Article(id=1151437190573159020, tenantId=1146029695717560320, journalId=1149652044408987649, issueId=1151437189243089177, articleNumber=null, orderNo=null, doi=10.19812/j.cnki.jfsq11-5956/ts.20250317002, pmid=null, cstr=null, oa=null, hot=null, price=null, onlineType=0, articleFormat=0, articleType=null, articleTypeStr=null, receivedDate=1742140800000, receivedDateStr=2025-03-17, revisedDate=null, revisedDateStr=null, acceptedDate=null, acceptedDateStr=null, onlineDate=1752453618900, onlineDateStr=2025-07-14, pubDate=1749916800000, pubDateStr=2025-06-15, doiRegisterDate=null, doiRegisterDateStr=null, onlineIssueDate=1752453618900, onlineIssueDateStr=2025-07-14, onlineJustAcceptDate=null, onlineJustAcceptDateStr=null, onlineFirstDate=null, onlineFirstDateStr=null, sourceXml=null, magXml=null, createTime=1752453618900, creator=13701087609, updateTime=1752453618900, updator=13701087609, issue=Issue{id=1151437189243089177, tenantId=1146029695717560320, journalId=1149652044408987649, year='2025', volume='16', issue='11', pageStart='1', pageEnd='320', issueExtLink='null', onlineDate='null', pubDate='null', beforeIssueId=null, nextIssueId=null, price=null, status=1, issueComplete=1, articleOrder=1, issueType=-1, specialIssue=0, createTime=1752453618584, creator=13701087609, updateTime=1767768054466, updator=13701087609, preIssue=null, nextIssue=null, ext={EN=IssueExt(id=1215670588966883492, tenantId=1146029695717560320, journalId=1149652044408987649, issueId=1151437189243089177, language=EN, specialIssueTitle=, coverIllustrator=null, specialIssueEditor=, specialIssueAbout=), CN=IssueExt(id=1215670588966883493, tenantId=1146029695717560320, journalId=1149652044408987649, issueId=1151437189243089177, language=CN, specialIssueTitle=, coverIllustrator=null, specialIssueEditor=, specialIssueAbout=)}, issueFiles=null}, startPage=140, endPage=145, ext={EN=ArticleExt(id=1151895328706932760, articleId=1151437190573159020, tenantId=1146029695717560320, journalId=1149652044408987649, language=EN, title=Rapid determination of thiabendazole residue in dairy products by molecular fluorescence differential addition method, columnId=1151895322692776479, journalTitle=Journal of Food Safety & Quality, columnName=Special Topic: Analysis and Monitoring of Toxic and Harmful Substances in Food, runingTitle=null, highlight=null, articleAbstract=

Objective To explore a method for rapid detection of thiabendazole residue in dairy products using molecular fluorescence differential addition method. Methods The optimal experimental conditions for the molecular fluorescence differential addition method were determined through single factor experiments, and this method was applied to determine the content of thiabendazole in dairy products. Results The residue of thiabendazole in dairy products ranged from 0.1580 to 0.1611 mg/kg, with a limit of detection was 0.0018 μg/mL and a limit of quantification was 0.0060 μg/mL. The relative standard deviation of the results was 0.75% (n=6), and the recovery rates ranged from 99.4% to 106.7%. A comparison with GB 23200.87—2016 National standards for food safety-Determination of thiabendazole residues in milk and dairy products-Fluorescence spectrophotometry showed no significant differences between the two methods based on F-test and t-test analysis. Conclusion This method eliminates the need to construct a calibration curve and measure blank solutions, offering advantages such as simplicity, sensitivity, rapidity, high recovery rates and accurate results. The proposed method is suitable for the rapid detection of thiabendazole residue in dairy products and provides a new detection technique for the determination of thiabendazole residues in dairy products.

