高效液相色谱-串联质谱法测定奶粉中的双氰胺与三聚氰胺

建立了同时测定奶粉中双氰胺与三聚氰胺含量的高效液相色谱-串联质谱法。采用CAPCELL PAK CR 1:4色谱柱(2.0 mm i.d×150 mm,5μm),V(乙腈):V(CH3COONH4溶液,2 mmol/L,用乙酸调pH=4)=95:5为流动相,流速0.3 mL/min,柱温35℃,质谱检测器进行检测,外标法定量。在上述条件下,两种目标化合物在10~500 ng/mL范围内,质量浓度与定量离子峰面积呈良好的线性关系。对奶粉样品进行添加回收,双氰胺与三聚氰胺的平均回收率分别为93.4%~97.0%,92.7%~98.1%,相对标准偏差(RSD)分别为3.7%~5.8%,3.4%~5.7%。该方法可用于奶粉样品中双氰胺与三聚氰胺含量的分析。     

化学发光法测定奶粉中的三聚氰胺

在碱性条件下,三聚氰胺对鲁米诺-铁氰化钾体系化学发光反应具有明显的增敏作用;据此建立了流动注射化学发光测定奶粉中三聚氰胺的分析方法,确定了优化测定条件,并将该方法用于测定奶粉中的三聚氰胺.结果表明,在优化测定条件下,三聚氰胺的质量浓度在1.0×10-6～4.0×10-5 g.L-1范围内与发光强度呈现良好的线性关系,检出限为2.6×10-7 g.L-1,11次平行测定4.0×10-6 g.L-1三聚氰胺的相对标准偏差为3.42%.与此同时,将所建立的方法用于奶粉中三聚氰胺的测定,结果令人满意.     

液相色谱-串联质谱联用法测定食品中三聚氰胺残留量

提出了液相色谱-串联质谱联用法测定动物源性食品中三聚氰胺残留量的方法。样品采用乙腈进行提取,饱和氯化钠溶液进行分配经反相色谱分离后,串联质谱法进行测定,同位素内标法进行定量,方法的测定下限(10S/N)为0.05 mg·kg~(-1),以5类食品样品为基体,加入4种不同浓度水平的三聚氰胺标准作回收试验,测得回收率在72.9$～104.1%之间,测定值的相对标准偏差(n=6)在4.7%～15.2%之间。     

超高效液相色谱-串联质谱法同时测定牛奶和奶粉中的二氰二胺、三聚氰胺和三聚氰酸

建立了一种超高效液相色谱-串联质谱法测定牛奶和奶粉中二氰二胺(DCD)、三聚氰胺(MEL)和三聚氰酸(CY)的分析方法。样品用5%(质量分数)三氯乙酸水溶液提取,无需净化和浓缩,用Acquity UPLC HSS T3色谱柱(50 mm×2.1 mm, 1.8 μm)分离,以乙腈和乙酸铵作为流动相进行梯度洗脱,电喷雾正、负离子(ESI^±)模式同时电离,多反应监测模式进行检测。结果表明:在5.0～200.0 μg/L范围内线性关系良好(r≥0.995)。牛奶中3种化合物在0.02、0.10、0.20 mg/kg的添加回收率为60.0%～105.8%,相对标准偏差(RSD, n=6)为4.2%～13.6%;奶粉中3种化合物在0.05、0.10、0.20 mg/kg的添加回收率为78.0%～115.0%, RSD (n=6)为2.7%～7.5%。牛奶中方法的定量限(信噪比为10)为0.02 mg/kg,奶粉中方法的定量限为0.05 mg/kg。该方法简单、快速、灵敏、准确,适合于牛奶、奶粉等乳及乳制品中DCD、MEL和CY的快速、高灵敏的分析检测。     

