磺胺地索辛抗原合成及多克隆抗体的制备

磺胺地索辛（Sulfadimethoxine，SDM）是具有对氨基苯磺酰胺结构的磺胺类药物的一种，是一种小分子药物，本身不具有免疫原件，不能直接免疫动物产生抗体，必须与载体蛋白偶联后，才具有免疫原性。     

牛奶中磺胺类药物免疫胶体金试纸条的制备

本研究将磺胺二甲嘧啶与载体蛋白偶联形成完全抗原,与混合磺胺类药物单克隆抗体反应制备试剂条。通过抗原抗体之间的免疫反应,采用胶体金检测方法,制备用于检测牛奶中磺胺类药物的试纸条,检测灵敏度最低可达100μg/kg,该检测限满足欧盟及我国对磺胺类药物检测的需求,且能够实现牛奶样品中多种磺胺类药物的现场快速检测。     

动物源性食品中磺胺类药物残留检测方法进展

磺胺类药物广泛应用于兽医临床治疗疾病和作为饲料添加剂促进动物的生长。磺胺类药物残留对人体存在潜在的威胁,目前常用的磺胺类药物残留检测方法有很多,包括各种微生物法、免疫法以及理化分析法。本文对这些方法加以综述。     

对氨基苯磺酰胺多克隆抗体的制备及特异性测定

为了制备对氨基苯磺酰胺多克隆抗体,试验采用N-羟基琥珀酰亚胺活性酯(NHS)法,将对氨基苯磺酰胺(SA)分别与牛血清白蛋白(BSA)耦联制备免疫抗原SA-BSA,与卵清蛋白(OVA)耦联制备检测抗原SA-OVA,用合成的SA-BSA耦联物免疫小鼠,制备对氨基苯磺酰胺多克隆抗体,并测定其特异性。结果表明:用合成的SA-BSA耦联物免疫小鼠获得了高效价的鼠源对氨基苯磺酰胺多克隆抗体,该抗体可与多种磺胺类药物发生交叉反应,与异类药物不发生反应,可作为动物源食品中磺胺类药物的多残留检测抗体。     

食源性动物饲料中磺胺类药物残留的检测及其研究

作为一类抗菌谱广、低毒且高效、价格低廉的药物,磺胺类药物近年来在食源性动物的饲料中开始被广泛应用。但磺胺类药物的使用具有严格的剂量要求,一旦使用不合理即很有可能残留在动物组织中,通过食物链过程最终会将不良影响传递给人类,对人体健康产生危害。加强对食源性动物饲料中磺胺类药物残留检测的研究,已经势在必行。本文即从磺胺类药物残留的来源与危害出发,对食源性动物饲料中磺胺类药物残留的检测方法进行研究。     

动物源性食品中磺胺类药物残留的危害及监控对策

长期食用磺胺类药物残留的动物源性食品会对人体产生严重的危害,因此动物源性食品中磺胺类药物残留的监控日益受到重视。本文阐述了磺胺类药物残留对人体的危害,列举目前我国磺胺类药物残留监控存在的问题,并对其提出相应的对策。     

检测磺胺类7种药物ELISA试剂盒的研制

以SAs-BSA的偶联物作为抗原免疫Balb/c小鼠,制备磺胺类药物单克隆抗体。与7种磺胺药(SD、SMM、SMZ、ST、PST、SMT、SPD)有很高的交叉反应性,并建立一种可检测样品中的SAs类药物的间接竞争ELISA法。试剂盒灵敏度为2.14μg/L,对SAs类药物的回收率在56.2%～72.1%之间,变异系数均20%,研制的ELISA试剂盒可进行磺胺类药物多残留快速检测。     

