【作者】 李洁；

【导师】 韩芹芹；

【作者基本信息】 昆明理工大学 ， 微生物学（专业学位）， 2018， 硕士

【摘要】 特异性强、灵敏度高的非法食品添加剂快速检测方法,对食品安全有着极其重要的意义。凡是在国家允许范围之外或者超标使用的添加剂都称为非法添加剂。吊白块本是一种常用的工业染料,为强致癌物,误食后,会对人体的肺、肝脏,肾脏等器官造成不同程度的损伤。《食品添加剂使用卫生标准》中明确规定,严禁在任何食品加工过程中添加或使用吊白块。但仍有部分商贩在米粉、豆皮、豆腐和鱼翅的生产加工过程中违法添加和使用吊白块来达到给食品增白、增韧的目的。目前国标实行的吊白块检测方法是基于检测吊白块的分解产物来间接检测吊白块,这些方法不仅操作繁琐,而且通过检测吊白块分解产物的方法会造成较大的偏差,因此亟需发展一种能直接快速检测吊白块的方法。通过指数富集的配体系统进化技术(Systematic Evolution of Ligands by Exponential Enrichment,SELEX)筛选得到的适配体,作为一种新型识别元件,在食品安全,药物研发,医学诊断等领域都有极其重要的应用。本课题组通过前期工作已经筛选出两条能特异性识别吊白块的核酸适配体SF-A09和SF-B09。免疫层析试纸条(Literal flow strip assay,LFSA)是一种利用可视化传感器来实现待测物快速检测的分析方法,具有操作简单、检测迅速、成本低廉以及可现场检测等特点。本课题组借助LFSA平台,以SF-A09和SF-B09识别元件为基础,针对试纸条的整个反应体系(包括重悬液配方,膜预处理溶液配方,膜材料型号,各溶液最适浓度)进行研究,成功建立了一种利用胶体金适配体试纸条检测吊白块的方法,并将该试纸条用于食品中吊白块的现场快速检测。实验室中确定了该试纸条对吊白块的检测限为1μg/m L,并采用高效液相色谱法(High Performance Liquid Chromatography,HPLC)进行结果确认,以保证检测结果的准确性和可靠性。本研究还对周边市场的部分食品进行抽样检测,再次确认了该试纸条在现场快速检测中的应用。同时考察了不同样品基质的基质效应,确定了米线、饵丝、豆腐皮的检测限均为1μg/m L,而腐竹的基质效应较为严重,为2μg/m L。综上所述,本研究以SF-A09和SF-B09适配体为元件研制的胶体金适配体试纸条可以用于米线等食品样品中吊白块的现场快速检测,为食品中非法添加剂吊白块的现场快速检测提供了新的技术手段,保障了食品市场的安全。更多还原

【Abstract】 Specificity,high sensitivity and rapid detection of illegal food additives and toxins have great significances for food safety.All additives that are outside the permitted scope of the standard or exceed the standard are called illegal additives.Rongalite is one kind of industrial dyestuff,it can cause allergies,intestinal irritation or other adverse reactions,even poisoning,kidney,liver damage or other diseases.In the national standard "food additives using health standards",hanging rongalite is excluded from the statutory food additives.However,there are still some traders who illegally add and use them in the production process of rice flour,bean curd,tofu and shark fin to achieve the purpose of whitening food.At present,the main detection methods of rongalite are detecting the decomposition products of rongalite.These methods not only operate cumbersome,but also cause large deviation by detecting the decomposition products.Therefore,it is urgent to develop a method for rapid detection of rongalite.The new identification elements,aptamers,screened by SELEX have extremely important applications on food safety,drug research and development,medical diagnosis and other fields.Our research group has screened two aptamers SF-A09 and SF-B09 that can specifically identify rongalite.Literal flow strip assay(LFSA)is a simple rapid and inexpensive analytical method that uses visualization sensors to rapidly detect analytes.Therefore,our group used the LFSA platform,based on SF-A09 and SF-B09 as the identification elements,and then compare and optimize the entire reaction system for the test strip,including the resuspension formulation,the membrane pretreatment solution formulation,the membrane material model,and the optimum concentration of the various solutions.A colloidal gold aptamer test strip was successfully established to detect hanging white pieces,and the test strip was used for on-site rapid detection of hanging white pieces in food.A sandwich lateral flow strip assay(LFSA)using a couple of aptamers functionalized with gold nanoparticles(AuNPs)was designed to assess the presence of rongalite in agrifood products.This system allows the successful and direct detection of rongalite in food samples with concentrations as low as 1 μg/mL,simply by observing the color change of LFSA control and test line.These results were confirmed by high performance liquid chromatography(HPLC).In addition,food samples from the surrounding market were purchased for testing.The application of LFSA in point-of-care testing(POCT)field was confirmed again.At the same time,the matrix effects of different sample were investigated.The detection limits of noodle,ersi,and skin of tofu is 1 μg/m L,and the detection limits of dried beancurd sticks is 2 μg/ m L.In summary,the self-assembled sandwich lateral flow strip assay(LFSA)using a couple of SF-A09 and SF-B09 aptamers functionalized with gold nanoparticles(AuNPs)can be used for on-site rapid detection of rongalite in rice noodle and other food samples,which provides new technical means to ensure the safety of the food market.更多还原