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Article(id=1216517520614412491, tenantId=1146029695717560320, journalId=1149652044408987649, issueId=1216517514570417012, articleNumber=null, orderNo=null, doi=10.19812/j.cnki.jfsq11-5956/ts.20250301002, pmid=null, cstr=null, oa=null, hot=null, price=null, onlineType=0, articleFormat=0, articleType=null, articleTypeStr=research-article, receivedDate=1740758400000, receivedDateStr=2025-03-01, revisedDate=null, revisedDateStr=null, acceptedDate=null, acceptedDateStr=null, onlineDate=1767969978717, onlineDateStr=2026-01-09, pubDate=1755187200000, pubDateStr=2025-08-15, doiRegisterDate=null, doiRegisterDateStr=null, onlineIssueDate=1767969978717, onlineIssueDateStr=2026-01-09, onlineJustAcceptDate=null, onlineJustAcceptDateStr=null, onlineFirstDate=null, onlineFirstDateStr=null, sourceXml=null, magXml=null, createTime=1767969978717, creator=13701087609, updateTime=1767969978717, updator=13701087609, issue=Issue{id=1216517514570417012, tenantId=1146029695717560320, journalId=1149652044408987649, year='2025', volume='16', issue='15', pageStart='1', pageEnd='322', issueExtLink='null', onlineDate='null', pubDate='null', beforeIssueId=null, nextIssueId=null, price=null, status=1, issueComplete=1, articleOrder=1, issueType=-1, specialIssue=null, createTime=1767969977276, creator=13701087609, updateTime=1768211590858, updator=13701087609, preIssue=null, nextIssue=null, ext={EN=IssueExt(id=1217530915467743720, tenantId=1146029695717560320, journalId=1149652044408987649, issueId=1216517514570417012, language=EN, specialIssueTitle=, coverIllustrator=null, specialIssueEditor=, specialIssueAbout=), CN=IssueExt(id=1217530915467743721, tenantId=1146029695717560320, journalId=1149652044408987649, issueId=1216517514570417012, language=CN, specialIssueTitle=, coverIllustrator=null, specialIssueEditor=, specialIssueAbout=)}, issueFiles=null}, startPage=110, endPage=117, ext={EN=ArticleExt(id=1216517521495216366, articleId=1216517520614412491, tenantId=1146029695717560320, journalId=1149652044408987649, language=EN, title=Detection of putrescine in Procambarus clarkii by carbon nanotubes-poly(neutral red) electrochemical competitive sensor, columnId=1216517518575980656, journalTitle=Journal of Food Safety & Quality, columnName=Special Topic: Application of Biosensors in Food Safety Detection, runingTitle=null, highlight=null, articleAbstract=

Objective To construct a carbon nanotube-poly(neutral red) modified electrode for the competitive electrochemical rapid detection of putrescine in Procambarus clarkii samples. Methods Putrescine was selected as a freshness indicator for Procambarus clarkii. A carbon nanotube-poly(neutral red) modified electrode was designed for putrescine detection. The relationship between the electrochemical behavior of the electrode and the concentration of putrescine was explored, and its application in electrochemical sensing of putrescine was investigated. After optimizing factors such as scan rate and electrolyte pH, the putrescine content in Procambarus clarkii was determined. Recovery experiments were conducted, and the results were compared with high performance liquid chromatography. Results The carbon nanotube-poly(neutral red) modified electrode exhibited a significant electrochemical inhibition effect towards putrescine, with a notable inverse relationship between the concentration of putrescine and the peak current of the electrode. The linear range was described by the equation ΔI=3.8ln[c(PUT)]+8.3, with a correlation coefficient of r=0.99. When applied to the detection of putrescine in crayfish samples, the results obtained by this method were highly consistent with those obtained by the high performance liquid chromatography. Conclusion The method can be employed for the detection of putrescine concentration in Procambarus clarkii and serves as a biosensor for putrescine, a freshness indicator of Procambarus clarkii.