, correspAuthors=Nan-Nan GUO, authorNote=null, correspAuthorsNote=null, copyrightStatement=null, copyrightOwner=null, extLink=null, articleAbsUrl=null, sourceXml=null, magXml=null, pdfUrl=null, pdf=null, pdfFileSize=null, pdfExtLink=null, richHtmlUrl=null, mobilePdfUrl=null, reviewReport=null, pdfFirstPage=null, abstractGraph=null, abstractGraphContent=null, abstractVideo=null, citation=null, cebUrl=null, magXmlContent=null, mapNumber=null, authorCompany=null, fund=null, authors=null, authorsList=Nan-Nan GUO), CN=ArticleExt(id=1151895333752680591, articleId=1151437190573159020, tenantId=1146029695717560320, journalId=1149652044408987649, language=CN, title=分子荧光差异加标法快速测定乳品中的噻菌灵残留量, columnId=1151895322898297380, journalTitle=食品安全质量检测学报, columnName=本期专题:食品中有毒有害物质分析与监测, runingTitle=null, highlight=null, articleAbstract=

目的 建立分子荧光差异加标法快速检测乳品中的噻菌灵残留的方法。方法 通过单因素试验确定分子荧光差异加标法的最佳试验条件并将其应用与乳品中噻菌灵的测定。结果 乳品中噻菌灵残留含量为0.1580~0.1611 mg/kg, 检出限为0.0018 μg/mL, 定量限为0.0060 μg/mL, 测定结果的相对标准偏差为0.75% (n=6), 加标回收率为99.4%~106.7%。与GB 23200.87—2016《食品安全国家标准 乳及乳制品中噻菌灵残留量的测定 荧光分光光度法》作对比, 经过F检验和t检验分析, 两者之间不存在显著性差异。结论 该方法无需绘制工作曲线和测定空白溶液, 具有操作简便、灵敏、快速、加标回收率高和结果准确等优点, 适用于乳品中噻菌灵残留的快速检测, 为测定乳品中噻菌灵残留提供了一种新的检测技术。

, correspAuthors=郭楠楠, authorNote=null, correspAuthorsNote=
* 郭楠楠(1986—), 女, 硕士, 副教授, 主要研究方向为食品安全快速检测分析新技术研究。E-mail:
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Development and application of method for determinatiaon of benzimidazoe residues in animal-derived foods[D]. Taian: Shandong Agricultural University, 2019., articleTitle=null, refAbstract=null)], funds=[Fund(id=1167036543953809587, tenantId=1146029695717560320, journalId=1149652044408987649, articleId=1151437190573159020, awardId=21B550008, language=CN, fundingSource=河南省高等学校重点科研项目(21B550008), fundOrder=null, country=null)], companyList=[AuthorCompany(id=1167036542460637337, tenantId=1146029695717560320, journalId=1149652044408987649, articleId=1151437190573159020, xref=null, ext=[AuthorCompanyExt(id=1167036542469025946, tenantId=1146029695717560320, journalId=1149652044408987649, articleId=1151437190573159020, companyId=1167036542460637337, language=EN, country=null, province=null, city=null, postcode=null, companyName=null, departmentName=null, remark=Zhengzhou City Food Safety Rapid Test Key Laboratory, College of Food Science and Engineering, Zhengzhou University of Science and Technology, Zhengzhou 450064, China), AuthorCompanyExt(id=1167036542473220251, tenantId=1146029695717560320, journalId=1149652044408987649, articleId=1151437190573159020, companyId=1167036542460637337, language=CN, country=null, province=null, city=null, postcode=null, companyName=null, departmentName=null, remark=郑州科技学院食品科学与工程学院, 郑州市食品安全快速检测重点试验室, 郑州 450064)])], figs=[ArticleFig(id=1167036543303692457, tenantId=1146029695717560320, journalId=1149652044408987649, articleId=1151437190573159020, language=EN, label=Fig.1, caption=Effects of sample volume (A), extraction frequency (B), reaction time (C), thiabendazole spiked concentration (D) on fluorescence intensity (n=6), figureFileSmall=0uvPVlF1e4rcmsKDYfrOhQ==, figureFileBig=gb/GAFQWONvrY9eGBDcPTg==, tableContent=null), ArticleFig(id=1167036543358218410, tenantId=1146029695717560320, journalId=1149652044408987649, articleId=1151437190573159020, language=CN, label=图1, caption=样品体积(A)、萃取次数(B)、反应时间(C)、加标质量浓度(D)对乳品样品荧光强度的影响(n=6)

