度(IC50)为20.19 ng/mL。方法特异性良好,与T-2毒素、脱氧雪腐镰刀菌烯醇、伏马毒素、赭曲霉毒素A多种真菌毒素交叉率小于10%。通过选择玉米、麦麸、大豆、小麦进行添加回收试验,AFB1的添加回收率在97.1%~108.7%之间,ZEN的添加回收率在92.8%~109.1%之间,相对标准偏差小于15%。选取经HPLC-MS/MS检测过的FAPAS标准质控样本进行测试,检测结果与其结果一致。在实际产品检测对比中,与市售胶体金免疫层析卡,ELISA试剂盒的检测结果基本一致。本方法操作简单快速、可定量,检测过程约25 min,适用于谷物样品中黄曲霉毒素B1和玉米赤霉烯酮的现场快速筛。     

IAC-HPLC法检测牛奶中黄曲霉毒素和玉米赤霉醇及类似物质量分数

建立了IAC-HPLC(免疫亲和柱净化-高效液相色谱)法检测牛奶中6种黄曲霉毒素和玉米赤霉醇及类似物的方法。样品经免疫亲和柱净化后,黄曲霉毒素用高效液相色谱——荧光检测器柱后衍生检测,玉米赤霉醇及其类似物用高效液相色谱——紫外检测器检测。结果表明,牛奶中黄曲霉毒素(M2,M1,G2,G1,B2,B1)的检测限分别为0.004,0.004,0.004,0.003,0.002,0.002μg/L,6种黄曲霉毒素的平均回收率在91.20%~113.8%之间,变异系数小于8.79%;玉米赤霉醇及其类似物的检测限均为0.05μg/L,平均回收率在54.22%~90.76%之间,变异系数小于9.44%。     

10种植物性饲料原料中3种霉菌毒素的污染分析和危害控制

通过对97份10种植物性饲料原料中黄曲霉毒素B1、玉米赤霉烯酮、呕吐毒素3种霉菌毒素的采样检测,分析了解3种霉菌毒素的污染状况,为饲料生产企业和养殖企业掌握植物性饲料原料中3种霉菌毒素的污染情况,减少霉菌毒素污染危害采取的应对措施提供参考。     

表面等离子共振技术快速检测食品中的 玉米赤霉烯酮毒素

目的建立基于表面等离子共振(surface plasmon resonance,SPR)技术快速检测食品中玉米赤霉烯酮毒素(zearalenone,ZEN)的方法。方法采用表面自组装技术(self-assembled monolayer,SAM)在金膜的表面修饰羧基基团,将ZEN抗原与牛血清白蛋白(albumin from bovine serum,BSA)偶联物(ZEN-BSA)通过共价键固定在芯片的表面,采用竞争法检测样品中的玉米赤霉烯酮毒素。结果该方法的检测限为8.2 ng/m L,ZEN单克隆抗体与呕吐毒素、黄曲霉毒素B_1、赭曲霉毒素、伏马毒素等没有交叉反应,与α-玉米赤霉烯醇和β-玉米赤霉烯醇交叉反应率分别为15.3%和11.5%。结论本方法具有简便、快速和高灵敏度等优势,在食品中真菌毒素的快速检测方面具有潜在的应用价值。     

竞争性酶联免疫法快速检测高产奶牛精料补充料中玉米赤霉烯酮

目的 建立竞争性酶联免疫方法检测饲料中玉米赤霉烯酮的分析方法。方法 样品经过60%的甲醇溶液提取, 提取液经过离心后, 取25 ?L提取液, 加入475 ?L稀释缓冲液混合后即为试样液, 取50 ?L试样液采用酶联免疫吸附方法进行检测。在450 nm波长处检测吸光度值。结果 本方法在3 h内完成了饲料中玉米赤霉烯酮含量的测定。在加标浓度为100、200和500 ?g/kg的加标水平下, 玉米赤霉烯酮酶在饲料中的加标回收率为99.5%~122.1%, 变异系数为1.4%~3.0%。结论 该方法快速、准确、灵敏, 适用于饲料中玉米赤霉烯酮的快速筛查检测。     

