表面增强拉曼光谱快速检测生鲜肉中的瘦肉精

为了快速检测生鲜肉中的瘦肉精,该研究利用表面增强拉曼光谱技术,以沙丁胺醇为检测目标物,建立了一种快速检测肌肉组织和肝脏中瘦肉精含量的方法。在碱性环境下利用乙酸乙酯对样品中沙丁胺醇进行提取,采用Savitzky-Golay 5点平滑法和自适应迭代重加权惩罚最小二乘法消除光谱噪声以及荧光背景对分析建模的影响。为检测方法的重复性,对50个相同沙丁胺醇质量分数(1 mg/kg)的肌肉组织样品进行信号采集,对沙丁胺醇特征峰强度进行分析,621、814、1 253、1 489、1 609 cm~(-1) 5个特征峰强度的相对标准偏差(RSD)为6.54%、6.07%、8.65%、7.44%、6.81%,说明该方法具有较好的重复性。建立沙丁胺醇标准溶液的预测模型,沙丁胺醇浓度与其特征峰强度相关性较好,决定系数R~2为0.968。对肌肉组织和肝脏中沙丁胺醇含量进行检测,检测范围分别为0.01~5和0.02~5 mg/kg,检出限分别为0.01和0.02 mg/kg,其含量与预测实测值决定系数为0.912和0.921。研究表明,该方法可以实现肌肉组织和肝脏中沙丁胺醇含量的定量预测。     

饲料中沙丁胺醇快速检测技术研究

本实验在现有的克伦特罗检测系统上建立了沙丁胺醇的酶联免疫吸附检测体系。结果表明,该方法在饲料中沙丁胺醇的检测中完全可行。     

一种新的沙丁胺醇人工抗原的合成及单克隆抗体的制备

【目的】为证明采用4-溴丁酸乙酯半抗原合成人工抗原可制备灵敏度高、特异性好的单克隆抗体.【方法】将硫酸沙丁胺醇(SAL)与4-溴丁酸乙酯反应制备半抗原(SAL-HS);通过N-羟基琥珀酰亚胺活化酯法合成免疫原与包被原,用聚丙烯酰胺凝胶电泳、紫外光谱和质谱方法对沙丁胺醇免疫原及包被原进行鉴定.免疫原免疫Balb/c小鼠,采用融合技术产生杂交瘤细胞株,体内诱生腹水法制备单克隆抗体,并鉴定其免疫学特性.【结果和结论】筛选出1株杂交瘤细胞株,其细胞培养上清液效价达1∶128 000,腹水效价达1∶2 560 000,免疫球蛋白亚型鉴定为IGg1;抗体对沙丁胺醇的半数抑制浓度(IC50)为:0.746 ng/mL;与其他同类结构药物的交叉反应率均小于0.015%.     

Study on Preparation Treatment for Salbutamol in Feed by Enzyme-linked Immunosorbent Assay

Objective of the study was to develop a preparation treatment for salbutamol in feed by enzyme-linked immunosorbent assay (ELISA).The assay optimized the influence factor of the preparation and used to extract salbutamol residues in feed.The results showed that the inhibition concentration (IC₅₀) for the developed ELISA was 0.606 ng/mL with the detection range of 0.221 to 1.658 ng/mL and R² was 0.9998.After optimization,pH 7.5,0.06 mol/L PBS with the dilution ratio of 10 was selected to extract salbutamol residues in feed and the limit of detection (LOD) was 5.0 ng/mL in feed.The recoveries ranged from 77% to 110% for the spiked samples (5 and 10 μg/kg),and RSD was less than 8%.     

免疫亲和色谱-气相色谱/质谱法(IAC-GC/MS)检测猪肝中的沙丁胺醇与克伦特罗

制备出抗沙丁胺醇的抗血清，提取、纯化IgG制备免疫亲和色谱柱，针对沙丁胺醇与克伦特罗的动态柱容量分别为400ng／mL基质和416ng／mL基质。猪肝样品匀浆后用稀盐酸提取，经免疫亲和色谱柱纯化，再用GC／MS检测，即为IAC-GC／MS法。对沙丁胺醇的检测限为0．5ng／g，定量限为2ng／g，对克伦特罗的检测限为0．8ng／g，定量限为2．5ng／g。空白组织按照1、5、10、20ng／g的浓度添加药物，沙丁胺醇在空白肝中的添加回收率为78．4％～106．9％，克伦特罗的添加回收率为77．1％～102．9％。本研究建立的检测猪肝中沙丁胺醇与克伦特罗的IAC-GC／MS法乃国内首次报道。     