注: 图中不同英文字母表示差异达到显著水平, P<0.05。

, figureFileSmall=0uvPVlF1e4rcmsKDYfrOhQ==, figureFileBig=gb/GAFQWONvrY9eGBDcPTg==, tableContent=null), ArticleFig(id=1167036543412744363, tenantId=1146029695717560320, journalId=1149652044408987649, articleId=1151437190573159020, language=EN, label=Table 1, caption=

Extraction yield under the different extraction times (n=6)

, figureFileSmall=null, figureFileBig=null, tableContent=
萃取次数 荧光值 萃取率/%
1 34.62 94.6
2 34.42 98.4
3 34.45 98.1
4 34.44 98.2
5 34.44 98.2
), ArticleFig(id=1167036543479853228, tenantId=1146029695717560320, journalId=1149652044408987649, articleId=1151437190573159020, language=CN, label=表1, caption=

不同萃取次数下的萃取率(n=6)

, figureFileSmall=null, figureFileBig=null, tableContent=
萃取次数 荧光值 萃取率/%
1 34.62 94.6
2 34.42 98.4
3 34.45 98.1
4 34.44 98.2
5 34.44 98.2
), ArticleFig(id=1167036543542767789, tenantId=1146029695717560320, journalId=1149652044408987649, articleId=1151437190573159020, language=EN, label=Table 2, caption=

Determination of the addition of standard volume (n=6)

, figureFileSmall=null, figureFileBig=null, tableContent=
样品 加标量/mL 样品噻菌灵质量
浓度/(μg/mL)
样品噻菌灵平均质量浓度/(μg/mL)
乳品 1.00、2.00 0.1345 0.1412
2.00、3.00 0.1289
3.00、4.00 0.1598
4.00、5.00 0.1376
5.00、6.00 0.1454
), ArticleFig(id=1167036543605682350, tenantId=1146029695717560320, journalId=1149652044408987649, articleId=1151437190573159020, language=CN, label=表2, caption=

加标体积的确定(n=6)

, figureFileSmall=null, figureFileBig=null, tableContent=
样品 加标量/mL 样品噻菌灵质量
浓度/(μg/mL)
样品噻菌灵平均质量浓度/(μg/mL)
乳品 1.00、2.00 0.1345 0.1412
2.00、3.00 0.1289
3.00、4.00 0.1598
4.00、5.00 0.1376
5.00、6.00 0.1454
), ArticleFig(id=1167036543676985519, tenantId=1146029695717560320, journalId=1149652044408987649, articleId=1151437190573159020, language=EN, label=Table 3, caption=

Determination of thiabendazole in sample

, figureFileSmall=null, figureFileBig=null, tableContent=
噻菌灵残留含量/(μg/g) 平均值
/(μg/g)
S/(μg/g) RSDs/%
标准曲线法 0.1622、0.1616、0.1608、0.1608、0.1604、0.1620、 0.1621 0.0001 0.42
差异加标法 0.1596、0.1611、0.1580、0.1589、0.1600、0.1608、 0.1599 0.0012 0.75
), ArticleFig(id=1167036543731511472, tenantId=1146029695717560320, journalId=1149652044408987649, articleId=1151437190573159020, language=CN, label=表3, caption=

样品中噻菌灵的测定值

, figureFileSmall=null, figureFileBig=null, tableContent=
噻菌灵残留含量/(μg/g) 平均值
/(μg/g)
S/(μg/g) RSDs/%
标准曲线法 0.1622、0.1616、0.1608、0.1608、0.1604、0.1620、 0.1621 0.0001 0.42
差异加标法 0.1596、0.1611、0.1580、0.1589、0.1600、0.1608、 0.1599 0.0012 0.75
), ArticleFig(id=1167036543777648817, tenantId=1146029695717560320, journalId=1149652044408987649, articleId=1151437190573159020, language=EN, label=Table 4, caption=

Results of comparison test

, figureFileSmall=null, figureFileBig=null, tableContent=
样品 噻菌灵含量/(mg/kg)
分子荧光差异加标法 GB 23200.87—2016法
平均值 0.1599 0.1621
S 0.0012 0.0001
RSDs/% 0.75 0.42
), ArticleFig(id=1167036543836369074, tenantId=1146029695717560320, journalId=1149652044408987649, articleId=1151437190573159020, language=CN, label=表4, caption=