高效毛细管电泳浊度法检测牛奶及奶粉中的三聚氰胺

建立高效毛细管电泳(HPCE)浊度法检测牛奶和奶粉中三聚氰胺的方法。样品中加入三氯乙酸水溶液,加热样品至沸腾后自然冷却,使蛋白质充分凝聚、沉降,并提取三聚氰胺。色谱条件为:毛细管柱长度50cm、内径75μm,分离电压13kV,进样量12.3kPa×3s,分离温度25℃,检测波长236nm。加标回收率为83%～98%之间,定量限为1mg/kg。测量结果的相对标准偏差为1.8%～3.4%(n=5)。该法可用于大量牛奶和奶粉样品中三聚氰胺的快速检测。     

亲水作用色谱-串联质谱测定蔬菜中灭蝇胺及其代谢物三聚氰胺

建立了亲水作用色谱-串联质谱测定蔬菜中灭蝇胺及其代谢物三聚氰胺的方法.蔬菜样品匀浆后经甲醇-水提取,取适量酸化后提取液经阳离子固相萃取柱净化,洗脱液用氮气吹干,残留物用5 mL流动相定容.采用亲水作用色谱(Hilic)分离,在电喷雾-选择反应监测模式下,进行定性和定量分析.以基质校正曲线计算,添加浓度为0.04和0.20 mg/kg时,灭蝇胺的回收率为84.2%～101.3%; 相对标准偏差(RSD)为4.5%～10.7%;三聚氰胺的回收率为72.5%～97.1%; RSD为4.3%～10.8%;灭蝇胺和三聚氰胺的定量限分别为0.01和0.005 mg/kg.利用本方法检测了多种蔬菜样品中灭蝇胺、三聚氰胺的含量.     

液相色谱-串联质谱法快速同时测定婴幼儿配方乳粉中氯霉素、三聚氰胺、甲硝唑和洛硝达唑

建立了婴幼儿配方乳粉中三聚氰胺、氯霉素、甲硝唑和洛硝达唑残留的高效液相色谱-串联四极杆质谱联用测定方法.样品经碱性乙腈提取,以MGⅢ-C18色谱柱(150 mm× 2.1 mm,5 μm)分离,流动相为60 mmol/L乙酸铵溶液(含0.4‰甲酸)和甲醇,梯度洗脱,流速为0.25 mL/min.采用多反应监测正离子或负离子模式,可以同时对婴幼儿配方乳粉中的三聚氰胺、氯霉素、甲硝唑和洛硝达唑进行快速定性和定量测定.在优化条件下,4种化合物检出限为0.3 ～ 50.0μg/kg,方法回收率为61.4％ ～ 96.2％,相对标准偏差(RSD)为1.8％ ～9.8％.采用本方法测定了多种市售婴幼儿配方奶粉.本方法操作简单、测定结果准确,可用于婴幼儿配方奶粉中三聚氰胺、氯霉素、甲硝唑和洛硝达唑残留的同时快速测定.     

气相色谱-质谱法快速测定牛奶中的三聚氰胺和三聚氰酸

建立了气相色谱-质谱法测定牛奶中三聚氰胺和三聚氰酸含量的分析方法。样品经二乙胺-乙腈-水溶液超声提取,离心,滤液用氮气吹干后加入硅烷化衍生试剂衍生,气相色谱-质谱测定,选择离子模式监测,外标法定量。在0.025～2 mg/kg范围内,目标物的峰面积与其质量浓度的线性关系良好(r>0.98);在0.5,1.0和2.5 mg/kg的添加水平,三聚氰胺和三聚氰酸的平均回收率分别为84%～87%和75%～102%,相对标准偏差(RSD)分别为5.7%～11.7%和4.9%～7.8%;三聚氰胺和三聚氰酸的检出限(LOD)分别为0.05 mg/kg和0.10 mg/kg。结果表明:该方法简便、快速、灵敏、准确,适合牛奶中三聚氰胺和三聚氰酸的确证和定量测定。     

表面解吸常压化学电离质谱法直接测定火锅底料中的痕量可卡因

采用表面解吸常压化学电离(SDAPCI)质谱法, 在无需样品预处理情况下, 直接测定了火锅底料中可卡因的含量. 采用取样针取样, 单次取样量在纳升级, 单个样品测定时间少于0.5 min, 回收率为92.9%～106.6%; 相对标准偏差(RSD)为4.7%～11.6%; 可卡因的检出限可达1.5×10-12 g/mL. 该方法适用于批量火锅底料等食品类样品的快速半定量检测.     

