磁性分子印迹聚合物提取-超高效液相色谱-串联质谱法测定乳及乳制品中的4 种伪蛋白

建立磁性分子印迹聚合物（magnetic molecular imprinted polymers,MMIPs）提取结合超高效液相色谱-串联质谱法测定乳及乳制品中4?种伪蛋白（三聚氰胺、环丙氨嗪、双氰胺和缩二脲）的方法。以缩二脲-13C2和环丙氨嗪-D4为印记分子,Fe3O4为磁性组分,制备具有特异识别性的MMIPs提取样品中的目标化合物,超高效液相色谱-串联质谱法检测。结果表明,4?种化合物在5～200?ng/mL范围内具有良好的线性关系,线性相关系数大于0.999。三聚氰胺、环丙氨嗪、双氰胺和缩二脲在牛奶样品中的检出限分别为0.10、0.02、0.05、0.50?μg/kg,在奶粉样品中的检出限分别为0.25、0.05、0.13、1.25?μg/kg。回收率为80.5%～96.1%,相对标准偏差为1.1%～9.2%。利用本方法对实际样品中4?种化合物进行测定,分析结果显示该方法所得到的数值准确、可靠。     

HPLC-MS/MS法快速测定乳制品中90种兽药的残留量

目的 建立高效液相色谱-串联质谱法（HPLC-MS/MS）测定乳制品中十大类90种兽药残留的检测方法。方法 试样用0.2%甲酸乙腈溶液提取,经Waters Prime HLB固相萃取柱净化,用Waters XBridge BEH-C18色谱柱洗脱分离,并使用电喷雾电离源,正负离子切换扫描模式进行检测,外标法定量。结果90种兽药在1 ng/mL～20 ng/mL范围内呈现良好的线性相关性,相关系数均大于0.995。90种兽药的方法检出限为0.3 ?g/kg,方法定量限为1 ?g/kg。在添加量为1 ?g/kg、2 ?g/kg和10 ?g/kg的加标回收实验下,90种兽药的加标回收率范围分别为66.0%～134.8%,64.0%～122.4%和62.4%～118.3%。同时做了方法精密度验证,有69%的兽药RSD<5%,有28%的兽药RSD在5%～10%,有3%的兽药RSD在10%～20%。结论 该方法简单、快速、高效且能同时准确定性和定量检测乳制品中十大类90种兽药的残留量。     

乳化/破乳法-气相色谱串联质谱法测定牛奶中12种亚硝胺残留

基于乳化/破乳法,本研究建立了一种气相色谱-串联质谱法定量检测牛奶中12种亚硝胺化合物含量的方法。对滤膜种类、提取剂种类以及提取次数等前处理条件的优化结果如下：滤膜种类为尼龙,提取剂为乙腈,提取次数为2次。样品经乙腈提取,蛋白沉淀,乳化/破乳法除脂后,采用HP-INNOWAX色谱柱分离,采用多反应离子监测（MRM）模式,内标法定量测定亚硝胺含量。结果表明,N-亚硝基吡咯烷（NPYR）的线性范围为2～100 ng/mL,其余亚硝胺化合物在1～100 ng/mL的浓度范围内呈现良好的线性关系,决定系数均大于0.995。在高、中、低3个水平加标试验中,回收率在90.90%～114.30%之间,日内精密度在1.76%～7.62%之间,日间精密度在2.75%～7.71%之间。NPYR的定量限为4μg/kg,其余化合物的定量限均为2μg/kg。方法的精密度与准确度良好,可满足牛奶中12种亚硝胺的准确定量需求。实际样品检测结果显示,6组牛奶样品中均检出了N-亚硝基二苯基胺（NDPhA）,含量在4.29～6.67μg/kg之间。     

超高效液相色谱-串联质谱法检测预包装畜禽肉制品中多种兽药残留

目的 建立预包装畜禽肉制品中54种兽药残留的超高效液相色谱-串联质谱检测方法。方法 样品经甲酸乙腈提取,Oasis PRIME HLB固相萃取柱净化,经Waters Acquity UPLC BEH C18色谱柱分离,以甲醇和0.1％甲酸水溶液为流动相进行梯度洗脱,电喷雾正离子(ESI+)模式电离,MRM-IDA-EPI扫描模式检测。结果 经方法学验证,54种化合物在0.5ng/mL～50 ng/mL范围内均呈现良好的线性关系,相关系数均?0.990,检出限0.5 μg/kg ～10 μg/kg,回收率为76.3%～92.4%,相对标准偏差为3.1%～15.2%。结论 本方法操作简便,回收率高。适用于预包装畜禽肉制品中54种兽药残留的同时定性、定量分析及确证。     