磺胺类药物广谱性多克隆抗体的制备

基于磺胺类药物(Sulfonamides,SAs)公共母环(对氨基苯磺酰胺)结构,合成了两种具有该结构的半抗原∶2-(对氨基苯磺酸氨基)-4-噻唑乙酸(TS)和对氨基苯磺酰胺基苯甲酸(SS).采用碳二亚胺法合成了免疫原和包被原,分别免疫10只新西兰大白兔,制备广谱性磺胺类药物抗体.应用间接竞争酶联免疫吸附分析(ELISA)检测血清的灵敏度和特异性,发现所有兔子对TS-BSA和SS-BSA均产生较高效价和特异性.其中SS-1号和TS-6号兔的血清灵敏度最好,效价分别为1∶80 000和1∶160 000,对10种磺胺类药物均有不同程度的识别能力.SS-BSA组IC50为257.9～6 590.9 ng/mL,而TS-BSA组IC50为12.9～586.1 ng/mL.结果表明,具有磺胺母环结构的半抗原TS和SS,可以应用于广谱性磺胺类抗体的制备.通过10只兔子免疫试验证明,应用SS获得的血清具有较高的簇特异性、相似的亲和性,优于TS.     

动物性食品中磺胺类药物残留检测方法研究进展

磺胺类药物是一类人工合成广谱抗菌药物,因为其疗效好且价格相对低廉,在动物养殖和疾病防治中应用较为广泛。但这类药物在使用后易以原型排出,且大剂量、长时间使用时会造成其在动物产品中的残留,对人的健康和环境造成危害。对动物性食品中磺胺类药物残留的检测是保证产品食用安全和减少环境污染的重要措施。磺胺类药物由于种类多、使用广泛、在不同动物性产品中的残留情况也不相同,在对其残留进行检测时,应选择合适方法对样品进行处理和检测。本文介绍了样品前处理的常用方法,如液液萃取等传统方法和固相萃取法、固相微萃取法以及基质固相分散法等的原理、优缺点等。对动物性食品中磺胺类药物的检测,应用较多的是高效液相色谱法、毛细管电泳法放射免疫法、免疫传感器发和酶联免疫法等,可使实现对产品中药物残留的定性和定量检测,但是也存在需要专门仪器、检测时间相对较长的问题;快速检测方法最常用的就是胶体金免疫层析方法,可以实现对中多种磺胺类药物的快速和定性的检测,是食品种抗菌药物检测技术研究的一个发展方向。通过对动物性食品中抗微生物残留进行初步筛查,确定为阳性的样品在进行实验室的定量检测,是保证产品安全、可操作的基本程序。研究高通量、敏感、快速、操作简便的动物性食品中磺胺类药物残留检测技术,也是一个重要方面。     

展青霉素人工抗原的制备与鉴定

为探讨展青霉素（patulin,PAT）不同抗原合成方法对完全抗原制备及抗体产生的影响,本试验利用琥珀酸酐联合活泼酯法,分别制备免疫原PAT-BSA和包被原PAT-OVA。为提高PAT与偶联蛋白的结合率,尝试将BSA修饰为EDA-BSA后与PAT进行偶联,并对比2种偶联方法制备的完全抗原免疫动物后抗血清的效价水平。同时利用琼脂糖凝胶电泳、非变性聚丙烯酰胺凝胶电泳、变性聚丙烯酰胺凝胶电泳、凝胶渗透色谱多种方法分析鉴定偶联结果,为PAT单克隆抗体的制备及免疫学检测方法的建立奠定了基础。     

磺胺母核人工半抗原的合成与鉴定

合成磺胺母核人工半抗原,并对其产物进行鉴定。采用4-氨基苯甲酸甲酯（PBPA）和对乙酰氨基苯磺酰氯（ASC）为原料,经过亲和取代,酯的水解反应合成磺胺药物共有的母核结构苯甲酸对氨基苯磺酰胺（SH）,应用质谱法（ESI—MS）与核磁共振氢谱法（1H—NMR）鉴定SH。质谱结果显示[M-]=291.03766,与理论值[M]-292相符,表明SH合成成功;核磁共振氢谱（1H—NMR）的数据与SH化合物的结构相符。磺胺母核人工半抗原合成成功,为研制磺胺药多残留检测试剂盒和胶体金免疫层析试纸条奠定基础。     

