不同前处理对植物酯酶抑制法检测蔬菜中有机磷及氨基甲酸酯类农药残留的影响

利用酶抑制法可以快速检测蔬菜中的有机磷和氨基甲酸酯类农药残留,这两类农药能抑制酯酶的活性,并影响酯酶对底物的催化能力。根据这一原理,采用麦麸酯酶作为酶源对酶抑制法快速检测农残前处理方法中农残提取液、提取方法、提取时间及样品提取液的处理方式进行优化。结果表明:用5%丙酮-磷酸盐缓冲液作为提取液,然后超声提取5 min后,将样品提取液再用0.45μm有机滤膜过滤后蔬菜中农药残留提取量可达到最高;且样品加标回收率为83.61%~102.81%,变异系数为1.20%~7.23%。因此,该前处理方法可满足于麦麸酯酶抑制法快速检测农药残留的需要。     

生物荧光酶抑制法快速定性测定粮食中有机磷类和氨基甲酸酯类农药残留

建立了生物荧光酶抑制法测定粮食及其制品中的有机磷类及氨基甲酸酯类的农药残留快速定性检测方法。样品通过乙酸乙酯：丙酮(v:v=95:5)提取离心净化,取50 μL上清液45℃氮气吹干,1 mL蒸馏水或去离子水复溶进行检测。结果表明,本方法测定粮食及其制品中的有机磷类及氨基甲酸酯类农药残留时,最低检测限为0.1 μg /kg,提取液与酶溶液的最佳反应时间是10 min,提取液与酶溶液的最佳反应温度为35℃。通过确立不同农药的限量抑制率,可以提高判定的准确度。本方法同时处理7个样品需要20 min,检测时间为15 min,平均每个样品5 min即可出结果,可用于大批量粮食及其制品中有机磷类及氨基甲酸酯类农药残留的快速筛选。     

酶联免疫吸附法(ELISA)检测婴幼儿配方食品中谷蛋白

介绍了利用ELISA法的醇溶谷蛋白试剂盒测定婴幼儿配方食品中的谷蛋白含量。样品经处理后,用提取液提取样品中谷蛋白,提取液经离心,稀释后,用酶联免疫吸附法直接测定。检测的线性范围是5～80μg/kg,相关系数是0.9988,回收率在115.6%～120.6%之间,证明可以用酶联免疫法测定婴幼儿配方食品中的谷蛋白含量。     

电感耦合等离子体质谱法同时测定与食品接触 陶瓷制品中12种元素溶出量

目的建立电感耦合等离子体质谱法(ICP-MS)同时快速测定与食品接触陶瓷制品铅、镉、镍、铬、砷、锑、锌、钴、铜、锰、铝、钡等12种元素溶出量的分析方法。方法采用4%醋酸溶液,在(22±2)℃室温条件下避光浸泡样品24 h±20 min,以45Sc、72Ge、115In、209Bi作为内标,优化仪器参数和试验条件,用电感耦合等离子体质谱法同时测定浸泡液中12种元素溶出量,内标法定量。结果该方法的检出限为0.10~10.0μg/L,样品的加标回收率为90.0%~106.0%,相对标准偏差0.86%~4.31%。结论方法操作简便,准确度好,灵敏度高。可适用于食品接触陶瓷多种痕量有毒有害元素溶出量的检测。     

改进的QuEChERS结合超高效液相色谱—串联质谱法同时测定水产品中19种兽药残留

目的:实现水产品中19种磺胺、氟喹诺酮类兽药同步检测。方法:利用1.0%甲酸乙腈溶液作为提取液,超声1 min对样品中目标分析物进行提取,采用QuEChERS多功能针式过滤器净化样品,样品净化与过滤除杂同步进行,采用超高效液相色谱结合三重四极杆质谱进行检测。结果:19种磺胺、喹诺酮类兽药的检出限为0.5～1.0μg/kg,与国家标准方法相比,检测灵敏度有所提高。添加回收率为81.0%～113.0%,相对标准偏差(RSD)为0.2%～11.7%。结论:该方法通过超声辅助萃取、QuEChERS多功能针式过滤器提高样品前处理效率,具有方法检测灵敏度高、准确度高、重现性良好等优点,适用于大批量水产品样品中磺胺、喹诺酮类兽药日常监测。     