Reproducibility of a commercially available kit utilizing enzyme-linked immunosorbent assay for determination of aflatoxin in peanut butter

The reproducibility of a commercially available kit utilising enzyme-linked immunosorbent assay (ELISA) for determination of total aflatoxins has been analysed. Peanut butter samples contaminated with aflatoxin at three concentrations (mean 5.8, 11.7 and 24.8 ng/g) were analysed on different occasions using four kits. Coefficients of variation at all concentrations ranged from 2.6% to 13.6% within replicate, 0.3% to 16.2% within plate and 6.7% to 10.2% between kits. Standard curve precision profiles showed a minimum coefficient variation (CV) of 2-4% and a working range (below 10% CV) covering almost the entire standard curve.     

Aflatoxin B1, zearalenone and deoxynivalenol in feed ingredients and complete feed from central China

Between 2012 and 2014, 2528 feed ingredient and complete feed samples were collected from central China. Numbers of 2083, 255 and 190 samples were analysed for aflatoxin B1 (AFB1), zearalenone (ZEN) and deoxynivalenol (DON), respectively, by high-performance liquid chromatography in combination with UV or fluorescence detection. The incidence rates of AFB1, ZEN and DON contamination of feed ingredients and complete feeds were 33.9%, 90.2% and 77.4%, respectively. The percentage of positive samples for AFB1 ranged from 13.1% to 97.1%. Cottonseed meal presented the most serious contamination by AFB1. ZEN and DON contamination levels of feeds ranged from 50% to 100%, indicating serious contamination over the studied 3-year period. This study demonstrates that AFB1, ZEN and DON contamination of feeds in central China is serious and differs over the years. Feeds are mostly contaminated with ZEN, followed by DON and AFB1.     

Evaluation of the effect of mycotoxin binders in animal feed on the analytical performance of standardised methods for the determination of mycotoxins in feed

Recently, the use of substances that can suppress or reduce absorption, promote the excretion of mycotoxins or modify their mode of action in feed, so-called mycotoxin binders, has been officially allowed in the European Union as technological feed additives. The influence of the addition of mycotoxin binders to animal feed on the analytical performance of the official methods for the determination of mycotoxins was studied and the results are presented. Where possible standardised methods for analysis were applied. Samples of 20 commercial mycotoxin binders were collected from various companies. The following mycotoxins were included in the study: aflatoxin B₁, deoxynivalenol, zearalenone, ochratoxin A, fumonisins B₁ and B₂, T-2 and HT-2 toxins. A binder (or binders combined in a group) was mixed with feed material containing the mycotoxin, and the feed material was analysed. For data evaluation, the mean values were compared by Student's t-test (an independent two-sample t-test with unequal sample sizes and equal variance). The repeatability standard deviation of each method was used as an estimate of method variability. No significant differences (p?=?0.05) in mycotoxin levels between binder-free material and the material containing different binders were found. Further, the possible effects of binder addition in combination with processing (pelletising) on the amount of aflatoxin B₁ determined in feed were studied. Three commercial mycotoxin binders containing hydrated sodium calcium aluminosilicate (HSCAS) as the main component were used in these experiments. Feed samples with and without mycotoxin binders were pelletised with and without steam treatment. After pelletising, materials were analysed for AFB₁. Only the combination pelletising and a mixture of binders added at a total level of 1.2% had a significant effect (41% reduction) on the amount of AFB₁ determined.     