对照试验测定结果

, figureFileSmall=null, figureFileBig=null, tableContent=
样品 噻菌灵含量/(mg/kg)
分子荧光差异加标法 GB 23200.87—2016法
平均值 0.1599 0.1621
S 0.0012 0.0001
RSDs/% 0.75 0.42
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分子荧光差异加标法快速测定乳品中的噻菌灵残留量
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郭楠楠 *
食品安全质量检测学报 | 本期专题:食品中有毒有害物质分析与监测 2025,16(11): 140-145
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食品安全质量检测学报 | 本期专题:食品中有毒有害物质分析与监测 2025, 16(11): 140-145
分子荧光差异加标法快速测定乳品中的噻菌灵残留量
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郭楠楠*
作者信息
  • 郑州科技学院食品科学与工程学院, 郑州市食品安全快速检测重点试验室, 郑州 450064

通讯作者:

* 郭楠楠(1986—), 女, 硕士, 副教授, 主要研究方向为食品安全快速检测分析新技术研究。E-mail:
Rapid determination of thiabendazole residue in dairy products by molecular fluorescence differential addition method
Nan-Nan GUO*
Affiliations
  • Zhengzhou City Food Safety Rapid Test Key Laboratory, College of Food Science and Engineering, Zhengzhou University of Science and Technology, Zhengzhou 450064, China
出版时间: 2025-06-15 doi: 10.19812/j.cnki.jfsq11-5956/ts.20250317002
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目的 建立分子荧光差异加标法快速检测乳品中的噻菌灵残留的方法。方法 通过单因素试验确定分子荧光差异加标法的最佳试验条件并将其应用与乳品中噻菌灵的测定。结果 乳品中噻菌灵残留含量为0.1580~0.1611 mg/kg, 检出限为0.0018 μg/mL, 定量限为0.0060 μg/mL, 测定结果的相对标准偏差为0.75% (n=6), 加标回收率为99.4%~106.7%。与GB 23200.87—2016《食品安全国家标准 乳及乳制品中噻菌灵残留量的测定 荧光分光光度法》作对比, 经过F检验和t检验分析, 两者之间不存在显著性差异。结论 该方法无需绘制工作曲线和测定空白溶液, 具有操作简便、灵敏、快速、加标回收率高和结果准确等优点, 适用于乳品中噻菌灵残留的快速检测, 为测定乳品中噻菌灵残留提供了一种新的检测技术。

噻菌灵  /  分子荧光  /  差异加标法  /  乳品

Objective To explore a method for rapid detection of thiabendazole residue in dairy products using molecular fluorescence differential addition method. Methods The optimal experimental conditions for the molecular fluorescence differential addition method were determined through single factor experiments, and this method was applied to determine the content of thiabendazole in dairy products. Results The residue of thiabendazole in dairy products ranged from 0.1580 to 0.1611 mg/kg, with a limit of detection was 0.0018 μg/mL and a limit of quantification was 0.0060 μg/mL. The relative standard deviation of the results was 0.75% (n=6), and the recovery rates ranged from 99.4% to 106.7%. A comparison with GB 23200.87—2016 National standards for food safety-Determination of thiabendazole residues in milk and dairy products-Fluorescence spectrophotometry showed no significant differences between the two methods based on F-test and t-test analysis. Conclusion This method eliminates the need to construct a calibration curve and measure blank solutions, offering advantages such as simplicity, sensitivity, rapidity, high recovery rates and accurate results. The proposed method is suitable for the rapid detection of thiabendazole residue in dairy products and provides a new detection technique for the determination of thiabendazole residues in dairy products.