胶束增敏分光光度法水相测定乳制品中三聚氰胺

建立了一种基于胶束增敏缔合反应水相测定乳制品中三聚氰胺的分光光度法,研究了在阴离子表面活性剂十二烷基硫酸钠(SDS)形成的胶束体系中,茜素红(AR)阴离子与三聚氰胺阳离子反应产物的紫外-可见光谱特性。实验发现,加入三聚氰胺后,体系λmax由426nm红移到516nm,加入SDS对缔合物有显著的增稳、增敏效应。三聚氰胺质量浓度在1.9～25mg/L范围内符合比尔定律,缔合物的表观摩尔吸光系数为6.9×103L.mol-1.cm-1。本方法用于液态奶及奶粉中三聚氰胺的测定,回收率分别为94.5%～104.8%和95.5%～100.4%,相对标准偏差分别为1.6%～2.2%和1.7%～2.6%。考察了酸效应、胶束对体系的作用机理,发现三聚氰胺与AR缔合物之间主要是通过静电引力和疏水作用力相结合。     

亲水作用色谱-电喷雾串联质谱法检测原料奶及奶制品中的三聚氰胺

建立了亲水作用色谱-电喷雾串联质谱测定原料奶及奶制品中三聚氰胺的方法。样品采用1%三氯乙酸水溶液-乙腈（体积比为1∶1）混合溶液提取,混合型阳离子交换反相固相萃取柱（MCX）富集净化,亲水作用色谱柱分离,电喷雾串联四极杆质谱仪进行检测。结果表明,三聚氰胺的质量浓度在0.05～10.0 mg/L范围内具有良好的线性关系。原料奶及奶制品中的三聚氰胺在0.5,2.5和10 mg/kg 3个添加水平下,平均回收率为76.3%～98.7%,相对标准偏差均小于6.8%;定量限（S/N＞10）为0.05 mg/kg。     

亲水作用色谱-串联质谱法同时测定液态奶中三聚氰酸和三聚氰胺

建立了亲水作用色谱-串联质谱同时测定液态奶中三聚氰酸和三聚氰胺的方法。液态奶样品经体积分数2.5%甲酸溶液提取、离心后乙腈稀释,亲水作用色谱柱分离,电喷雾串联四极杆质谱检测器检测,分别在负、正离子模式下测定三聚氰酸和三聚氰胺。三聚氰酸和三聚氰胺分别在0.5～100μg/L、0.1～50μg/L范围内线性关系良好。在0.25～15mg/kg、0.1～7.5mg/kg添加水平范围内,三聚氰酸平均回收率为84.5%～98.0%(RSD为2.1%～6.1%),三聚氰胺平均回收率为85.5%～88.9%(RSD为3.2～5.8%)。三聚氰酸、三聚氰胺定量限分别为0.25mg/kg、0.1mg/kg。     

气相色谱-串联质谱法测定食品中的三聚氰胺

建立了使用气相色谱-串联质谱(GC-MS/MS)测定食品中三聚氰胺含量的分析方法。利用三氯乙酸超声萃取样品中的三聚氰胺,经乙酸锌溶液沉淀蛋白质后,将提取液离心、过滤、净化、衍生后采用GC-MS/MS的多反应监测模式（MRM）进行检测。方法的重现性良好,6次重复测定的峰面积的相对标准偏差小于9%;最低定量检测限（S/N=10）为1 μg/kg,在0.1~100 μg/L范围内具有良好的线性关系。应用所建立的方法测定了奶粉、奶糖、含乳饮料、饼干等食品中的三聚氰胺含量,为三聚氰胺滥用的检测和判断提供了方法。     

Development of melamine certified reference material in milk using two different isotope dilution mass spectrometry techniques