含噻唑环的磺胺母核合成与鉴定

详细介绍了以2-氨基-4-噻唑乙酸（ATAA）和对乙酰氨基苯磺酰氯（ASC）为原料,经过酯化、亲和取代、酯的水解反应合成含噻唑环的磺胺母核即N^1-[4-（羧甲基）-2-噻唑]对氨基苯磺酰胺（TS）的过程及采用碳二亚胺法（EDC法）合成完全抗原TS-牛血清白蛋白（TS-BSA）的方法。应用质谱法鉴定TS,紫外扫描鉴定完全抗原TS-BSA。质谱结果显示[M^-]：311．92,与理论值[M]=313．01相符,表明Ts合成成功;紫外扫描特征峰值发生偏移,证明完全抗原偶联成功,计算其结合比为11：1。     

恩诺沙星免疫抗原的制备与鉴定

通过N-羟基琥珀酰亚胺活化酯法合成恩诺沙星免疫抗原,用聚丙烯酰胺凝胶电泳、紫外光谱和酶联免疫法分析对恩诺沙星免疫抗原的结构、分子量、免疫原性等特征进行鉴定。经紫外光谱分析恩诺沙星-牛血清白蛋白偶联比为5:1,紫外光谱峰形发生变化并位移。间接酶联免疫法分析证明其免疫兔所获得的抗体与恩诺沙星有良好的特异性,半数抑制浓度为60ng/mL。实验结果表明成功合成了恩诺沙星免疫抗原,并获得了抗恩诺沙星抗体,为ELI SA或TRFI A等免疫方法的进一步研究提供了试验基础。     

The biotransformation of sulfadimethoxine, sulfadimidine, sulfamethoxazole, trimethoprim and aditoprim by primary cultures of pig hepatocytes

The in vitro biotransformation of three sulfonamides, trimethoprim and aditoprim, was studied using primary cultures of pig hepatocytes. Incubation of monolayer cultures with sulfadimethoxine (SDM), sulfamethoxazole (SMX) and ¹⁴C-sulfadimidine (SDD) resulted in the formation of the corresponding N 4-acetylsulfonamide to different extents, depending upon the molecular structure of the drug. Addition of the acetylsulfonamides to the cells showed that these compounds were deacetylated, each to a different extent. A relatively low degree of acetylation (in the case of SDD) was paralleled by extensive deacetylation (i.e. AcSDD), whereas extensive acetylation (i.e. SMX) was in concert with minor deacetylation (i.e. AcSMX). The addition of bovine serum albumin to the medium resulted in a decrease in conversion of sulfonamides as well as acetylsulfonamides. The main metabolic pathway of ¹⁴C-trimethoprim (TMP) was O -demethylation with subsequent conjugation. Two hydroxy (demethyl) metabolites were formed, namely 3'- and 4'-demethyl trimethoprim, which were both glucuronidated while 3'-demethyl trimethoprim was also conjugated with sulphate. The capacity to form conjugates with either glucuronic acid or sulphate was at least as high as the capacity for O -demethylation since more than 90% of the metabolites were excreted as conjugates in the urine of pigs. Addition of ¹⁴C-aditoprim (ADP) to the hepatocytes led to the N -demethylation of ADP to mono-methyl-ADP and di-desmethyl-ADP. During the incubation another three unknown ADP metabolites were formed. In contrast to TMP, no hydroxy metabolites or conjugated metabolites of aditoprim were formed. These in vitro results were in agreement with the in vivo biotransformation pattern of the studied sulfonamides and trimethoprim in pigs.     

兽药残留免疫检测技术应用进展

免疫检测技术（Immunoassays,IAs)是利用抗原与抗体在体外的特异性反应建立起来的检测技术。兽药属于小分子物质（MW＜1000D），没有免疫原性，必须与大分子物质交联成人工抗原才能获得免疫原性。本文从兽药人工抗原的制备、抗体的制备、免疫检测方法的建立等三个方面对免疫技术在兽药残留检测的应用进行了综述，并展望了兽药免疫检测技术的发展趋势和前景。     