thiabendazole  /  molecular fluorescence  /  differential standard addition method  /  dairy products
郭楠楠. 分子荧光差异加标法快速测定乳品中的噻菌灵残留量. 食品安全质量检测学报, 2025 , 16 (11) : 140 -145 . DOI: 10.19812/j.cnki.jfsq11-5956/ts.20250317002
Nan-Nan GUO. Rapid determination of thiabendazole residue in dairy products by molecular fluorescence differential addition method[J]. Journal of Food Safety & Quality, 2025 , 16 (11) : 140 -145 . DOI: 10.19812/j.cnki.jfsq11-5956/ts.20250317002
噻菌灵(thiabendazole, TBZ), 化学名称为2-(1,3-噻唑-4-萘)苯并咪唑, 是高效性、广谱性、低毒性、持效期长和最早市场化的的内吸性杀菌剂。既可用于人、畜肠道的驱虫药剂, 又可用于工业防霉剂以及多种作物的真菌病害防治及果蔬的防腐保鲜, 在农药、医药及工业领域有重要应用[1-3]
很多国家对苯并咪唑类杀菌剂的最大残留限量(maximum residue limit, MRL)都有严格的规定[4], 其中GB 2763—2021《食品安全国家标准 食品中农药最大残留限量》中规定乳品中噻菌灵的最大残留限量标准为0.2 mg/kg。而乳及乳制品是人们日常生活中不可或缺的, 因此对于乳品中的苯并咪唑类杀菌剂的检测非常重要。
目前检测苯并咪唑类药物的方法有液相色谱法[5]、液相色谱-质谱法[6-7]、毛细管电泳-质谱法[8]、荧光分析法[9-10]、免疫分析法[11-12]、气相色谱-质谱法[13-15]及一些快速分析检测法[16-19]等。液相色谱方法灵敏度较高, 能够满足筛选检测需要, 但分离效果不理想, 分析时间长; 质谱方法因价格及维护费用较贵, 不能普及; 气相色谱法需要将样品衍生后才能进行检测, 操作复杂, 且常需与其他方法联用, 对于苯并咪唑类杀菌剂而言, 熔点相对偏低。气相色谱-质谱法由于苯并咪唑类药物难以气化, 需衍生后测定。而生物法因为检出限太高, 不能达到检测要求[20-22]
荧光分光光度差异加标法是在相同体积的样品溶液背景条件下加入同质量浓度、不同体积的标准溶液后测定被检测物质, 实现了同时测定样品和标准溶液, 能够有效扣除环境、试液背景, 减少了荧光的干扰, 消除了一些反应的干扰[23]。该方法不需要测定空白、绘制标准曲线, 灵敏度较高、准确度高、快速简便、避免引入较大误差, 操作简便, 快速准确, 成本较低, 检出限低、荧光化学物质稳定, 样品和试剂用量少且无需过柱除杂, 工作效率得到极大提高[24]。但目前鲜少见文献报道将其用于测定乳品中的噻菌灵残留, 本研究拟采用分子荧光差异加标法测定乳品中的噻菌灵残留, 以期为乳品中噻菌灵残留的检测提供一种相对更简便、快速、准确的方法。
970CRT型荧光分光光度计(配有10 mm四通石英比色皿一个, 上海仪电分析仪器有限公司); KQ-5200B机械型超声波清洗仪器(东莞市科桥超声设备有限公司); JA2003A万分之一分析天平(上海豪晟科学仪器有限公司); 101-0ABS型电热鼓风干燥箱(西安莫吉娜仪器制造有限公司); HH-6型数显恒温水浴箱(金坛市华峰仪器有限公司); QL-901型涡旋仪(北京海天友诚科技有限公司); DDSJ-318型电导率仪(上海洪纪仪器设备有限公司)。
乳品, 产地河南省, 购于郑州市某超市。
噻菌灵标准品(分析纯, 上海麦克林生化科技有限公司); 氢氧化钾(分析纯, 郑州派尼化学试剂厂); 盐酸(分析纯, 苏州化学试剂有限公司); 乙酸乙酯(分析纯, 天津市德恩化学试剂有限公司); 试验用水为4.5 L/桶的怡宝饮用纯净水, 电导率为2.82 μS/cm。
(1)皂化。用分析天平称取乳品试样10.0000 g(精确至0.0001 g)于圆底烧瓶中, 加入7 mL 50% (m/V)氢氧化钾溶液, 在烧瓶上端接上冷凝管(下端进水、上端出水, 水速均匀), 在沸腾的水浴上皂化回流40 min, 取下, 充分冷却。