双抗夹心酶联免疫吸附法检测荞麦过敏蛋白

提取制备荞麦过敏原蛋白(TBt)并免疫动物,分别制备鼠多克隆抗体和兔多克隆抗体,建立了双抗夹心酶联免疫吸附检测法(ELISA)用于检测食品中的荞麦过敏原。SDS-PAGE结果表明,纯化的TBt纯度达到98%以上,鼠抗血清效价为1∶6400。建立的双抗夹心ELISA法对荞麦过敏原蛋白的最低检测限为0.16μg/m L,线性范围为0.16~16μg/m L。并采用建立的双抗夹心ELISA法对市场出售的15种食品中荞麦过敏成分进行了检测。结果显示,该方法对绝大多数食品中的荞麦过敏原都有很好的特异性及灵敏性,说明该法可用于食物过敏原的诊断及食品中微量荞麦过敏成分的检测。     

环介导等温扩增技术快速检测食品过敏原牡蛎成分

目的建立环介导等温扩增(loop-mediated isothermal amplification, LAMP)方法快速检测食品中的过敏原牡蛎成分。方法根据国家生物技术信息中心的牡蛎线粒体序列,通过Primer Explorer version 5.0软件设计引物并筛选出LAMP特异性扩增引物。并进一步对反应体系优化,对该方法的灵敏度、特异性以及稳定性进行验证。对10种牡蛎阳性样品、14种阴性样品、4类牡蛎相关食品进行检测。结果该方法可以检测出含牡蛎成分0.1%, DNA浓度为0.01 ng/μL的样品。在实验时间上大幅缩短,反应可在25～45 min内结束,并且可以在微量体系下完成,对于食品相关产品的检出率为100%。结论该方法操作简单、成本较低、特异性高、灵敏度好,适用于食品中过敏原牡蛎成分的检测。     

双抗体夹心ELISA法测定食物中牛奶过敏原蛋白成分

建立双抗体夹心酶联免疫吸附测定法(ELISA)测定食物中牛奶过敏原成分.通过提取牛奶过敏原蛋白,免疫小鼠制备抗牛奶过敏原的多抗,免疫印迹法鉴定多抗与牛奶过敏原蛋白的反应,并建立双多抗央心ELISA法.结果表明,该法检测牛奶过敏原蛋白的最低检出限为25μg/L,标准曲线在25～1000μg/L范围内线性良好;同时对市场上食物标签标注含有牛奶、未舍牛奶成分以及标注不详共12种食品进行检测,结果10种食品检测结果与食物标签标注内容相符,而2种标示不详的食品检测结果一个呈现阳性,一个呈现阴性.这说明本方法对于未标注牛奶过敏原成份的食品检测具有一定的实际应用价值.     

脐橙皮中果胶的提取工艺优化

采用单因素分析法分析果胶提取工艺,选取液料比、提取温度、提取时间、提取液pH值四个因素对脐橙皮果胶的关键因素进行优化,通过绘制折线图得到果胶提取的最佳条件。结果表明:脐橙皮的最佳工艺参数为液料比20︰1,pH为0.8,提取温度75℃,提取时间80 min,乙醇最终浓度60%,沉淀时间60 min。在此条件下,脐橙皮果胶得率为22.4%,为生产加工脐橙皮果胶提供了低廉、高效、简单的提取方法。     

食品中牛奶过敏原成分LAMP检测方法的建立

目的建立食品过敏原牛奶成分LAMP(loop-mediated isothermal amplification)检测方法,并与实时荧光PCR(real-time PCR)检测方法比对。方法针对牛线粒体细胞色素b(cyt-b)基因设计LAMP引物并建立反应体系,在特异性和灵敏度方面与real-time PCR检测方法比对。结果本研究建立的LAMP方法检测9份不同品牌的牛奶和羊奶及其加工制品,没有出现交叉反应,具有良好的特异性。该方法的检测灵敏度为0.5%,与real-time PCR方法检测灵敏度相当。检测了69份实际样品,检测结果与real-time PCR检测结果一致。结论本研究建立的食品过敏原牛奶成分LAMP检测方法简单经济,检测结果可靠,可有效缩短检测时间,适用于过敏原牛奶成分的检测,具有良好的应用前景。     