QuEChERS-超高效液相色谱-串联质谱法同时测定畜禽肉中11 种喹诺酮类兽药残留

建立一种超高效液相色谱-串联质谱法测定畜禽肉中恩诺沙星、环丙沙星、氧氟沙星、培氟沙星、诺氟沙星、洛美沙星、沙拉沙星、双氟沙星、司帕沙星、氟罗沙星和达氟沙星共11 种喹诺酮类兽药残留的检测方法。畜禽肉样品经2%甲酸-乙腈均质提取,离心沉淀后上清液经QuEChERS净化剂净化后,经快速溶剂蒸发系统浓缩,采用Waters BEH C18色谱柱（2.1 mm×50 mm,1.7 μm）分离,以0.1%甲酸-水和0.1%甲酸-乙腈为流动相进行梯度洗脱,电喷雾离子源电离,多反应监测模式采集信号,内标法定量。结果表明：11 种喹诺酮类兽药在2～80 ng/mL质量浓度范围内线性关系良好（R2＞0.99）,方法的检出限和定量限分别为0.5～2.0 μg/kg和1.5～6.0 μg/kg;空白样品加标量为10、25、50 μg/kg 3 个水平时,11 种喹诺酮类兽药的平均加标回收率为74.52%～106.83%,相对标准偏差为1.8%～8.7%（n=6）。该方法检测过程简单快捷、准确度好、灵敏度高,适用于畜禽肉中喹诺酮类兽药残留的检测。     

QuEChERS EMR Lipid净化结合同位素稀释-超高效液相色谱-串联质谱法同时测定畜肉中17种β-受体激动剂

目的：建立了一种采用QuEChERS EMR Lipid净化结合同位素稀释-超高效液相色谱-串联质谱技术同时测定畜肉中17种β-受体激动剂药物残留的检测方法。方法：样品加入pH为5.2的乙酸铵缓冲溶液,以β-葡萄糖醛苷酶/芳基硫酸酯酶酶解,经5%甲酸乙腈提取,QuEChERS EMR Lipid净化,采用CORTECS?UPLC?C18柱（3.0 mm×100 mm,1.6μm）,0.1%甲酸水和甲醇梯度洗脱,在分时段多反应监测（scheduled MRM,sMRM）模式下测定,内标法定量。结果：17种β-受体激动剂在0～20 ng/mL浓度范围内线性相关系数均大于0.99,方法检出限为0.01～0.09μg/kg,定量限为0.05～0.31μg/kg。在0.5、2.0、5.0μg/kg 3个加标浓度水平下的平均回收率为81.7%～111.8%,RSD为0.8%～10.3%(n=6)。100批市售生鲜畜肉中未检出β-受体激动剂。结论：该法前处理步骤简便,净化效果良好,缩短酶解时间,提高了样品检测效率,内标法定量精准可靠,适用于大批量畜肉样品中多种β-受体激动剂药物残留的同时检测。     

基于磁性免疫传感器的牛乳中四环素残留检测方法的构建

建立一种高灵敏磁性光学免疫传感检测方法,可用于生鲜乳中四环素残留的快速检测。方法的检出限为0.03 ng/mL,定量限为0.20 ng/mL;四环素质量浓度在0.25～32 ng/mL时,荧光信号值与质量浓度之间具有良好的线性关系,其回归方程决定系数（R2）为0.995 9;本方法准确度高,四环素不同添加水平条件下的回收率在82.6%～97.0%之间;该方法具有良好的重复性,测定结果的日内精密度和日间精密度均在5.0%以内;本方法与高效液相色谱-串联紫外检测结果高度吻合。     

