六种生物药物分子的高分辨太赫兹光谱研究

利用太赫兹时域光谱技术,在77～295 K之间,分别测量了两种异黄酮(金雀异黄酮和鹰嘴豆芽素A)、两种瘦肉精(盐酸克伦特罗和沙丁胺醇)和两种人参皂苷(Rg2和Rg3)等三类药物生物分子随温度变化的低频振动光谱,发现结构和功能相似的同类样品在室温环境下的太赫兹光谱均存在明显的不同。当温度从295 K冷却到77 K,测量样品的特征峰增加,鹰嘴豆芽素A可以观察到13个高分辨的特征吸收峰;并伴随线宽变窄,可以为1.9×10-5 mol用量的人参皂苷20-(R)-Rg3提供较准确的吸收峰。得到的高分辨太赫兹指纹谱为快速、无损鉴定该类药物生物分子提供一种可靠的新方法。     

测定克喘素免疫偶合物分子结合摩尔比的紫外-可见分光光度法

以牛血清白蛋白（BSA）、卵清白蛋白（OVA）为载体，采用重氮法，制出克喘素（CL）的两种免疫偶合物免疫原CL－BSA和包被抗原CL－OVA；通过紫外光谱定性证明偶合物偶联成功，通过可见光谱法定量测定偶合物的分子结合摩尔比，结果表明CL－BSA与CL－OVA的分子结合摩尔比分别为13．19：1、7．53：1，方法简便快捷，灵敏度高。     

人尿中克仑特罗的测定

本文采用直接液－液萃取和ＴＭＳ衍生化方法，利用ＧＣ／ＭＳ选择离子检测技术对人尿中的克仑特罗进行了分析研究，建立了灵敏的分析方法。给出了克仑特罗的回收率、检测限及人尿中的排泄曲线。     

衍生固相萃取GC-MS检测牛可食组织中克伦特罗的残留量

盐酸克伦特罗(clenbuterol,CLB),俗名"瘦肉精",化学名为"2-[(叔丁氨基)甲基]-4-氨基-3,5-二氯苯甲醇盐酸盐"(相对分子质量313.5),是一种β-兴奋剂。近年来动物性产品中CLB残留对人类健康造成的危害和对生态食物链所造成的破坏已经引起高度重视。1986年开始,欧美等发达国家已严禁畜牧生产中应用CLB。中国农业部在1997年3月以[农牧发(1997)3号文]严令禁止β-肾上腺素能激动剂在动物生产中应用。但目前国内外非法使用情况仍存在。国际上发展了一系列畜禽     

质谱快速分析猪肉中痕量沙丁胺醇及克伦特罗

采用内部萃取电喷雾电离质谱( iEESI-MS)技术,在无需样品预处理的前提下,采用标准加入法直接对猪肉组织中沙丁胺醇与克伦特罗进行定性和定量分析。结果表明,本实验对猪肉组织中沙丁胺醇与克伦特罗具有较高的灵敏度,单个样品单一指标的检测时间少于30 s。在0.01~1000μg/kg浓度范围内,信号强度对数(Y)与浓度对数(X)具有较好的线性关系,定量限分别为6.2和9.8 ng/kg。本方法分析速度快、样本耗量少、灵敏度高,适用于猪肉中痕量沙丁胺醇与克伦特罗等“瘦肉精”的快速检测。     

Newly Combined Method of Molecularly Imprinted Solid-Phase Extraction with ELISA for Rapid Detection of Clenbuterol in Animal-Tissue Samples

Abstract

A new method using molecularly imprinted polymers (MIPs) as specific adsorbent materials coupled with ELISA analysis is being reported for the first time for the detection of clenbuterol (CLB) residue in the pig muscles. After optimization of the posttreatments, the total amount of template bleeding in the CLB MIPs was decreased to only 3.0 ng CLB/60 mg MIPs, which is 10 times lower than that of the previous report. Moreover, compared to the methods of C18-ELISA and single ELISA, the combined molecularly imprinted solid-phase extraction (MISPE)–ELISA exhibited high precision and robust accuracy for CLB at all three spiked levels of 0.5, 5.0, and 10.0 ng g^?1.     