Detection of shrimp-derived components in food by real-time fluorescent PCR

Crustaceans such as shrimp and crabs and their products are important allergens in food, and allergic reactions due to the consumption of shrimp and crabs are frequently reported. However, the chemical properties of shrimp-derived allergens, except for Pen a I, are still unclear. Therefore, it is important to establish a more sensitive and specific method for detecting the composition of foods containing shrimp. In the present study, we developed a real-time fluorescent PCR to identify the specific shrimp-derived components in food. The primers and TaqMan probes for real-time fluorescent PCR were designed based on 16S rRNA genes through comparing a large number of nucleic acid sequences from different species of shrimp that have been published by the National Center for Biotechnology Information. In total, 56 kinds of samples, including different kinds of shrimp, crab, fish, shellfish, and octopus, were subjected to detection by real-time PCR. The results indicated that real-time fluorescent PCR could successfully identify the shrimp-derived components. In order to explore the effect of food processing on detection sensitivity, fish powder containing shrimp powder was treated by heating at 133°C for 30 min. The limit of detection of shrimp-derived components in fish powder was 0.05% (wt/wt).     

Application of antibodies for the identification of polysaccharide gum additives in processed foods

Anti-carbohydrate antibodies with specificities for polysaccharide gums were isolated from the serum of rabbits that were immunized with a solution of the gums and Freund's complete adjuvant. The primary objective was to test an immunological method for the detection of the polysaccharide gums as additives to processed foods. Analysis involved the extraction of food with phosphate buffer and the testing of the extract for a reaction with anti-gum antibodies by the agar diffusion method. Reaction by a specific gum with the homologous antibodies establishes the presence of the gum in the food. The method is a novel application of antibodies. The antibody method is highly specific for a gum and thus possesses advantages over other methods of analysis for polysaccharide gums as additives in processed foods.     

Effects of Microwave and Ultrasound Assisted Extraction on the Recovery of Soy Proteins for Soy Allergen Detection

The extraction of soy proteins for soy allergen detections is conventionally achieved with PBS buffer for at least 2 h at room temperature or 4 °C. This method has been reported to be inefficient due to time consumption and inadequate protein extraction resulting in false negative allergen detection and mislabeling of foods containing allergenic proteins. This study investigated the application of microwave (MAE) and ultrasound assisted extraction (UAE) techniques to extract and improve recovery of allergens from various soy matrices. Soy proteins were extracted from raw soy flour, soy protein isolate (SPI) and soy milk using MAE at 60, 70, and 100 °C for 5 and 10 min and UAE at 4 and 23 °C for extraction times of 1, 5, and 10 min with PBS, Laemmli and urea buffers. Extracts were analyzed for total proteins, protein profile, and antibody‐based detection (ELISA) of soy proteins. Conventional extraction with each of the buffers was used as controls. Overall, proteins recovered from MAE and UAE samples were higher than recoveries from the controls in all soy matrices. Under all extraction conditions, Laemmli and urea buffer recovered more proteins than PBS. Electrophoresis analysis of protein showed bands around 75, 50, and 33 kDa indicating the presence of soy allergenic proteins β‐conglycinin and glycinin, in all samples. Using sandwich ELISA, control and UAE extracts resulted in high soy protein detection but this reduced in MAE extracts.     