酶联免疫法测定食品和饲料中的黄曲霉毒素

本实验应用酶联免疫方法对食品和饲料中的黄曲霉毒素进行测定,奶粉中黄曲霉毒素M1的多次重复测定的结果的变异系数小于12%,回收率接近90%。说明本试剂盒测定结果比较精确,重现性较好。食品辅料及饲料中的黄曲霉毒素B1的测定结果的变异系数小于15%,说明实验结果的重现性较好。     

进口玉米酒糟粕中15种真菌毒素的检测和风险分析

建立了液相串联质谱检测玉米酒糟粕中15种真菌毒素的检测方法,并对结果分布进行了讨论。检测的真菌毒素包括黄曲霉毒素B1等15种真菌毒素。这些毒素采用溶剂提取,多功能净化柱净化,液相串联质谱检测。方法均经过优化和验证,满足进口玉米酒糟粕的检测要求。对67个进口玉米酒糟粕样品的检测结果表明,玉米赤霉烯酮、脱氧雪腐镰刀菌烯醇、T-2毒素、HT-2毒素等真菌毒素均有检出。本试验对这些数据进行了统计分析,提出了对进口玉米酒糟粕的真菌毒素风险分析结果。     

多厂家商品化黄曲霉毒素M1检测试剂盒质量评价与分析

目的对市场上11个厂家商品化黄曲霉毒素M_1快速检测试剂盒进行质量评价。方法定性检测试剂盒验证其灵敏度、特异性、假阳性率、假阴性率、符合率;定量试剂盒验证其范围和线性、准确度、精密度,并与实验室参考方法比较,分别对定性、定量试剂盒进行质量评价。结果 9个厂家定性试剂盒灵敏度、特异性及符合率均为100%,假阳性率及假阴性率均为0;厂家编号10的定量试剂盒准确度和精密度较差,厂家编号11的定量试剂盒线性、准确度及精密度较好,可以满足检测需求。结论黄曲霉毒素M_1快速检测试剂盒在定性检测方面较成熟,能够参与市场监督;定量试剂盒产品不够成熟,使用前应严格验证其指标。     

Mycotoxin contamination of sorghum and its contribution to human dietary exposure in four sub-Saharan countries

This research aimed at evaluating the safety, and the type, level and prevalence of mycotoxins in grain sorghum of four sub-Saharan African (SSA) countries (Burkina Faso, Ethiopia, Mali and Sudan). A multi-analyte LC-MS/MS method for quantification of 23 mycotoxins (nivalenol, deoxynivalenol, fusarenon X, neosolaniol, 3-acetyl deoxynivalenol, 15-acetyl deoxynivalenol, diacetoxyscirpenol, roquefortine C, HT-2 toxin, alternariol, T-2 toxin, FB1, FB2, FB3, zearalenone, aflatoxin G₁, aflatoxin G₂, aflatoxin B₁, aflatoxin B₂, sterigmatocystin, OTA, altenuene, alternariol monomethylether) was applied to different sorghum matrices. Of the 1533 analysed samples, 33% were contaminated with at least one of the following mycotoxins: aflatoxins, fumonisins, sterigmatocystin, Alternaria toxins, OTA and zearalenone. Country of origin, colour, source and collection period of sorghum samples significantly influenced the type, level and prevalence of mycotoxins. Sterigmatocystin (15%), fumonisins (17%) and aflatoxins (13%) were the most prevalent. FB1 (274 ± 585 µg/kg) had the highest mean concentration followed by FB2 (214 ± 308 µg/kg) while diacetoxyscirpenol (8.12 ± 19.2 µg/kg) and HT-2 (11.9 ± 0.00 µg/kg) had the lowest concentrations. Neosolaniol, fusarenon-X, 3-acetyl deoxynivalenol, 15-acetyl deoxynivalenol, T-2 toxin, nivalenol and roquefortine C were not detected in any of the samples. Sudan had the lowest prevalence and mean concentration of all mycotoxins. Pink sorghum had the highest concentrations of fumonisins and aflatoxins. Mycotoxins from Aspergillus spp. and Alternaria spp. are the mycotoxins of concern in SSA grain sorghum with regard to prevalence, concentration and possible health risk from exposure. Based on the performed risk characterisation, daily consumption of sorghum containing aflatoxins, alternariol, alternariol monomethyl ether, sterigmatocystin and OTA could result in exceeding the established health-based guidance values for these toxins.     