Detection of β2-agonists in milk replacer

₂-Agonist drugs may be illegally used as growth promoters for feedlot calves, when mixed into milk replacer immediately before feeding. To check for the presence of clenbuterol, salbutamol and terbutaline in such food, an analytical system was established using a screening method based on two commercial qualitative competitive ELISA tests, with antibodies raised against the arylamino group and thet-butyl group. The extraction procedure was based on precipitation of the milk samples with acetonitrile followed by filtration. The absence of any significant interference by other substances in the filtrate allowed detection of ₂-agonist drugs in spiked samples at the lowest concentration having a repartitioning effect (50 ppb for clenbuterol, mabuterol and terbutaline, 500 ppb for salbutamol). In view of a false positive response with tetracycline in milk samples and a cross-reaction between clenbuterol and mabuterol, an HPLC-MS technique was developed which, after extraction and purification of the samples with SPE C18 Polar Plus, was able to confirm the presence of these drugs. The good recovery after extraction (ranging from 84% to 90.2%) and the low detection limit with this method (250 ng/ml for clenbuterol, mabuterol and terbutaline, and 2.5 µg/ml for salbutamol) allowed easy confirmation and simultaneous detection of the four ₂-agonists at the lowest concentrations at which they are used in adulterated milk for calves.Abbreviations B optical density of the sample - B maximal optical density in total absence of competition - %B/B ₀ percentage of inhibition - ELISA enzyme-linked immunosorbent assay - EIA enzyme immunoassay - HPLC-MS high-performance liquid chromatography-mass spectrometry - m/z mass to charge ratio - ppb parts per billion - ppm parts per million - SPE solid-phase extraction     

沙丁胺醇残留的酶联免疫检测方法的建立

用自主制备的沙丁胺醇特异性抗体建立了针对沙丁胺醇残留的间接竞争酶联免疫检测方法,对该检测体系的灵敏度、准确度、精密度、特异性等性能进行了测定,并与高效液相色谱方法进行了对比性分析。结果显示:沙丁胺醇药物残留的间接竞争酶联免疫检测体系的检测范围为1～80ng/mL,灵敏度为0.58ng/mL,检测限为1ng/mL,回收率为70～99%,与盐酸克仑特罗、硫酸特布他林交叉反应率分别为107%、10%,与盐酸莱克多巴胺及肾上腺素的交叉反应率小于0.01%;采用HPLC方法进行沙丁胺醇药物残留的检测时,其检测范围为10μg/mL～200μg/mL,检测限为1μg/mL,回收率为80～95%。该方法与高效液相色谱法相比灵敏度较高,特异性强,检测结果准确度相近,但是在检测结果稳定性方面逊与HPLC方法。     

酶联免疫吸附（ELISA）法测定生猪尿液中的沙丁胺醇

将标准品、样品和和酶标记物一并加入到微孔板中孵育,样品中的游离沙丁胺醇与酶标记的沙丁胺醇竞争结合酶标板微孔中固定相化的沙丁胺醇特异性抗体,通过清洗步骤洗掉未结合的酶标记沙丁胺醇,再通过酶的专一性显色剂显色,微孔显蓝色,加入反应终止液使颜色由蓝色转为黄色。在450nm处测量吸光度（OD）值,并设置标准曲线,通过标准曲线测定样品中沙丁胺醇的含量,结果显示：OD值的大小与样品中沙丁胺醇的含量成反比。     

分子印迹膜修饰丝网印刷电极快速测定饲料中的沙丁胺醇

以裸丝网印刷电极(SPE)为载体原位合成分子印迹膜(MIM),制得分子印迹膜修饰丝网印刷电极(MIM-SPE),从而建立MIM-SPE检测沙丁胺醇(SAL)的方法。首先通过SPE获得标准液中SAL的电化学行为图像,并计算得到标准曲线:I=2.9818+0.14302c,R=0.998,检测限为0.9mg/L,检测的线性范围为1.0~6.0mg/L;再用修饰电极测定SAL,得到标准方程:I=8.9433+0.79294c,R=0.996,检测的线性范围为0.50~8.0mg/L,检测限为0.2mg/L。该方法平均回收率可达86.7%,准确率为100%,存在相似干扰物的情况下,检出量为94.8%以上,结果比较理想。另外,此方法检测单个样品用时仅为150s,因此MIM-SPE可以方便快速检测SAL,检测成本较低,适用于现场的大批样品快速筛选工作。     

β-兴奋剂沙丁胺醇及其检测技术研究进展

在畜禽生产中滥用β－兴奋剂将对人体健康造成极大的损害，国内外对肉食品中β－兴奋剂残留的检测分析都非常重视。本文对β－兴奋剂沙丁胺醇的各种检测方法的原理进行了较为详细的阐述，综述了其在肉食品沙丁胺醇残留检测中的应用及国内外研究状况。免疫标记技术以其快速、灵敏、简便，仪器分析技术以其准确可靠的特性分别被广泛应用于沙丁胺醇的抽查和确证工作。展望未来，在各种检测技术中，以酶联免疫吸附分析方法为基础的ELISA检测试剂盒和免疫胶体金试纸条的应用前景将最为广阔。