Effect of roasting history and buffer composition on peanut protein extraction efficiency

Peanut is a major allergenic food. Undeclared peanut (allergens) from mis-formulation or contamination during food processing pose a potential risk for sensitized individuals and must be avoided. Reliable detection and quantification methods for food allergens are necessary in order to ensure compliance with food labelling and to improve consumer protection. The extraction of proteins from allergenic foods and complex food products is an important step in any allergen detection method. In this study, the protein extraction efficiency of various buffers prepared in-house and some extraction buffers included in some commercial allergen enzyme-linked immunosorbent assay (ELISA) test kits for peanut determination in food products were tested. In addition, the effect of roasting history on the extractability of peanut protein was investigated by the biuret and the bicinchoninic acid (BCA) assays. Elevated roasting temperatures in food processing were found to have a major impact on protein extraction efficiency by reducing protein yields of oil and dry roasted peanuts by 50-75% and 75-80%, respectively, compared with the raw material. Extraction buffers operating in the higher pH range (pH 8-11) showed best yields.     

中式油炸食品中丙烯酰胺测定及降低方法初探

研究高效液相色谱法分析测定中式油炸食品中丙烯酰胺方法。样品预处理条件:油炸样品粉碎后用0.1%甲酸水溶液(料液比1:5)进行三次重复提取,提取液经高速离心、冷冻、C_(18)固相萃取小柱纯化,最后采用高效液相色谱仪进行测定。测定色谱条件:流动相为甲醇—0.02 mol/L乙酸胺溶液(5:95,v/v)、流速为0.9 ml/min、进样量为20μl、保留时间约4.887 min,检测器为紫外检测器,波长为210 nm、柱温25℃±0.5℃;试验结果表明,丙烯酰胺色谱图峰面积与其浓度在0.5～5.0μg/mL范围内呈良好线性关系,其最低检出限为10 ng/mL,回收率为94.5%～106.5%,相对标准偏差3.2%～4.5%;该法回收率高、精密度好,重复性好。利用该法测定几类市售中式油炸食品和自制油炸食品中丙烯酰胺含量,结果表明,市售中式油炸食品丙烯酰胺检出率为100%,仅含量水平有较大差异;试验也表明,通过优化工艺技术条件和配方如调节pH值或添加抗氧化剂等可降低油炸食品中丙烯酰胺含量。     

大豆过敏原夹心ELISA和竞争ELISA方法的建立及其在加工食品中应用研究

本文以大豆混合过敏原为目标,建立了快速、便捷检测大豆过敏原的夹心酶联免疫吸附方法（sandwich-enzyme linked immunosorbent assay）和间接竞争酶联免疫吸附方法（indirect competitive enzyme-linked immunosorbent assay）,通过实际加工样品的回收实验、加标食品回收实验以及对真实食物样本的检测,对这两种方法进行了比较,确定了各自的适用范围。结果表明,夹心ELISA方法标准品浓度在0.0078~30 μg/mL范围内呈现出良好的线性关系,曲线方程为y=0.2333x+0.0692,决定系数R2=0.995。竞争ELISA方法的检测范围为10~100000 ng/mL,最低检测限为10 ng/mL。对购入橙汁进行加标回收实验,夹心ELISA检测后的回收率要高于竞争ELISA检测后的回收率,达100%以上;而对成分和加工方式都比较复杂的巧克力、牛肉酱、面包或蛋糕来说,竞争ELISA检测后的回收率要高于夹心ELISA检测后的回收率。对发酵类食物进行检测,竞争ELISA方法检测到的浓度要高于夹心ELISA,而对成分比较简单的食物比如芝麻糊、豆奶等进行检测时,夹心ELISA的检测浓度要略高于竞争ELISA。综上,竞争ELISA方法更适用于食物基质复杂,经过深度加工的食品,而夹心ELISA方法更适用于食物成分简单,轻加工后的食品,两种方法在各自的适用范围内均能实现较准确的检测。     

Quantitative analysis of shrimp allergen in food matrices using a protein chip based on sandwich immunoassay