, correspAuthors=Shuo DUAN, Ji-Wang CHEN, 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=Shuo DUAN, Ke ZHAN, Huang DAI, Yan LIU, Ji-Wang CHEN), CN=ArticleExt(id=1216517526733902221, articleId=1216517520614412491, tenantId=1146029695717560320, journalId=1149652044408987649, language=CN, title=碳纳米管-聚中性红电化学竞争性传感器检测小龙虾中腐胺, columnId=1216517518756335730, journalTitle=食品安全质量检测学报, columnName=专题：生物传感器在食品安全检测中的应用, runingTitle=null, highlight=null, articleAbstract=

目的 构建碳纳米管-聚中性红修饰电极用于小龙虾样品中腐胺的竞争性电化学快速检测。方法 以腐胺作为小龙虾鲜度特征物, 并针对腐胺设计了碳纳米管-聚中性红修饰电极, 探索电极电化学行为与腐胺浓度之间的关系, 并将其应用于腐胺的电化学传感研究。在优化了扫描速度、电解液pH等相关因素后, 测定小龙虾中的腐胺含量, 进行回收率实验, 并与高效液相色谱法进行对比。结果 水滑石-碳纳米管-聚中性红修饰电极对腐胺表现出明显的电化学抑制现象, 且腐胺的浓度与电极自身的峰电流对数值呈现出显著的反比关系, 其线性关系方程式为ΔI=3.8ln[c(PUT)]+8.3, 相关系数r=0.99。将其应用于小龙虾实际样品的腐胺检测, 该方法得到的腐胺含量与高效液相色谱法得到的结果高度一致。结论 该方法适合用于小龙虾中腐胺浓度的检测, 可以作为小龙虾鲜度特征物腐胺的生物传感器。

, correspAuthors=段烁, 陈季旺, authorNote=null, correspAuthorsNote=
*陈季旺(1970—), 男, 博士, 教授, 主要研究方向为肉制品加工与质量控制。E-mail: ;
段烁(1988—), 男, 博士后, 副教授, 主要研究方向为电化学在食品安全中的应用。E-mail:
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注: A. 电沉积NR的循环伏安图; B. 3种电极的拉曼光谱对比图; C. CNT-GCE的扫描电镜图; D. PNR-CNT-GCE的扫描电镜图。

, figureFileSmall=RkDHYHlOUXCqM4X+3pVuxg==, figureFileBig=eVAmVnN2X43PDv9qD+eSgQ==, tableContent=null), ArticleFig(id=1217127901053501632, tenantId=1146029695717560320, journalId=1149652044408987649, articleId=1216517520614412491, language=EN, label=Fig.3, caption=Comparison of electrochemical performances for different electrodes, figureFileSmall=9eeJbuEZaW/NWRI9C/3gEw==, figureFileBig=PK3XaU9ekzNCDVsinL3fXQ==, tableContent=null), ArticleFig(id=1217127902320181444, tenantId=1146029695717560320, journalId=1149652044408987649, articleId=1216517520614412491, language=CN, label=图3, caption=不同电极的电化学性能比较

注: A. 4种电极在铁氰化钾电解液中的循环伏安图; B. 4种电极在腐胺电解液中的计时电量图。

, figureFileSmall=9eeJbuEZaW/NWRI9C/3gEw==, figureFileBig=PK3XaU9ekzNCDVsinL3fXQ==, tableContent=null), ArticleFig(id=1217127902454399180, tenantId=1146029695717560320, journalId=1149652044408987649, articleId=1216517520614412491, language=EN, label=Fig.4, caption=Study on the electrochemical behavior of putsamine on electrodes, figureFileSmall=Pu+uCh8dxZsKvA30+Wy1/w==, figureFileBig=238bsKPpnAWq+3QRmYRP9g==, tableContent=null), ArticleFig(id=1217127902592811219, tenantId=1146029695717560320, journalId=1149652044408987649, articleId=1216517520614412491, language=CN, label=图4, caption=腐胺在电极上的电化学行为研究

注: A. 不同腐胺浓度下的循环伏安图; B. 不同pH下腐胺浓度与电流变化值的线性关系图。

, figureFileSmall=Pu+uCh8dxZsKvA30+Wy1/w==, figureFileBig=238bsKPpnAWq+3QRmYRP9g==, tableContent=null), ArticleFig(id=1217127902748000478, tenantId=1146029695717560320, journalId=1149652044408987649, articleId=1216517520614412491, language=EN, label=Fig.5, caption=Electrochemical detection of putsamine, figureFileSmall=YKHPUYGF60fCeA0Fhzf6yg==, figureFileBig=nm5ex6LShebZOoijoyguoQ==, tableContent=null), ArticleFig(id=1217127902919966951, tenantId=1146029695717560320, journalId=1149652044408987649, articleId=1216517520614412491, language=CN, label=图5, caption=腐胺的电化学检测