Transference of Dietary Veterinary Drugs into Eggs

Twelve veterinary drugs, bacitracin (BC), chloramphenicol (CAP),chlortetracycline (CTC), oxytetracycline (OTC), tylosin (TS), amprolium (APL), furazolidone (FZD), nicarbazine (NCZ), ormetoprium (OMP), sulfadimidine (SDD), sulfadimethoxine (SDM) and sulfamonomethoxine (SMM) were fed to laying hens for 14 days, each at a dietary concentration of 500 mg/kg. The concentrations of the drugs in the eggs from these birds were determined at 2-day intervals for 14 days after the start of feeding. The relationship between the concentrations of the drugs (mg/kg) in the eggs and the number of days after the start of feeding was analysed by regression and covariance analyses. The concentrations of the drugs in the eggs became constant after 4 days for OTC, TS, FZD and all the sulfonamides, and after 6 days for CAP, APL, NCZ and OMP. No BC or CTC was detected in the eggs. The transfer rates of the 10 drugs (excluding BC and CTC) from the feed to eggs varied from 0.005% for TS up to 1.540% for SDD.     

氯羟吡啶抗原合成和多克隆抗体制备

本研究以氯羟吡啶原药为基础,合成了带羧基的氯羟吡啶衍生物,经质谱鉴定结构正确,并用活性酯法和氯甲酸异丁酯法合成抗原,紫外测定偶联比率和最终抗原蛋白浓度,免疫制备抗血清并测定血清特异性和灵敏度,为建立氯羟吡啶酶联免疫法(ELISA)奠定坚实基础。     

甲硝唑人工抗原的制备及其ELISA检测

采用碳二亚胺法将甲硝唑半抗原与牛血清白蛋白(BSA)连接制备人工免疫原,同样方法将其与卵清蛋白(OVA)连接制备人工包被原.经紫外扫描分析,两种方法合成的免疫原和包被原的结合比分别为8∶1、2∶1和6∶1、2∶1;动物免疫试验分析偶联物,小鼠抗体效价达1∶5 000,与3种硝基咪唑类药物的交叉反应率均小于10%.表明制备的抗体可以用于甲硝唑残留的检测,同时为检测试剂盒的研制奠定了基础.     

氨基脲人工抗原的合成及其多克隆抗体的制备

目的合成氨基脲人工抗原并制备其多克隆抗体。方法以氨基脲(SEM)和对醛基苯甲酸(CP)为原料合成半抗原(CP-SEM),并采用碳化二亚胺法和混合酸酐法将其分别与牛血清白蛋白(BSA)和鸡卵清蛋白(OVA)偶联,合成氨基脲人工抗原。以氨基脲人工抗原(CPSEM-BSA)为免疫原免疫Balb/c小鼠,利用间接ELISA法测定其抗体效价。结果经薄层层析、红外光谱和元素分析等方法鉴定合成半抗原即为CP-SEM。紫外扫描、聚丙烯酰胺凝胶电泳结果表明人工抗原合成成功。免疫获得抗氨基脲的多克隆抗体,其抗体效价为1:64000。结论氨基脲人工抗原合成成功,并获得抗氨基脲的多克隆抗体。     

The sulfonamide-diaminopyrimidine story*

This paper is devoted to chemotherapy in the sense that Paul Ehrlich (Nobel Prize winner in 1908) coined the word: to describe the cure of infectious diseases by chemical agents without injury to the organism infected. This approach, essentially that of selective toxicity, is applicable to the investigations performed by Gerhard Domagk (Nobel Prize winner in 1939), which resulted in the development of Prontosil rubrum. This agent was active in vivo but not in vitro. Trefouel supposed that the in vivo action was due to a metabolite of the drug (sulfanilamide), a hypothesis later proved by Fuller in 193 7. Sulfanilamide was a simple agent, easy to manufacture and free of patent rights. Thus, more than 5400 derivatives were synthesized and studied in the decades that followed. Research on the side-effects of sulfonamides resulted in the development of diuretics and antidiabetogenic agents. The resurgence of interest in sulfonamides in following years has been associated with the development of diaminopyrimidines by Hitchings (Nobel Prize winner in 1988). It soon became evident that combinations of these drugs produced potent synergistic effects. Trimethoprim, a potent antibacterial agent of this series, was selected for combining with sulfonamides such as sulfadiazine. This line is still continued and even recently new diaminopyrimidine derivatives such as aditoprim and baquiloprim have been synthesized having superior properties in farm animal species. Thus, even nowadays the principles of Ehrlich continue to have a major impact on the development of new veterinary drugs.