不同前处理方法对猪组织中喹诺酮类兽药残留检测效果对比

采用超高效液相色谱-三重四极杆质谱和飞行时间质谱技术,比较EMR-L、Oasis PRiME HLB和液液萃取法对猪组织中16?种喹诺酮类兽药残留检测效果的影响。目标兽药以基质匹配工作曲线进行定量,在1.0～100.0?μg/L范围内线性关系良好,在5、10、20?μg/kg加标量条件下使用超高效液相色谱-串联质谱检测,EMR-L和Oasis?PRiME?HLB方法处理的猪组织中加标回收率（71.7%～106.9%、70.7%～118.3%）、相对标准偏差（0.2%～16.8%、0.1%～17.8%）、定量限（0.13～0.56、0.27～1.36?μg/kg）和检出限（0.04～0.23、0.08～0.41?μg/kg）均满足方法学要求。针对不同基质成分样品,采用不同的前处理技术减少假阳性结果的产生,以适用于日益增长的动物源性食品兽药残留检测的需求。本研究采用的2?种方法操作简便、灵敏度高、重复性好,满足对猪组织中多兽药残留的定性、定量分析。     

基于金纳米颗粒信号放大ELISA检测食品中恩诺沙星

以金纳米颗粒（gold nanoparticles,AuNPs）为信号放大载体,利用辣根过氧化物酶（horseradish peroxidase,HRP）标记的二抗为信号探针,建立一种基于AuNPs的信号放大酶联免疫检测方法（AuNPs signal amplification enzyme-linked immunosorbent assay,AuNPs-HRP-IgG ic-ELISA）,检测食品中恩诺沙星（enrofloxacin,ENR）。该方法对ENR的检测限（IC15）为5×10－4?ng/mL,半数抑制浓度（IC50）为0.24?ng/mL,检测范围为0.16～500?ng/mL,与建立的传统ELISA方法（IC50=8.76?ng/mL）相比,显著提高了检测灵敏度,并且该方法与环丙沙星、氧氟沙星、诺氟沙星交叉反应率均小于0.1%,具有良好的特异性。该AuNPs-HRP-IgG?ic-ELISA方法的实用性得到了样品添加回收实验和商业化ELISA检测试剂盒的验证,其在牛奶样品中的加标回收率可达80.52%～102.66%,适用于实际牛奶样本中ENR的快速灵敏检测,也为建立其他食品危害物质的精准检测技术开发提供参考。     

破乳诱导萃取-ICP-MS法测定食用油中镉的含量

建立破乳诱导萃取结合电感耦合等离子体质谱（inductively coupled plasma mass spectrometry,ICP-MS）法测定食用油中重金属镉含量的方法。优化关键实验参数（表面活性剂种类、表面活性剂质量浓度、硝酸体积分数以及食用油取样量）,应用优化的实验条件检测实际食用油样品。优化的实验条件为Triton?X-114作表面活性剂质量浓度0.09?g/mL、硝酸体积分数5%、食用油取样量2.0?g。该方法食用油中镉的检出限为0.031?ng/g,标准曲线的相关系数大于0.999?9,回收率在90.3%～105.0%之间。与传统的微波消解法对比,2?种方法得到的结果无显著性差异。此破乳诱导萃取法是一种简单、快速、绿色的样品前处理方法,结合ICP-MS可以实现食用油中镉的灵敏、准确检测。     

高分辨质谱定性筛查牛奶和奶粉中兽药残留

目的 建立高分辨质谱定性筛查牛奶和奶粉中兽药残留的高通量方法。方法 样品经乙腈、QuEChERS盐包提取, 采取改良的3种不同方式[QuChERS、高分辨质谱(high resolution mass spectrometry, EMR)、固相萃取(solid phase extraction, SPE)]净化提取液, 用液相色谱联用高分辨质谱(liquid chromatography-high resolution mass spectrometry, LC-HRMS)筛查了牛奶和奶粉中138种兽药残留, 建立兽药信息数据库, 并用截断值(Vcut-off, 判断阳性和阴性样品的临界值)作为评估筛查方法的指标。结果 改良的QuChERS可以获得更高的回收率, 当截断值(Vcut-off)设定为0.2时, 在2种加标水平下, 有109种兽药通过了筛查方法验证的要求, 假阳性和假阴性概率均低于5%。结论 此方法适用于牛奶和奶粉中多兽药残留的定性筛查, 一针进样可以得到109种兽药残留的筛查信息, 方法假阳性和假阴性概率在规定的目标物浓度下都低于5%。     

超高效液相色谱-串联质谱法测定牛乳中6种兽药残留

目的:建立高效液相色谱-串联质谱法(high performance liquid chromatography-tandem mass spectrometry,HPLC-MS/MS)测定牛奶样品中6种兽药残留(硝碘酚腈、碘醚柳胺、氯氰碘柳胺、托曲珠利、三氯苯达唑、水杨酸钠)的检测方法.方法:样品经过乙腈溶液提取,以...     