注: A. 不同腐胺浓度下的差分脉冲伏安图; B. 腐胺浓度与电流变化值之间的线性关系图。

, figureFileSmall=YKHPUYGF60fCeA0Fhzf6yg==, figureFileBig=nm5ex6LShebZOoijoyguoQ==, tableContent=null), ArticleFig(id=1217127903200985330, tenantId=1146029695717560320, journalId=1149652044408987649, articleId=1216517520614412491, language=EN, label=Table 1, caption=

Comparison bewteen electrochemical detection values and HPLC detection values for putsamine concentration in Procambarus clarkii samples

, figureFileSmall=null, figureFileBig=null, tableContent=
小龙虾
样品		 腐胺含量/(mg/kg) 差值
/(mg/kg)		 相对标准
偏差/%
电化学检测值 HPLC测定值
T0-1 3.38 3.41 0.03 0.81
T0-2 3.62 3.68 0.06 1.69
T0-3 3.77 3.89 0.12 3.29
T0-4 3.96 3.95 0.01 0.26
T0-5 3.45 3.47 0.02 0.58
T3-1 20.55 20.71 0.26 1.27
T3-2 19.33 19.60 0.27 1.40
T3-3 18.93 18.80 0.13 0.38
T3-4 20.22 20.30 0.08 1.05
T3-5 21.12 21.25 0.13 0.48
T7-1 56.11 56.53 0.42 0.74
T7-2 50.33 49.22 1.11 2.26
T7-3 48.43 49.84 1.41 2.84
T7-4 58.53 59.80 1.27 2.11
T7-5 62.91 64.04 1.13 1.78
), ArticleFig(id=1217127903372951805, tenantId=1146029695717560320, journalId=1149652044408987649, articleId=1216517520614412491, language=CN, label=表1, caption=

小龙虾样品中腐胺含量的电化学检测值与HPLC检测值的比较

, figureFileSmall=null, figureFileBig=null, tableContent=
小龙虾
样品		 腐胺含量/(mg/kg) 差值
/(mg/kg)		 相对标准
偏差/%
电化学检测值 HPLC测定值
T0-1 3.38 3.41 0.03 0.81
T0-2 3.62 3.68 0.06 1.69
T0-3 3.77 3.89 0.12 3.29
T0-4 3.96 3.95 0.01 0.26
T0-5 3.45 3.47 0.02 0.58
T3-1 20.55 20.71 0.26 1.27
T3-2 19.33 19.60 0.27 1.40
T3-3 18.93 18.80 0.13 0.38
T3-4 20.22 20.30 0.08 1.05
T3-5 21.12 21.25 0.13 0.48
T7-1 56.11 56.53 0.42 0.74
T7-2 50.33 49.22 1.11 2.26
T7-3 48.43 49.84 1.41 2.84
T7-4 58.53 59.80 1.27 2.11
T7-5 62.91 64.04 1.13 1.78
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碳纳米管-聚中性红电化学竞争性传感器检测小龙虾中腐胺
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段烁 1, 2, * , 占可 2 , 戴煌 2 , 刘言 2 , 陈季旺 2, *
食品安全质量检测学报 | 专题：生物传感器在食品安全检测中的应用 2025,16(15): 110-117
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食品安全质量检测学报 | 专题：生物传感器在食品安全检测中的应用 2025, 16(15): 110-117
碳纳米管-聚中性红电化学竞争性传感器检测小龙虾中腐胺
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段烁1, 2, * , 占可2, 戴煌2, 刘言2, 陈季旺2, *
作者信息
  • 1 重庆化工职业学院环境与质量检测系, 重庆 401228
  • 2 武汉轻工大学食品科学与工程学院, 武汉 430023

通讯作者:

*陈季旺(1970—), 男, 博士, 教授, 主要研究方向为肉制品加工与质量控制。E-mail: ;
段烁(1988—), 男, 博士后, 副教授, 主要研究方向为电化学在食品安全中的应用。E-mail:
Detection of putrescine in Procambarus clarkii by carbon nanotubes-poly(neutral red) electrochemical competitive sensor
Shuo DUAN1, 2, * , Ke ZHAN2, Huang DAI2, Yan LIU2, Ji-Wang CHEN2, *
Affiliations
  • 1 Environment and Quality Test Department, Chongqing Chemical Industry Vocational College, Chongqing 401228, China
  • 2 College of Food Science and Engineering, Wuhan Polytechnic University, Wuhan 430023, China
出版时间: 2025-08-15 doi: 10.19812/j.cnki.jfsq11-5956/ts.20250301002
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摘要
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目的 构建碳纳米管-聚中性红修饰电极用于小龙虾样品中腐胺的竞争性电化学快速检测。方法 以腐胺作为小龙虾鲜度特征物, 并针对腐胺设计了碳纳米管-聚中性红修饰电极, 探索电极电化学行为与腐胺浓度之间的关系, 并将其应用于腐胺的电化学传感研究。在优化了扫描速度、电解液pH等相关因素后, 测定小龙虾中的腐胺含量, 进行回收率实验, 并与高效液相色谱法进行对比。结果 水滑石-碳纳米管-聚中性红修饰电极对腐胺表现出明显的电化学抑制现象, 且腐胺的浓度与电极自身的峰电流对数值呈现出显著的反比关系, 其线性关系方程式为ΔI=3.8ln[c(PUT)]+8.3, 相关系数r=0.99。将其应用于小龙虾实际样品的腐胺检测, 该方法得到的腐胺含量与高效液相色谱法得到的结果高度一致。结论 该方法适合用于小龙虾中腐胺浓度的检测, 可以作为小龙虾鲜度特征物腐胺的生物传感器。

小龙虾  /  腐胺  /  电化学传感器  /  鲜度  /  纳米材料

Objective To construct a carbon nanotube-poly(neutral red) modified electrode for the competitive electrochemical rapid detection of putrescine in Procambarus clarkii samples. Methods Putrescine was selected as a freshness indicator for Procambarus clarkii. A carbon nanotube-poly(neutral red) modified electrode was designed for putrescine detection. The relationship between the electrochemical behavior of the electrode and the concentration of putrescine was explored, and its application in electrochemical sensing of putrescine was investigated. After optimizing factors such as scan rate and electrolyte pH, the putrescine content in Procambarus clarkii was determined. Recovery experiments were conducted, and the results were compared with high performance liquid chromatography. Results The carbon nanotube-poly(neutral red) modified electrode exhibited a significant electrochemical inhibition effect towards putrescine, with a notable inverse relationship between the concentration of putrescine and the peak current of the electrode. The linear range was described by the equation ΔI=3.8ln[c(PUT)]+8.3, with a correlation coefficient of r=0.99. When applied to the detection of putrescine in crayfish samples, the results obtained by this method were highly consistent with those obtained by the high performance liquid chromatography. Conclusion The method can be employed for the detection of putrescine concentration in Procambarus clarkii and serves as a biosensor for putrescine, a freshness indicator of Procambarus clarkii.