Food allergy is increasingly becoming a serious concern these days. With packaged foods becoming the norm of the day, food allergy cases out of accidental consumption are becoming rampant, thereby generating great risks for the subjects involved and prompting food authorities in different countries to formulate new regulations about displaying food allergen data on food labels. Detection of food allergens is conventionally carried out by ELISA or PCR tests. These techniques are limited in that they can only detect one or few allergens at one time. Therefore, in the present study a novel sandwich protein chip assay was developed for quantitation of shrimp allergens in food matrixes. The shrimp allergen model used 3D aldehyde slides as the solid carrier, rabbit antisera as the capture reagent, and biotin-labeled monoclonal antibody as the detector reagent. Resulting antigen–antibody complexes were visualized in the presence of commercial strepavidin labeled with Cy3 to produce fluorescence for quantification. With the LOD of the protein chip being 0.054 mg tropomyosin/kg, the protein chip can quantify down to 0.096 mg tropomyosin/kg. The protein chip was not found to be sensitive to other kinds of foods but cross-reacted to some extent with allergens of some other crustaceans. The recoveries ranged from 69.2 to 99.9%, while the intra- and inter-assay coefficients of variation were <13% and <19%, respectively. It seems that the new assay is reliable enough to detect shrimp allergens in food and food products and help minimize the instances of shrimp allergy. It is also possible to use the protein chip for simultaneous detection of other food allergens.     

超高效液相色谱-串联质谱法测定肉制品中的虾过敏原

目的：建立超高效液相色谱-串联质谱（ultra performance liquid chromatography-tandem mass spectrometry,UPLC-MS/MS）法检测肉制品中虾过敏原的定量方法。方法：选取基质较为复杂的肉制品（西式火腿、香肠和肉丸）作为研究对象。样品经磷酸盐缓冲液（Phosphate buffer solution,PBS）超声提取30 min,离心（10000 r/min,15℃,10 min）后加入乙腈去除脂肪,取样液加入内标肽段（2.5μmol/L,40μL）和胰蛋白酶（1 mg/mL,10μL）在37℃下酶解16 h后上液质进行分析,样品经T3柱进行分离,0.1%甲酸-水溶液和乙腈梯度洗脱,多反应监测（multiple reaction monitoring,MRM）正离子模式采集数据,内标法定量。结果：采用该方法测定肉制品中虾过敏原蛋白含量,其中定量肽段在0.001～2.0μmol/L范围内,线性关系良好,决定系数R2为1.0000,检出限为0.67 mg/kg,定量限为2.00 mg/kg;在三个加标浓度水平下,回收率为83.2%...     

食品中常见过敏原及检测技术研究进展

近年来，食物过敏发生率呈现上升趋势，由食物过敏引发的食品安全问题引起广泛关注。目前对于食物过敏尚无有效治疗手段，避免摄入含过敏原食物是最有效的预防方式。因此，过敏原检测与标识对过敏人群具有重要的警示意义。本文介绍了8 类常见致敏食品中主要过敏原的结构与致敏特点，综述了现阶段用于过敏原检测的主要技术，包括基于蛋白水平的免疫学检测技术、基因水平的分子生物学检测技术以及质谱技术，分析了各方法的优势与局限性，并对过敏原检测技术的发展方向进行了展望。     

电化学分析法检测动物源食品中氯丙嗪的3种前处理方法比较

目的对猪肉、鸡肉、牛肉、猪肝、虾肉、蜂 蜜、鱼肉、鸡蛋和牛奶共9种动物源食品中氯丙嗪电化学检测的前处理方法进行了选择和优化,方法 样品分别采用溶剂提取法、QuECHERS法（Quick、Easy、Cheap、Effective、Rugged、Safe,QuEChERS法）和固相萃取法（solid-phase extraction,SPE）进行了电化学检测比较分析。结果表明,猪肉、鸡肉、牛肉、猪肝、虾肉和蜂蜜可用溶剂提取法进行处理,氯丙嗪的加标回收率均在81.6%以上;而鱼肉和鸡蛋样品需采用QuECHERS法进行处理,回收率分别达到88.5%～95.8%和89.7%～98.2%;牛奶样品用溶剂提取法和QuECHERS 法处理时均有基质干扰,采用固相萃取法进行净化后,回收率达到80.6%～93.0%。不同类型的动物源食品由于肌肉的纤维组织、脂肪含量、水分含量、蛋白质的种类和含量等存在差别,需分别采用适宜的前处理方法才能获得满意的测定结果。结论 该研究可为不同类型动物源食品中氯丙嗪电化学检测方法的应用提供参考。