供港原料奶兽药残留危害及控制

讨论了原料奶中药物残留对人类健康以及乳制品加工造成的危害，介绍了中国大陆和香港两地关于原料奶中药物残留的相关规定，提出了控制供港原料奶中药物残留的5项措施，包括提高饲养和防疫管理水平、规范兽药管理、加强牛群管理、使用高效低毒的药物以及加强药残检测。     

Screening of veterinary drug residues in food by LC-MS/MS. Background and challenges

Regulatory agencies and government authorities have established maximum residue limits (MRL) in various food matrices of animal origin for supporting governments and food operators in the monitoring of veterinary drug residues in the food chain, and ultimately in the consumer’s plate. Today, about 200 veterinary drug residues from several families, mainly with antibiotic, antiparasitic or antiinflammatory activities, are regulated in a variety of food matrices such as milk, meat or egg. This article provides a review of the regulatory framework in milk and muscle including data from Codex Alimentarius, Europe, the U.S.A., Canada and China for about 220 veterinary drugs. The article also provides a comprehensive overview of the challenge for food control, and emphasizes the pivotal role of liquid chromatography-mass spectrometry (LC-MS), either in tandem with quadrupoles (LC-MS/MS) or high resolution MS (LC-HRMS), for ensuring an adequate consumer protection combined with an affordable cost. The capability of a streamlined LC-MS/MS platform for screening 152 veterinary drug residues in a broad range of raw materials and finished products is highlighted in a production line perspective. The rationale for a suite of four methods intended to achieve appropriate performance in terms of scope and sensitivity is presented. Overall, the platform encompasses one stream for the determination of 105 compounds in a run (based on acidic QuEChERS-like), plus two streams for 23 β-lactams (alkaline QuEChERS-like) and 10 tetracyclines (low-temperature partitioning), respectively, and a dedicated stream for 14 aminoglycosides (molecularly-imprinted polymer).     

Development of a multi-class method for the identification and quantification of residues of antibiotics, coccidiostats and corticosteroids in milk by liquid chromatography-tandem mass spectrometry

Even if coccidiostats are not generally employed in dairy cows, maximum levels for these substances in milk samples have been set up in Commission Regulation 124/2009. Due to public health concerns regarding the presence of veterinary drug residues in milk, the aim of the present work is to report a multi-class high performance liquid chromatography-tandem mass spectrometry method for the simultaneous extraction, identification and quantification of seven coccidiostats, seven antimicrobial agents, three corticosteroids and an antifungal agent. Validation was conducted following the guidelines established in the European Commission Decision 2002/657/CE at the maximum concentration permitted in milk by the European Union for each drug. The method was successfully applied in 100 raw milk samples collected from ten local dairy farms and 15 milk samples bought in supermarkets. This method could be applied in routine analysis of milk samples by the dairy industry, replacing the current, non-differentiating, screening method.     

Multiresidue method for pesticides and veterinary drugs in bovine milk using GC/MS and LC/MS/MS

A simple, sensitive and selective method with gas chromatography/mass spectrometry (GC/MS) and liquid chromatography/tandem mass spectrometry (LC/MS/MS) has been developed to detect 342 pesticides and veterinary drugs contaminating bovine milk at the maximum residue limits (MRLs) defined in the "positive list system". Sample preparation was performed by extracting the analytes with acetonitrile, followed by salting-out with sodium chloride. For some pesticides, the extract was further cleaned up by n-hexane partitioning and PSA cartridge column chromatography. GC/MS-EI or -NCI was used to determine pesticide residues, while LC/MS/MS-ESI was applicable to the determination of pesticide and veterinary drug residues. The variation of the recoveries of these drugs at MRL was relatively wide; however the relative standard deviations of the recovery of each drug were within 28%, suggesting that the present method is good enough for use as a screening test for contaminants at the MRLs. These results show that this method is useful for multiresidue analysis of numerous pesticides and veterinary drugs in bovine milk.     