(2)提取。将烧瓶里的溶液转入125 mL分液漏斗中, 用10 mL水洗涤圆底烧瓶, 洗液也转入分液漏斗。加15 mL乙酸乙酯于分液漏斗中, 轻摇0.5 min, 静置分层。将下层溶液转入另一分液漏斗, 取15 mL乙酸乙酯溶液提取下层溶液, 剧烈振摇1 min, 静置分层, 舍去下层溶液, 合并两次乙酸乙酯提取液。
(3)净化。用吸量管取20 mL 0.05% (m/V)氢氧化钾溶液于分液漏斗中洗涤乙酸乙酯提取液, 剧烈振摇1 min, 静置分层后, 舍去下层溶液。再向此分液漏斗中加入20 mL 0.05% (m/V)氢氧化钾溶液轻摇洗涤乙酸乙酯提取液, 舍去水层。用10 mL盐酸溶液提取乙酸乙酯层。合并盐酸提取液于10 mL容量瓶中, 并用0.1 mol/L的盐酸溶液定容。待用荧光分光光度计测定[25-27]
噻菌灵待测样品: 用移液枪向3个10 mL刻度比色管中分别加入3份1.00 mL样品待测液, 在比色管上写上编号1、2、3。向第1份和第2份待测溶液中分别加0.15 μg/mL噻菌灵标准溶液3.00 mL和4.00 mL, 第3份不加噻菌灵标准溶液, 3份均用0.1 mol/L盐酸溶液定容至刻度, 待仪器检测。在完全相同条件下进行3次平行测定, 检验无可疑值后取平均值, 按式(1)~(4)计算样品中噻菌灵残留的含量。
Ix+1=K(ρx+ρs1)
Ix+2=K(ρx+ρs2)
${{\rho }_{x}}=\frac{{{I}_{x+1}}\times {{\rho }_{s2}}-{{I}_{x+2}}\times {{\rho }_{s1}}}{{{I}_{x+2}}-{{I}_{x+1}}}$
式中: Ix+1, Ix+2为加标过滤液的相对荧光强度; ρx为样品中噻菌灵残留的质量浓度, μg/mL; ρs1, ρs2为两个加标分析液中因加标增加的噻菌灵残留的质量浓度, μg/mL, ρs2>ρs1
ω=$\frac{{{\rho }_{x}}\times V}{m}$
式中: ω为样品中噻菌灵残留的含量, μg/g; V为样品试液的体积, mL; m为样品质量, g。
参考GB 23200.87—2016《乳及乳制品中噻菌灵残留量的测定 荧光分光光度法》, 选择最大激发波长307 nm, 最大发射波长359 nm, 灵敏度为2, 扫描速度高速, 激发波长为狭缝10 nm, 发射波长狭缝为10 nm作为测定样品时的仪器条件。
实验数据平行测定6次, 用SPSS 2021进行显著性分析, 用WPS Office、Origin 8.0软件制表及绘图, 结果以平均值±标准偏差表示。
首先向5个10 mL棕色容量瓶中分别加入10.00、15.00、25.00、50.00和100.00 μL的100 μg/mL噻菌灵标准溶液, 其次用0.1 mol/L盐酸溶液进行定容, 配制成质量浓度分别为0.10、0.15、0.25、0.50和1.00 μg/mL噻菌灵标准待测溶液。在EX=307 nm、EM=359 nm, 灵敏度为2, 扫描速度高速, 激发波长为狭缝10 nm, 发射波长狭缝为10 nm的条件下, 对噻菌灵系列标准溶液的相对荧光强度进行测定。以噻菌灵的质量浓度为横坐标(X, μg/mL), 以噻菌灵对应的相对荧光强度为纵坐标(Y), 得出噻菌灵标准溶液的回归直线方程为Y=55.801X+24.378, r2=0.9994。在测定浓度范围(0.1~1.0 μg/mL)内标准曲线线性良好。
在与标准曲线一致的条件下测定样品分析液, 同时进行空白测定, 由绘制标准曲线所得到的回归直线方程计算出样液中噻菌灵的质量浓度。按公式(5)计算噻菌灵的残留含量。
X=CV/m
式中: X为样品中噻菌灵的含量, mg/kg; C为通过标准曲线计算所得的样品中噻菌灵残留的质量浓度, μg/mL; V为定容后待测液的体积, mL; m为样品的质量, g。
为了防止比色皿发生荧光污染, 因此在开始试验前将比色皿洗净后, 放在8 mol/L的硝酸中[28]浸泡0.5 h, 依次用蒸馏水、怡宝水冲洗后再进行使用。将比色皿内外壁的水用不含荧光物质的吸水纸吸干, 在不加任何溶液的条件下测定其荧光值, 发现在EX为307 nm、EM为359 nm条件下, 空的比色皿也有荧光值, 6次平行测定结果为21.83, 标准偏差为0.063, 在做试验时应尽可能避免比色皿荧光的干扰。