A new certified reference material (CRM) of melamine in milk GLHK-11-02 was developed aiming to address the great demand from the testing community after the melamine crises. The material was prepared by adding an appropriate quantity of melamine into the skimmed milk samples and the final product was in the form of fine lyophilized powder. Characterization of the material relied on two newly developed gravimetric isotope dilution mass spectrometry (IDMS) methods, one using liquid chromatography-tandem mass spectrometry (LC-MS/MS) and another gas chromatography-mass spectrometry (GC-MS). Experimental parameters with crucial effects on the performance of the two IDMS methods were thoroughly investigated. These included purity of standard used, equilibration time of isotopes, efficiency of extraction methods as well as possible interferences from the matrix and melamine analogues. Precision was found to be excellent with a coefficient of variation of 2.5% for the LC-IDMS/MS (n=46) and 1.9% for the GC-IDMS (n=30) respectively. Using one-tail Student's t-test at 95% confidence interval, analytical data sets generated from the two methods were found to exhibit no significant difference. Measurement accuracy of the methods was further verified through an Asia Pacific Metrology Program (APMP) pilot study. Analytical results of the present LC-IDMS/MS for the two milk test samples at the concentration level of about 0.45 and 3.5 mg kg(-1) were proven to be very good. There were excellent overlaps between our results and the assigned reference values, and the absolute deviation was less than 3.2%. Both the LC-IDMS/MS and GC-IDMS methods were shown to be sufficiently reliable and accurate for certification of the melamine CRM. Certified value of melamine in dry mass fraction in GLHK-11-02 was 1.14 mg kg(-1). Expanded uncertainty due to sample inhomogeneity, long term and short term stability and variability in the characterization procedure was at 7.1% or 0.08 mg kg(-1). The CRM is primarily used to provide a complete method validation for and to improve the technical competence of melamine analysis to food and chemical testing laboratories.     

Analysis of melamine in milk powder by using a magnetic molecularly imprinted polymer based on carbon nanotubes with ultra high performance liquid chromatography and tandem mass spectrometry

A new magnetic molecularly imprinted polymer was coupled with ultra high performance liquid chromatography and tandem mass spectrometry for the selective determination of melamine in milk powder. The magnetic molecularly imprinted polymer has been prepared by using carbon nanotubes as the matrix, Fe₃O₄ particles as the magnetic ingredient, melamine as the template molecule, methacrylic acid as the functional monomer, ethylene glycol dimethacrylate as the cross‐linker and polyvinylpyrrolidone as the dispersant. The polymer was characterized with scanning electron microscopy, Fourier transform infrared spectroscopy and a physical property measurement system. The isothermal adsorption, kinetics adsorption, and selectivity were studied to evaluate the rebinding properties of the magnetic molecularly imprinted polymer. Various parameters affecting the extraction efficiency such as the amount of polymer, extraction time, and eluting solution were evaluated. The limit of detection was 0.00075 mg/kg. The relative standard deviations of the intraday and interday precision are 0.4–2.7 and 2.3–5.1%, respectively. The proposed method was successfully applied to determine melamine in different milk powder samples from different provenances, and satisfactory recoveries of 89.0–95.6% were obtained. This method has great significance for quality control and is simple and suitable for the rapid determination of melamine in milk powder.     

Determination of melamine and cyanuric acid in powdered milk using injection-port derivatization and gas chromatography–tandem mass spectrometry with furan chemical ionization

A reliable, sensitive and eco-friendly injection-port trimethylsilylated (TMS) derivatization and gas chromatography–tandem mass spectrometry (GC–MS/MS) with furan chemical ionization (furan-CI) method was developed to determine melamine and cyanuric acid in powdered milk samples. The effects of several parameters related to the TMS-derivatization process (i.e., injection-port temperature, residence time and volume of silylating agent) and of various CI agents were investigated. Addition of a solution (3 μL) of bis(trimethyl)silyltrifluoroacetamide (BSTFA) containing 1% trimethylchlorosilane (TMCS) reagent to a 20-μL extract from the powdered milk sample gave an excellent yield of the tris-TMS-derivatives of melamine and cyanuric acid at an injection-port temperature of 90 °C. Furthermore, using furan as the CI agent in conjunction with tandem mass spectrometry provided the greatest sensitivity and selectivity of detection. The limits of quantitation (LOQs) for melamine and cyanuric acid were 0.5 and 1.0 ng/g in 0.5-g of powdered milk samples, respectively. The recoveries from spiked samples – after simple ultra-sonication with 5% dimethyl sulfoxide in acetonitrile coupled with n-hexane liquid–liquid extraction – ranged from 72% to 93% with relative standard deviations of lower than or equal to 18%. In three of four real powdered milk samples, melamine was detected at concentrations ranging from 36 to 1460 ng/g; and cyanuric acid was detected in two of these samples at concentrations of 17 and 180 ng/g.     