超高效液相色谱-四极杆/静电场轨道阱高分辨质谱法测定动源性食品中33种兽药残留及质谱库的建立

目的建立超高效液相色谱-四极杆/静电场轨道阱高分辨质谱法测定动源性食品中33种兽药残留的检测方法。方法采用乙腈-水(80:20, V:V)为提取溶剂, QuEChERS方法净化,在Full MS-ddMS2正离子扫描模式下,快速对动源性食品中的目标化合物进行检测。结果在r=70000,扫描范围80~1000m/z内,33种目标化合物的线性相关系数均大于0.99,基质加标回收率范围为70.4%~97.3%,相对标准偏差(relativestandard deviation, RSD)为2.1%~6.0%(n=6)。结论建立的方法适用于动源性食品中33种兽药残留快速检测。同时,本研究建立的33种兽药高分辨质谱库,可应用于非目标化合物的定性筛查。     

Analysis of antibiotic residues in raw and commercial milk in Punjab,India vis‐à‐vis human health risk assessment

The present study quantitatively analyzed the occurrence of antibiotic residues in 524 milk samples (492 raw milk samples from dairy farms and 32 commercial milk samples) from Punjab, India using high‐performance liquid chromatography (HPLC). Tetracyclines, sulphonamides, fluoroquinolones (commonly used in veterinary practice in Punjab), and chloramphenicol (banned in food animals in India) were targeted and analyzed using multiresidue HPLC method. Out of 492 dairy farms milk samples, 78 (16%) were found to be positive for antibiotic residues with 20 (4%) exceeded the maximum residue limits (MRLs). Among 32 commercial milk samples, 4 (12.5%) were positive for antibiotic residues with one (3.1%) sample violated the MRL. Assessment of human health risks revealed that the current levels of antibiotic residues in milk pose no significant toxicological effects on the health of consumers. However, the results highlighted an immediate need of effective strategies for prevention of nonprudent use of antibiotics in veterinary practice for safeguarding consumer's health.     

Multiclass methods for the analysis of antibiotic residues in milk by liquid chromatography coupled to mass spectrometry: A review

Milk is an important and beneficial food from a nutritional point of view, being an indispensable source of high quality proteins. Furthermore, it is a raw material for many dairy products, such as yoghurt, cheese, cream etc. Before reaching consumers, milk goes through production, processing and circulation. Each step involves potentially unsafe factors, such as chemical contamination that can affect milk quality. Antibiotics are widely used in veterinary medicine for dry cow therapy and mastitis treatment in lactating cows, which can cause the presence of antimicrobial residues in milk. In order to ensure consumers’ safety, milk is analyzed to make sure that the fixed Maximum Residue Limits (MRLs) for antibiotics are not exceeded. Multiclass methods can monitor more drug classes through a single analysis, so they are faster, less time-consuming and cheaper than traditional methods (single-class); this aspect is particularly important for milk, which is a highly perishable food. Nevertheless, multiclass methods for veterinary drug residues in foodstuffs are real analytical challenges. This article reviews the major multiclass methods published for the determination of antibiotic residues in milk by liquid chromatography coupled to mass spectrometry, with a special focus on sample preparation approaches.     

QuEChERS在动物源性食品兽药残留检测中的研究进展

QuEChERS是基于分散固相萃取建立起来的一种农药多残留快速(quick)、简单(easy)、便宜(cheap)、高效(effective)、耐用(rugged)和安全(safe)的分析方法。其基本流程为采用乙腈或含体积分数1%乙酸的乙腈提取,加入无水硫酸镁和氯化钠吸水并促使提取液分层,上清液加入硅胶基伯胺仲胺键合相吸附剂(PSA)、C18或石墨化炭黑(GCB)等吸附剂除去基质干扰物,最后用气-质联用(GC-MS)或液-质联用(LC-MS/MS)分析测定。在兽药残留分析中,QuEChERS主要用于牛奶、动物源性食品肌肉或肝脏组织中磺胺类、喹诺酮类、苯并咪唑类等兽药的分析。本文对QuEChERS在兽药残留领域的应用进展做一综述,以期为兽药残留分析方法及思路提供一定的参考。