称取乳品样品, 按照1.3步骤进行处理, 分别移取1.00、2.00、3.00、4.00、5.00、6.00、7.00、8.00、9.00、10.00 mL样品分析液于10个10 mL具塞比色管中, 用0.1 mol/L盐酸溶液定容至刻度。在EX为307 nm、EM为359 nm条件下进行测定, 测定结果如图1A
图1A可知, 样品的相对荧光强度随着加入样品体积的增加量呈现先增加后下降的趋势, 由此可得定容时加入8 mL样品分析液, 用0.1 mol/L盐酸溶液定容至刻度时, 样品的荧光值达到最大, 为33.42。因此将样品的体积确定为8 mL。
将步骤1.3中净化舍弃的水层和乙酸乙酯层混合在一起并入分液漏斗中, 进行再次净化、萃取、静止, 将得到盐酸层定容于10 mL容量瓶, 再在EX为307 nm、EM为359 nm条件下进行测定, 重复上述操作步骤得到如图1B试验结果。由图1B可知, 第一次萃取样品的荧光值小于后面4次萃取的荧光值, 并且萃取两次后荧光值开始逐渐稳定, 因此在对样品净化时需进行2~3次萃取。
将处理后的样品用0.1 mol/L盐酸溶液定容后, 及时计时, 设定反应时间分别为0、5、10、20、30、40、50 min, 反应后在设定的仪器条件下对样品的荧光强度进行测定, 试验结果如图1C。由图1C可得出: 随着反应时间的增加, 样品待测液的相对荧光强度无明显波动、呈平缓变化趋势, 由此可得反应时间的长短对样品的荧光强度并无太大影响。
称取乳品样品, 按照1.3步骤进行处理, 分别移取1.00 mL样品液于12个10 mL具塞比色管中, 依次分别加入1.00、2.00、3.00、4.00、5.00和6.00 mL的0.15 μg/mL和0.25 μg/mL噻菌灵标准溶液, 并用0.1 mol/L盐酸溶液的溶液定容。在最大激发波长307 nm, 最大发射波长359 nm, 灵敏度为2的条件下进行测定, 结果如下图1D。由图1D可知, 样品的相对荧光强度随着加入噻菌灵标准溶液体积的增加呈现先增加后下降的趋势。并且在加入0.15 μg/mL噻菌灵标准溶液时的荧光值大于0.25 μg/mL噻菌灵标准溶液的荧光值, 因此选择0.15 μg/mL噻菌灵标准溶液作为加标质量浓度。
为检验是否完全萃取, 对上述试验萃取后的水溶液再次重复上述的萃取操作, 其他的依次操作, 测定荧光值求得萃取率, 结果如表1。由表1可知, 样品的萃取率在94.6%~98.4%之间。
称取乳品样品, 按照1.3.1步骤进行处理, 分别移取1.00 mL样品液于6个10 mL具塞比色管中, 依次分别加入1.00、2.00、3.00、4.00、5.00和6.00 mL的0.15 μg/mL噻菌灵标准溶液, 并用0.1 mol/L盐酸溶液的溶液定容。在最大激发波长307 nm, 最大发射波长359 nm, 灵敏度为2的条件下进行测定, 按照式(1)、(2)计算乳品中噻菌灵的质量浓度(μg/mL)[29]。由表2可知, 在加标量为3.00 mL、4.00 mL时, 样品中噻菌灵的的质量浓度和国标法最接近, 因此, 本试验选择0.15 μg/mL噻菌灵标准溶液添加量3.00 mL、4.00 mL作为最佳加标量。
按试验方法, 在最合适的仪器工作条件下对样品中噻菌灵的相对荧光强度进行测定, 分别利用标准曲线法和差异加标法对样品进行6次平行对照测定, 通过检验没有可疑值后, 然后由公式进行计算, 测定结果和标准偏差(S)、相对标准偏差(relative standard deviation, RSD), 结果见下表3。由表3的结果可以看出测定结果的RSD均小于1.0%, 表明该方法精密度较好, 并且可以得出标准曲线法和分子荧光差异加标法所测得结果差异较小, 因此可以认为分子荧光差异加标法适用于对乳品中噻菌灵的残留进行测定。
对质量浓度为0.10 μg/mL的噻菌灵标准溶液分别进行11次平行试验测定, 由工作曲线方程获得结果, 按3倍S/N计算噻菌灵的最低检出限0.0018 μg/mL, 按10倍S/N计算噻菌灵定量限0.0018 μg/mL, 此方法低于GB 23200.87—2016规定的检出限, S为样品测定结果的标准偏差[30]