酶联免疫法测定猪肉、猪肝和猪尿中盐酸克伦特罗

对4种国产及进口的酶联免疫法测试盒测定猪肉、猪肝及猪尿中克伦特罗残留量的性能及效果作了对比试验。试验时以上述3种样品为基体,加入1,2,3,5,10μg.kg-1(或μg.L-1)等5个浓度水平的克伦特罗标准溶液,然后用上述测试盒进行检测。从所得的结果可知,国内外的测试盒所得测定值间存在较大差异;猪尿样品的检测结果表明这些测试盒所采用的方法比较适合,但由猪肉及猪肝样品的检测结果可知,其中一些测试盒所采用的方法尚有改进的必要。     

Simultaneous determination of clenbuterol, salbutamol and ractopamine in milk by reversed-phase liquid chromatography tandem mass spectrometry with isotope dilution

A simple, sensitive and reliable analytical method was developed for the simultaneous determination of clenbuterol (CLB), salbutamol (SAL) and ractopamine (RAC) in milk by ultra high performance liquid chromatography–positive electrospray ionization tandem mass spectrometry (UHPLC–ESI-MS/MS) with isotope dilution. Samples were directly purified through HLB cartridge. Then the eluate was dried under nitrogen and residues were redissolved in mobile phase. Samples were analyzed by LC–MS/MS on an Acquity UPLC^® BEH C₁₈ column with gradient elution. The samples were quantified using clenbuterol-D₉, salbutamol-D₃ and ractopamine-D₆ as internal standards. The proposed method was validated according to the European Commission Decision 2002/657/EC determining specificity, decision limit (CCα), detection capability (CCβ), recovery, precision, linearity, robustness and stability. CCα values were 0.054, 0.006 and 0.008 μg/kg for CLB, SAL and RAC, respectively. CCβ values were 0.058, 0.007 and 0.009 μg/kg for CLB, SAL and RAC, respectively. The mean recoveries, repeatability (expressed as coefficient of variation, CV_r), and reproducibility (CV_R) varied from 95.8 to 106.2%, from 3.60 to 6.44% (CVr), and from 4.77 to 7.53% (CV_R), respectively. The method is demonstrated to be suitable for the determination of clenbuterol, salbutamol and ractopamine in milk. The total time required for the analysis of one sample, including sample preparation, was about 45 min.     

On the use of dispersed nanoparticles modified with single layer beta-cyclodextrin as chiral selecor to enhance enantioseparation of clenbuterol with capillary electrophoresis

A new strategy for chiral separation by capillary electrophoresis employing modified-nanoparticles as chiral selector is described for clenbuterol analysis. Nanoparticles modified with β-cyclodextrin (β-CD) form a large surface area platform to serve as a pseudostationary chiral phase, which can be applied for the enhancement of the enantioseparation. The application of four kinds of nanoparticles was investigated (multi-walled nanotubes (MWNTs), polystyrene (PS), TiO₂ and Al₂O₃) modified with single layer β-CD as chiral selector in the enantioseparation of clenbuterol by capillary electrophoresis (CE). Successful clenbuterol enantioseparation could be achieved with the β-CD-modified MWNTs as chiral selector. X-ray diffraction (XRD) and Fourier transform infrared spectroscopy (FTIR) confirmed the β-CD modification of the nanoparticles. The effects of nanoparticles, surfactant, chiral selector (β-CD) and run buffer were studied in relation to the enantiomeric separation of clenbuterol. This study opens attractive perspectives for the use of modified nanoparticles for chiral separational purposes in CE.     

Surface Plasmon Resonance‐based Inhibitive Immunoassay Coupled with Dummy Template Molecularly Imprinted Polymer Solid Phase Extraction for On‐line Analysis of Trace Clenbuterol

A new indirect inhibitive immunoassay using surface plasmon resonance (SPR) coupled with molecularly imprinted polymer (MIP) was developed and applied for the analysis of trace clenbuterol (CL). A MIP coating using phenylephrine as the dummy template was synthesized in a flexible quartz capillary (30 cm×0.25 mm i.d.) by in situ polymerization technique, which then was used as the online solid phase extraction (SPE) tube before SPR detection. The thickness of the coating was 198 nm on average, illustrated by scanning electron microscope (SEM) image. The mechanism for adsorption of CL on dummy template MIP was found to be a Freundlich isotherm and pseudo‐second‐order model. The immunoassay was conducted on BIAcore 3000 biosensor automatically without pre‐treatment. The calibration curve was generated by linear fit in the range of 0.1–10 ng L^‐1 and 10–100 ng L^‐1. The detection limit was 0.1 ng L^–1, which is super sensitive. This method was directly applied for the analysis of real‐world samples without pretreatment.