胶体金免疫层析法快速检测三聚氰胺

采用戊二醛法制备三聚氰胺-BSA偶联抗原,胶体金标记三聚氰胺单克隆抗体,建立了检测三聚氰胺的胶体金免疫层析试验。结果显示,该试验具有良好的敏感性,胶体金免疫层析试验对鲜奶、奶粉和饲料样品中三聚氰胺的最低检测量分别为1.0、2.0和2.5μg/g,该法适合现场快速检测三聚氰胺。     

Monitoring of melamine contamination in fat watery milk by the photoluminescence analysis

A photoluminescence method to detect the toxic melamine contamination in fat watery milk has been proposed. Despite the quite different luminescence origins of milk and melamine patterns, their wide emission spectra under UV excitation are similar and in the range of 2.2-3.5 eV. The complex milk photoluminescence spectrum composed of riboflavin, furosine, lactulose, Vitamin E and tryptophan emitting species can be modified if milk pattern is undergone by acid treatment (for example, in vinegar). At the same time the melamine emission is not subjected to any modification in vinegar. It allows quantitatively discriminating the melamine contamination in milk in linear range, at least, 0.05-7 g/l from different photoluminescence spectra of milk (water) with and without melamine. Limit of melamine detection achieves 0.01 g/l.     

Detection of Melamine in Powdered Milk Using Surface-Enhanced Raman Scattering with No Pretreatment

We detected a trace amount of melamine in powdered milk using surface enhanced Raman scattering (SERS) on gold surfaces at an excitation wavelength of 632.8 nm. A detection limit of ～100 ppm (μg/g) melamine in milk was attained within a few minutes by the gold nanoparticle substrate from chemical reduction; whereas, better sensitivity, as low as ～200 ppb (ng/g), was achieved by the roughened gold substrate. Our method has the advantage of fast and sensitive detection of melamine in powdered milk without pre-treating the samples.     

Fast quantitative determination of melamine and its derivatives by matrix-assisted laser desorption/ionization time-of-flight mass spectrometry

A simple, sensitive and fast method for the determination of melamine and its derivatives in milk powder using matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF-MS) was developed. Neither time-consuming sample preparation, nor special target plates, or other extra equipment are necessary. The common matrix sinapinic acid (SA) was used with a dried-droplet preparation. Detection limits (signal-to-noise (S/N) ratio = 3) for standard solutions of melamine, ammeline and cyanuric acid were 10, 25 and 10 μg/L, respectively. The limit of quantification (LOQ) for melamine was 25 μg/L and excellent linearity (R(2): 0.9990) was maintained over the range of 10-2000 μg/L. Ammeline and cyanuric acid were analyzed with an LOQ of 50 μg/L and also excellent linearity (R(2): 0.9997 and R(2): 0.9998). Good accuracy and precision were obtained for all concentrations within the range of the standard curve. The developed method was successfully used for the determination of melamine, ammeline and cyanuric acid in milk powder samples with a simple sample preparation. The LOQ of melamine was 0.5 μg/g. Ammeline and cyanuric acid were detectable at 0.5 and 5 μg/g. This method showed excellent accuracy, precision and linearity and significantly reduces the needed analysis time, as only approximately 10 s/sample measuring time is required. To the authors' knowledge, this is the first published method to quantify melamine and derivatives by MALDI-TOF-MS.