Procambarus clarkii  /  putrescine  /  electrochemical sensor  /  freshness  /  nanomaterial
段烁, 占可, 戴煌, 刘言, 陈季旺. 碳纳米管-聚中性红电化学竞争性传感器检测小龙虾中腐胺. 食品安全质量检测学报, 2025 , 16 (15) : 110 -117 . DOI: 10.19812/j.cnki.jfsq11-5956/ts.20250301002
Shuo DUAN, Ke ZHAN, Huang DAI, Yan LIU, Ji-Wang CHEN. Detection of putrescine in Procambarus clarkii by carbon nanotubes-poly(neutral red) electrochemical competitive sensor[J]. Journal of Food Safety & Quality, 2025 , 16 (15) : 110 -117 . DOI: 10.19812/j.cnki.jfsq11-5956/ts.20250301002
正文
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0 引言
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小龙虾, 学名克氏原螯虾, 由于肉质鲜嫩、蛋白含量高, 深受消费者喜爱, 最近十年在我国广泛养殖。中国小龙虾产业发展报告(2024)显示, 我国小龙虾养殖产量占全国淡水养殖总产量的9.26%, 位列我国淡水养殖品种第4位, 淡水虾类养殖第1位, 是我国农业和食品产业的重要组成部分[1]。由于小龙虾生长周期存在季节性, 通常仅在夏季繁殖生长, 导致夏季以外的季节产量较低甚至停滞。因此, 小龙虾通常需要冷冻贮藏以保证其他季节的小龙虾消费和生产供应。在贮藏过程中, 小龙虾新鲜度会逐渐下降, 营养价值降低, 还会产生细菌等微生物、生物胺、嘌呤等有害物质, 危害消费者的身体健康[2-3]。因此, 为了保障小龙虾相关食品的食品安全和营养价值, 对小龙虾产品的新鲜度检测显得极为重要。
目前, 新鲜度检测主要是通过检测某项或多项与新鲜度密切相关的指标(即鲜度特征物)来实现间接检测[4]。例如, 电导率[5]、细菌[6]、黄嘌呤和次黄嘌呤[7-8]、三甲胺[9]、生物胺[10]等。在包括小龙虾在内的各种水产品新鲜度检测研究中, 由于其鲜度特征物通常是源于水产品自身滋生的微生物或代谢出的生物分子(如嘌呤[11]、尿酸、三甲胺、组胺或腐胺等), 因此, 针对生物类鲜度特征物的生物传感器成为了鲜度检测的热点[12-13]。
生物传感器由于可以利用生物识别元件(如酶、抗体、核酸等)对目标物进行特异性识别, 因而其具有高特异性等优点, 在环境监测、食品安全检测、临床诊断等领域有着广泛的应用[14-15]。近年来, 基于生物传感器的食品新鲜度检测也逐渐进入研究人员的视野。例如, 1992年加拿大就开发了生物传感设备用于牛羊肉等食品的鲜度检测[16]。彭文杰[17]综述了生物传感器在食品安全监测中的应用, 其中针对食品鲜度检测的生物传感器研究占很大比重。然而, 食品等生物样品中存在复杂的蛋白质、脂类、无机盐和各种激素等, 这些物质均可能对生物传感器的检测产生干扰[18-19]。许多分析化学研究人员开发了各种手段以提升生物传感器在复杂体系中的抗干扰能力[20]。其中, 构建自身具有响应信号的竞争型生物传感器, 是解决复杂基质干扰的有效途径[21]。竞争型传感器通过检测传感器自身信号, 以目标分子带来的信号变化来实现目标物浓度检测[22]。由于生物样品复杂基质等外部因素带来的扰动会同时影响目标分子和传感器本身, 因此以目标分子带来的信号变化作为定量策略, 可以有效提高生物传感器在复杂条件下的准确度和可靠性[23]。目前, 竞争型生物传感器(如竞争性酶联免疫分析)在医学和生物检测中已经十分成熟[24], 其在复杂基质干扰下的选择性和稳定性已被广泛认可。但在食品检测, 尤其是食品鲜度检测中, 竞争性(电化学)生物传感器的研究和应用依然相对较少。
基于王汝娟等[25]和田娟等[26]关于小龙虾生物胺组分的研究, 以及本课题组围绕小龙虾限度特征物检测的前期数据[27], 本研究筛选了腐胺作为小龙虾鲜度特征物进行电化学研究, 在电极上负载碳纳米管(carbon nanotubes, CNT)并沉积了聚中性红(poly neutral red, PNR), 以CNT为导电介质, 以PNR为信号响应分子, 以腐胺在电极上的吸附导致的电化学信号变化实现腐胺的分析传感。本研究为小龙虾腐胺快速检测技术提供了新的思路, 为小龙虾鲜度快速评价提供了新的技术支撑。