鲜食玉米籽粒发育期间污染真菌分离鉴定及真菌毒素含量分析

为探讨鲜食玉米籽粒发育期间真菌污染和真菌毒素积累情况,选择天紫23、京科糯2000和黑糯6号3?个品种鲜食玉米,在玉米授粉后第0、12、23、35、55天采样对污染真菌进行分离鉴定,并对黄曲霉毒素B1、呕吐毒素和玉米赤霉烯酮含量进行对比分析。结果从3?个品种鲜食玉米中,共分离出1?606?株真菌,经鉴定,隶属13?个属,20?个种;玉米籽粒发育期间主要污染真菌为青霉属（Penicillium）、曲霉属（Aspergillus）、镰刀菌属（Fusarium）、毛霉属（Mucor）和篮状菌属（Talaromyces）,其次是交链孢属（Alternaria）、根霉属（Rhizopus）、帚枝霉属（Sarocladium）、笄霉属（Choanephora）,还有少部分茎点霉（Phoma）、黑孢霉（Nigrospora）、横梗霉（Lichtheimia）和枝孢霉（Cladosporium）;鲜食玉米授粉后23?d（乳熟期）黄曲霉毒素B1、呕吐毒素和玉米赤霉烯酮的含量最低,是食用的最佳时期;污染真菌数量与毒素积累不具备相关性,黑糯6号品种受真菌毒素污染最轻,其次是天紫23和京科糯2000,但测试的鲜食玉米品种真菌毒素含量均远低于国家限量。     

Natural occurrence of zearalenone and trichothecene toxins in maize-based animal feeds in zambia

A year-long collection of maize-based animal feed samples from the National Milling Company and mouldy maize collected from farmers fields near Lusaka were analysed for Fusarium mycotoxins. In the survey, 148 samples were tested for zearalenone, deoxynivalenol and nivalenol, and 60 samples for T-2 toxin and diacetoxyscirpenol. Zearalenone was present up to 0.6 mg kg^?1 in 17% of the feed samples, and deoxynivalenol was found at I-0 mg kg^?1 in 1.4 % of these samples. This is the first report of these toxins in animal feeds in Zambia. Zearalenone was also found in 57.6 % of the 33 mouldy maize samples collected at levels ranging from 0.08 to 6.0 mg kg^?1 (mean concentration 1.11 mg kg^?1), and 49.5% of these samples contained deoxynivalenol at levels ranging from 0.5 to 16.0 mg kg^?1 (mean concentration 5.56 mg kg ^?1). T-2 toxin and diacetoxyscirpenol were not detected.     

Mycotoxin occurrence in feed and feed raw materials worldwide: long‐term analysis with special focus on Europe and Asia

During an 8‐year period, 17 316 samples of feed and feed raw materials from all over the world were analysed for contamination with aflatoxins, ochratoxin A, zearalenone, deoxynivalenol and fumonisins. Overall, 72% of the samples tested positive for at least one mycotoxin and 38% were found to be co‐contaminated. Mycotoxin concentrations were generally low and the majority of the samples were compliant with the most stringent EU guidance values or maximum levels for mycotoxins in feed. However, in their present state these regulations do not address co‐contamination and associated risks. Long‐term trends are difficult to establish as strong yearly variations were observed regarding mycotoxin prevalence and contamination levels. In some cases unusual weather conditions can be linked with high observed mycotoxin loads. An exception to this rule is South‐East Asia, where a steady increase of aflatoxin prevalence has been observed. The percentage of aflatoxin‐positive samples in this region rose from 32% in 2005 to 71% in 2011. © 2013 Society of Chemical Industry