化学发光酶免疫法检测猪肉中氯丙嗪残留

采用棋盘滴定法确定包被抗原浓度和抗体稀释倍数,通过单因素实验优化了竞争反应时间、磷酸盐缓冲液浓度、甲醇含量、pH值等参数,建立了氯丙嗪的间接竞争化学发光酶免疫检测方法,并考察了方法的特异性、灵敏度和稳定性.结果表明,最佳反应条件为包被抗原浓度为0.05 μg/L;氯丙嗪抗体稀释32000倍;体系缓冲液为含10％甲醇、0.1 mol/L磷酸盐缓冲液(pH 7.0).本方法的IC50为0.12 μg/L;检出限为0.02 μg/L;线性范围0.02～24.78 μg/L;批内和批间相对标准偏差均小于10％.与其它结构类似物没有明显交叉反应.猪肉检测的平均回收率为87.4％～105.6％,与高效液相色谱方法的相关性良好.本方法具有较高的灵敏度和较好的稳定性.     

Generation of anti-azoxystrobin monoclonal antibodies from regioisomeric haptens functionalized at selected sites and development of indirect competitive immunoassays

Azoxystrobin is a modern strobilurin fungicide used around the world to combat prime diseases affecting highly valuable crops. Accordingly, residues of this chemical are frequently found in food, even though mostly under maximum tolerated levels. We herein describe the development of an indirect competitive immunoassay for the determination of azoxystrobin residues. A panel of monoclonal antibodies displaying subnanomolar affinity to azoxystrobin was generated using, as immunizing haptens in mice, four functionalized derivatives carrying the same spacer arm located at different rationally chosen positions. This collection of antibodies was thoroughly characterized with homologous and heterologous antigens, and the immunoassay consisting of monoclonal antibody AZo6#49 and the coating conjugate OVA–AZb6, which displayed an IC₅₀ value of 0.102 μg L⁻¹ and a LOD of 0.017 μg L⁻¹, was eventually optimized. The response to different pH and ionic strength conditions of the specific assay was studied using a biparametric approach. In addition, the influence of Tween 20 and organic solvents over the assay parameters was also evaluated. After optimization, the developed immunochemical assay was applied to the analysis of azoxystrobin in spiked juices of relevant fruits and vegetables, showing excellent recoveries between 2 and 500 μg L⁻¹.     

Ultrasensitive aptamer-based bio bar code immunomagnetic separation and electrochemiluminescence method for the detection of protein

An ultrasensitive aptamer-based bio bar code immunomagnetic separation and electrochemiluminescence (IM-ECL) method for the detection of protein is developed. The target protein is captured by biotin-labeled aptamer (biotin probe) and [Ru(bpy)₃]²⁺ (TBR)-Au bio bar code-labeled aptamer (ECL nanoprobe), to form a double aptamer–protein sandwich complex. The complex is then immobilized on the streptavidin microbeads through biotin–streptavidin linkage and detected by ECL assay. The ECL signal of the target protein is amplified by the TBR-bio bar code DNAs. As an example, platelet-derived growth factor B-chain homodimer (PDGF-BB) was detected by the method. Experimental results show that the detection limit of the assay is 1 pM of PDGF-BB. A calibration curve with a linearity range from 1 pM to 10 nM is established, thus, make quantitative analysis possible. The method has been used to detect PDGF-BB in fetal calf serum with minimum background interference. Due to the wide availability of aptamer for numerous proteins, this aptamer-based bio bar code IM-ECL method holds great promise in protein detection.     

高效液相色谱柱后衍生测定鸡组织中甲基盐霉素残留量

建立了鸡组织中甲基盐霉素的高效液相色谱柱后衍生化分析方法.样品经异辛烷提取,离心后上层有机相过硅胶固相萃取小柱,洗脱液浓缩后用V(甲醇)∶ V(水)=90∶ 10混合液溶解.采用Inertsil ODS-3 C18柱,以V(甲醇): V(乙酸)∶ V(水)=94∶ 3∶ 3为流动相,香草醛为衍生剂进行高效液相色谱柱后衍生分析,520 nm检测,外标法定量.方法检出限为6 μg/kg; 定量限为20 μg/kg; 添加浓度在20～1800 μg/kg范围内,平均添加回收率为76.4%～93.1%; 批内相对标准偏差(RSD)在2.6%～8.9%之间; 批间相对标准偏差(RSD)在4.7%～9.7%之间.样品浓度在0.07～10.0 mg/L范围内与峰面积呈良好的线性关系,r>0.9993.     