1 材料与方法
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1.1 材料与试剂
原料: 鲜活小龙虾购买于当地市场。
腐胺标准品、精胺标准品、亚精胺标准品(纯度>98%)、CNT(纯度>95%)(美国Sigma-Aldrich化学试剂有限公司); 乙腈、丙酮、正己烷、乙醚、正丁醇、三氯甲烷、乙酸、乙酸铵、谷氨酸钠、碳酸氢钠、三氯乙酸、硝酸钠、异丙醇、中性红(neutral red, NR)、铁氰化钾(化学纯, 国药集团化学试剂有限公司); Nafion溶液(纯度5%, 北京伊诺凯化学试剂网); 去离子水(实验室自制)。
1.2 仪器与设备
CHI760E型电化学工作站(美国辰华仪器有限公司上海分公司); ACQUITY H-Class型高效液相色谱仪、C18型双向色谱柱(100 mm×2.5 mm, 5 μm)(美国Waters科技有限公司); TSX40086LA型超低温冰箱(美国赛默飞科技有限公司上海分公司); SB-5200DTN超声波清洗机、XHF-DY型高速分散器(宁波新芝生物科技股份有限公司); FC5718型高速冷冻离心机[奥豪斯仪器(常州)有限公司]; ZETASIZER NANO S90纳米粒度电位仪(英国马尔文帕纳科公司); INVIA-QONTOR显微拉曼光谱仪(英国Renishaw公司); S400型pH计、LA204/A型分析天平(精密度0.1 mg) (瑞士梅特勒-托利多仪器有限公司); TM 4000 PLUSⅡ扫描电子显微镜(日本Hitachi有限公司)。
1.3 实验方法
1.3.1 小龙虾样品处理及生物胺含量测定
小龙虾样品处理为避免样品间差异过大, 小龙虾样品由同一家超市同批次购买, 筛选质量接近(单只约30 g±5 g)的鲜活小龙虾共2 kg。参考小龙虾工业化加工中的实际贮藏条件, 将鲜活小龙虾超声清洗后置于-50 ℃环境中冰冻2 h致死, 然后-18 ℃下冻藏, 得到小龙虾模拟冷冻样品。测试前1 d将冷冻小龙虾样品置于4 ℃冰箱解冻。小龙虾样品解冻后, 进行去壳、匀浆处理, 参考GB 5009.208—2016《食品安全国家标准 食品中生物胺的测定》以高效液相色谱法(high performance liquid chromatography, HPLC)测定腐胺、组胺、亚精胺等生物胺类含量。经过测定, 小龙虾中腐胺含量与水产品鲜度指标总挥发性盐基氮显著相关(相关系数为0.94), 因此本研究选取腐胺作为小龙虾鲜度特征物进行检测研究。
1.3.2 电化学传感器制备
用去离子水超声洗涤玻碳电极(glassy carbon electrode, GCE)至表面光洁无污染后晾干。称取2 mg CNT置于5 mL离心管中, 加入2 mL异丙醇、10 μL Nafion溶液, 超声直至液体混合完全分散。准确移取超声处理后的分散液10 μL分5次滴加至GCE电极表面, 待分散液彻底干燥即可得到CNT修饰的GCE (CNT-GCE)。
PNR以电化学聚合法负载在电极上, 方法参考文献[20]。准确称取2.3 mg NR置于电解池中, 加入20 mL含0.1 mol/L NaNO3的磷酸缓冲盐溶液(0.1 mol/L, pH 6)使其完全溶解, 将制备好的CNT-GCE置于电解池中并作为工作电极。以50 mV/s的扫描速率在-0.8 V至0.8 V电位范围内反复进行循环伏安扫描。扫描完成后, NR即可在电极上形成PNR, 得到PNR-CNT修饰电极(PNR-CNT-GCE)。该电极即为用于腐胺检测的电化学传感器。同时准备了GCE、CNTs-GCE、PNR-GCE用于对照。
1.3.3 电化学测试方法及原理
本研究所用到的电化学测试方法包括循环伏安测试(cyclic voltammetry, CV)、计时电量测试(chroncoulometry, CC)和差分脉冲伏安测试(differential pulse voltammetry, DPV)。以上3种测试均采用三电极体系, 以Ag/AgCl电极为参比电极, 以铂丝电极为对电极, 以修饰前或修饰后的GCE为工作电极进行测试。其中循环伏安测试和计时电量测试所采用的电解液为含有1 mmol/L的铁氰化钾的磷酸缓冲溶液(phosphate buffer solution, PBS)作为电解液, 用于对比电极在修饰前后的性能差异; 而差分脉冲伏安测试则以含有腐胺标准品或小龙虾提取液样品的PBS电解液, 用于实现小龙虾中腐胺的电化学检测。当样品中没有腐胺存在时, 电极上的PNR会因为吸附电解液中带负电的磷酸根而产生较强的电化学还原峰; 当加入不新鲜的小龙虾样品时, 腐胺分子会与磷酸根结合, 导致原有的电化学还原峰出现变化, 根据还原峰变化程度, 即可判断小龙虾中腐胺含量, 进而判断小龙虾新鲜与否。实验原理如图1所示。