准确移取经步骤1.3.1所处理的样品分析液8.00 mL于10 mL容量瓶中, 分别加入质量浓度为0.15 μg/mL的噻菌灵标液1.00、2.00、3.00、4.00 mL, 用0.1 mol/L盐酸溶液定容, 对以上溶液分别进行6次平行对照测定, 通过检验没有可疑值后, 计算加标回收率[31]。样品加标回收率为99.4%~106.7%, 符合GB/T 27404—2008《试验室质量控制规范 食品理化检测》中的相关规定。
通过比较分子荧光差异加标法、GB 23200.87—2016的标准曲线法的平均值、标准偏差(S)、RSD、F检验和t检验的结果来判断这两种方法是否有显著差异, 两种方法的测定结果如表4所示。
表4可知, 分子荧光差异加标法和GB 23200.87—2016法中的标准曲线法的平均值相差不大, 这两种方法的相对标准偏差均小于1.0%。通过对测定结果进行F检验和t检验, 结果表明两种方法之间不存在显著性差异, 结论的置信度为95%。
将荧光分光光度法与差异加标法分析方法结合, 对乳品中噻菌灵残留的含量进行测定, 在最佳仪器工作条件下测定乳品中噻菌灵残留的含量为0.1580~0.1611 mg/kg, 噻菌灵的检出限为0.0018 μg/mL, 定量限为0.0060 μg/mL, 测定的噻菌灵加标回收率为99.4%~106.7%, 测定的RSD小于1.0%。分子荧光差异加标法与GB 23200.87—2016对照, 通过F检验和t检验结果表明: 两种方法的相对标准偏差和平均值均不存在显著性差异。此方法具有灵敏度高、检出限低、受基质干扰影响小、荧光化学物质稳定、荧光强度高的特点。分子荧光差异加标法为检测乳品中噻菌灵的残留提供了一种新的分析技术。
  • 河南省高等学校重点科研项目(21B550008)
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2025年第16卷第11期
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doi: 10.19812/j.cnki.jfsq11-5956/ts.20250317002
  • 接收时间:2025-03-17
  • 首发时间:2025-07-14
  • 出版时间:2025-06-15
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  • 收稿日期:2025-03-17
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河南省高等学校重点科研项目(21B550008)
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    郑州科技学院食品科学与工程学院, 郑州市食品安全快速检测重点试验室, 郑州 450064

通讯作者:

* 郭楠楠(1986—), 女, 硕士, 副教授, 主要研究方向为食品安全快速检测分析新技术研究。E-mail:
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2种不同金属材料的力学参数

Family
属数
Number of
genus
种数
Number of
species
占总种数比例
Percentage of
total species (%)

Genus
种数
Number of
species
占总种数比例
Percentage of total
species (%)
鹅膏菌科Amanitaceae 2 11 5.26 鹅膏菌属 Amanita 10 4.78
小菇科 Mycenaceae 2 12 5.74 丝盖伞属 Inocybe 5 2.39
多孔菌科 Polyporaceae 8 14 6.70 蜡蘑属 Laccaria 5 2.39
红菇科 Russulaceae 3 23 11.00 小皮伞属 Marasmius 6 2.87
小菇属 Mycena 11 5.26
光柄菇属 Pluteus 5 2.39
红菇属 Russula 17 8.13
栓菌属 Trametes 5 2.39
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