液相色谱串联质谱法测定水果中复硝酚钠残留量

以甲醇提取样品,采用超高效液相色谱串联质谱法(UPLC–MS/MS)测定水果中复硝酚钠的残留量。以甲醇–10 mmol/L乙酸铵水溶液(体积比为60∶40)为流动相,质谱采用电喷雾负离子MRM检测模式。对硝基苯酚钠和5-硝基愈创木酚钠的线性范围为0.05~2.00 mg/L,检出限为0.01 mg/kg,邻硝基苯酚钠的线性范围为2.5~100.0 mg/L,检出限为0.5 mg/kg,线性相关系数均大于0.995。实际样品中对硝基苯酚钠、邻硝基苯酚钠和5-硝基愈创木酚钠的加标回收率分别为83.0%~93.4%,81.0%~87.4%,83.0%~91.8%,测定结果的相对标准偏差小于7%(n=6)。该法操作简单、快捷,精密度、准确度高,适用于水果中复硝酚钠的残留分析。     

Residue and dissipation of zinc thiazole in tobacco field ecosystem

A high-performance liquid chromatography with ultraviolet (HPLC-UV) detection method after derivatisation was developed for the first time for the novel fungicide zinc thiazole residue in tobacco samples. Field trials in two different locations were conducted to investigate the dissipation and residue of zinc thiazole in tobacco leaves and soil. The average recoveries of zinc thiazole were in the range of 82.5%–93.9% with relative standard deviations (RSDs) of 1.2%–9.1%. The zinc thiazole showed a rapid dissipation rate in fresh tobacco leaves with the half-lives of 1.1–1.6 days. The terminal residues of zinc thiazole in cured tobacco leaves and soil were 2.8–28.0 mg kg^?1and <0.05 mg kg^?1, respectively. The results could be used to establish the maximum residue limits (MRLs) and provide guidance for the scientific use of zinc thiazole in agriculture.     

Multiresidue determination of 160 pesticides in wines employing mixed-mode dispersive-solid phase extraction and gas chromatography-tandem mass spectrometry

A new multiresidue method for the efficient screening, identification and quantification of over 160 pesticides belonging to different chemical classes in red, rose and white wines have been developed. The analysis was based on gas chromatographic-tandem quadrupole mass spectrometric determination (GC-QqQ-MS/MS). An optimization strategy involved the selection of buffering conditions and sorbents for dispersive-solid phase extraction (dispersive-SPE) in order to achieve acceptably high recoveries and reduce co-extractives in the final extracts. As a result, the optimized procedure allowed us to obtain consistent recoveries of the target pesticides including problematic ones such as captan, chlorothalonil, dichlofluanid, folpet and tolylfluanid. The attained recoveries were typically between 80 and 110% (89% on average) with RSD values typically lower than 10% (8% on average) at three spiking levels of 0.01, 0.05 and 0.2 mg kg⁻¹. Linearity was studied in the range between 0.005 and 0.2 mg kg⁻¹ using pesticide standards prepared both in pure solvent and in the presence of matrix, showing coefficients of determination (R²) higher than 0.99 for all the pesticides except for desmedipham, thiabendazole and thiamethoxam in pure solvent. The study of the ratio of the slopes obtained in solvent and in matrix provided information about the matrix effects, which was <10%, 10-20% and >20% for 33, 36 and 31% of the studied pesticides, respectively. To improve accuracy, matrix matched standards were always used for calculation of the quantification results. The expanded uncertainties were estimated by using a “top-down” approach as being 17% on average (coverage factor k = 2, confidence level 95%). Finally, the method was used with success to detect and quantify pesticide residues in commercial wines.     

Analytical methods for pesticide residues in rice

Rice consumption has increased worldwide over recent decades, as it has become one of the most common foods. Although the analysis of environmental samples coming from rice areas has been well documented, there is less information regarding the analysis of pesticide residues in rice-grain samples.Rice (paddy, brown and white) can be considered a complex matrix, leading to difficulties in the application of the different multiresidue methods described in the literature. This review addresses and compares the principal extraction and clean-up methodologies [e.g., liquid-liquid extraction, solid-phase extraction, pressurized-liquid extraction, QuEChERS (quick, easy, cheap, effective, rugged and safe), gel-permeation chromatography and supercritical-fluid extraction - with QuEChERS-based methods being the most frequently employed].Traditionally, the determination of pesticide residues in rice has been based on gas chromatography with mass spectrometry (MS). But the application of new classes of pesticides has driven laboratories to increase the use of liquid chromatography with tandem MS. The limits of detection and quantification are in the ranges 0.09-90 μg/kg and 1-297 μg/kg, respectively, for the methodologies reported. These values agree with the current internationally-accepted maximum residue limits (MRLs).Based on the European Union (EU) database, more than 3000 analyses of pesticide residues in rice have been performed by official EU laboratories over the past decade. Of these, 6% reported pesticide residues above the MRLs.Physico-chemical properties can explain the occurrence of pesticides in rice commodities: lipophilic pesticides are frequently found in brown rice, whereas fungicides are mainly found in milled rice. Carbendazim, malathion, iprodione, tebuconazole, quinclorac and tricyclazole are the pesticides most frequently found in white rice, while buprofezin, hexaconazole, chlorpyrifos and edifenphos are most commonly found in paddy rice.Pesticide-residue concentrations can be affected during rice processing - with concentrations generally lower in the final products. However, few studies focusing on primary processing have addressed the setting of precise values applicable for the processing factors.     

Analytical methods applied to the determination of pesticide residues in foods of animal origin. A review of the past two decades

Pesticides are widely used in agriculture and can be transferred to animals in a number of ways. Consequently, reliable analytical methods are required to determine pesticide residues in foods of animal origin. The present review covers published methods and research articles (1990-2010) in which pesticide residues have been extracted from meat and meat products, milk and dairy products, fish and seafood, and eggs, then cleaned up, and isolated by chromatographic techniques to be identified and quantified by various detection methods. Recovery rates, quantification limits, the matrix effect and related parameters have all been considered. Lastly, future developments in this field are outlined.