1.3.4 电极表征
电极材料的水合粒径测试、电极的拉曼光谱和扫描电子显微镜测试等均委托武汉轻工大学分析测试中心进行。
1.4 数据处理
本研究的电化学测试图、拉曼光谱测试图以及线性方程拟合图由OriginPro 2021版完成, 线性方程拟合模式为FitLine模式, Pearson相关系数要求大于0.99。本研究的分析方法检出限(limit of detection, LOD)计算公式为LOD=3σ/S, 其中σ是多次空白测量的标准差(n>7), S是方法的灵敏度(线性方程斜率)。分析检测数据取小数点后2位。
2 结果与分析
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2.1 电极的制备与表征
PNR在电极上的负载通过循环伏安法实现电化学聚合(见图2)。在聚合过程中, 聚合曲线在0.05到0.20 V范围内, 出现一对弱的氧化还原峰, 这归结于NR自身的氧化还原过程; 而在-0.60到-0.40 V范围内, 出现一对强的氧化还原峰, 这归结于PNR与电解液中的磷酸盐吸附结合而形成的中间体。该中间体电化学信号较强, 而且还能随着电聚合次数的增多而增强, 聚合10循环后从30 μA逐渐增加到65 μA左右, 可以作为竞争性生物传感器的主要检测信号。本研究将电聚合得到的PNR-CNT-GCE与未聚合的CNT-GCE和空白GCE进行一系列表征用于对比其性质。拉曼表征结果显示, 3种电极均出现碳的D峰(约1350 cm-1)和G峰(约1580 cm-1), 这归结于GCE本身的碳基底[28]。而加入CNT后, PNR-CNT-GCE和CNT-GCE的G峰有了显著的增高, DG峰强度比下降, 这归结于CNT引入了碳缺陷[29]。此外, 局部放大图显示, PNR-CNT-GCE在800、600和460 cm-1处出现3个峰, 分别归因于NR结构中C-C-N、C-N的伸缩振动, 以及C-C-C的面内弯曲振动[30]。这证明了PNR成功聚合在电极上。图C和图D分别为CNT-GCE和PNR-CNT-GCE的扫面电子显微镜图。在微观条件下, CNT呈现交错相连的管线结构, 这有利于与检测目标物接触并改善电极导电性。在PNR聚合后, CNT的表面变得光滑且厚度增加, 这意味着NR已经聚合在了CNT表面。将电极表面的CNT和PNR-CNT颗粒回收后进行了水合粒径测试和表面电荷测试。CNT的水合粒径为1345 nm, Zeta电位为-16.3 mV, 而聚合后的PNR-CNT水合粒径为4592 nm, Zeta电位为-5.5 mV。水合粒径的增大表明聚合后CNT颗粒变大, 这与扫面电镜图结果一致。Zeta电位正向移动归结于带正电的PNR聚合后中和了CNTs表面带负电的羧基, 使得整体电位向正向移动[31]。
2.2 传感器的电化学表征及分析传感研究
在含有铁氰化钾的PBS电解液中进行CV测试, 用于对比电极的电化学性质, 结果如图3A所示。铁氰化钾在4种电极GCE、CNTs-GCE、PNR-GCE与PNR-CNTs-GCE上均表现出良好的电化学响应, 在0.0 V到0.3 V范围内均出现一对氧化还原峰。其中PNR-CNTs-GCE表现出最高的氧化峰电流和还原峰电流, 意味着PNR-CNTs具有良好的电化学传感性能, 可用于构建生物传感器。而PNR-CNT-GCE和CNT-GCE在-0.7 V至-0.4 V范围内还存在一对氧化还原峰, 这是PNR与磷酸根离子结合的导电中间体, 该信号可以用于作为生物传感器的自身检测信号。将4种电极应用于含有腐胺的PBS电解液中进行计时电量测试, 结果见图3B所示。4种电极的CC曲线在Y轴上的截距与参与反应的目标物的量成正比, PNR-CNT-GCE展现出远超另外3种电极的电量值, 表明该电极对腐胺的结合能力最强, 有更多的腐胺参与了反应, 这对于构建用于腐胺检测的电化学传感器极为有利。
以上研究证明了PNR-CNT-GCE在腐胺检测中的潜力。以PNR-CNT-GCE为工作电极, 构筑电化学传感器, 将其应用于腐胺的电化学研究。在含有不同浓度的腐胺标准品的PBS电解液(pH=5)中, 以PNR-CNT-GCE为工作电极进行循环伏安测试, 结果如图4A所示。所有的CV曲线在-0.8 V左右均出现一个还原峰, 且还原峰的峰强度随着腐胺浓度的增加而逐渐减小(还原峰电流为负值, 所以还原峰强减小意味着峰值增大)。这是由于腐胺的吸附影响了PNR与磷酸根的结合, 降低了中间体的氧化还原行为。基于这一现象, 将电流的变化值(ΔI, μA)与电解液中腐胺的质量浓度[c(PUT), mg/L]进行作图, 得出其线性关系方程为ΔI=2.5ln[c(PUT)]+8.7, 相关系数r=0.99。进一步研究了在不同pH的电解液中, PNR-CNT-GCE对腐胺的电化学响应, 结果见图4B。当pH为5时, 其线性方程斜率最高, 表明所制备的电化学传感器在pH为5时对腐胺的响应最为灵敏, 即最佳电解液为pH为5的PBS溶液。同时本研究也探讨了CNT用量、电聚合时间等因素, 获得了传感器的最佳测试条件。
为进一步提升传感器的灵敏度, 采用差分脉冲伏安法(differential pulse voltammetry, DPV)对腐胺含量进行检测。在最优条件下, 在含有不同浓度腐胺的电解液中进行DPV测试, 结果如图5所示。腐胺质量浓度在0至50 mg/L范围内, 随着腐胺浓度的升高, 还原峰强度逐渐降低(因还原峰电流数值为负, 电流值逐渐升高)。以电流的变化值(ΔI, μA)与电解液中腐胺的质量浓度[c(PUT), mg/L]进行作图, 得到其线性方程为ΔI=3.8ln[c(PUT)]+8.3, 相关系数r=0.99。
2.3 传感器在小龙虾样品腐胺检测中的应用
根据线性方程和实际实验结果, 计算出本方法构建的腐胺传感器的最低检出限(limit of detection, LOD)为0.08 mg/kg, 与现有的文献[32]报道值相比, 本方法的LOD具有一定优势。新鲜小龙虾样品中腐胺含量超过3 mg/kg, 腐败小龙虾样品中腐胺浓度通常超过50 mg/kg[33], 远高于本方法的LOD (0.08 mg/kg), 因此, 本方法具有实际应用价值。
为保证传感器的稳定性和重复性等指标, 按照1.3.2中的方法制作了7根相同的PNR-CNT-GCE分别作为工作电极, 对同一小龙虾样品中的腐胺进行电化学检测, 其相对标准偏差为4.7%, 小于国家规定的5%; 采用同一根工作电极, 对含有相同浓度的腐胺电解液重复测定7次, 相对标准偏差为1.8% (n=7)。这表明本方法建立的腐胺传感器, 具有良好的稳定性和重复性。在测试用的电解液中添加小龙虾样品中常见的干扰物质(组胺、精胺、亚精胺、谷氨酸、精氨酸等), 峰电流最大偏差(组胺)为4.2%, 最小偏差(谷氨酸)为0.3%, 均小于国家规定的5%, 这表明采用的竞争性传感器对小龙虾基质中常见干扰物有着显著的稳定性和抗干扰能力。
将本方法构建的传感器应用于小龙虾实际样品的检测中, 并与GB 5009.208—2016规定的HPLC对比。对储藏0、3和7 d的小龙虾样品(分别命名为T0、T3和T7), 本方法和HPLC方法测得的结果见表1所示。与GB 5009.208—2016的HPLC相比, 本方法测得的结果相对标准偏差均小于5%, 符合要求。
3 结论
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本研究以CNT为导电介质, 电聚合制备了PNR-CNT结构, 利用PNR与磷酸根离子结合能力产生电化学信号, 并以腐胺对PNR-磷酸的阻碍作用实现了竞争性电化学传感, 并基于此建立了竞争性电化学传感器。结果表明, 该方法具备良好的检出限、稳定性和重复性, 与GB 5009.208—2016测得结果极为接近。综上所述, 本研究构建的竞争性电化学传感器, 在小龙虾中腐胺含量的检测上具有显著优势。
基金
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  • 重庆市自然科学基金面上项目(2024NSCQ-MSX0774)
  • 重庆市教委科研项目青年项目(KJQN202404503)
  • 农村农业部农产品产地处理装备重点实验室开放课题项目(2016 NYZD2202)
文献
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2025年第16卷第15期
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文章信息
doi: 10.19812/j.cnki.jfsq11-5956/ts.20250301002
  • 接收时间：2025-03-01
  • 首发时间：2026-01-09
  • 出版时间：2025-08-15
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出版历史
  • 收稿日期：2025-03-01
基金
重庆市自然科学基金面上项目(2024NSCQ-MSX0774)
重庆市教委科研项目青年项目(KJQN202404503)
农村农业部农产品产地处理装备重点实验室开放课题项目(2016 NYZD2202)
作者信息
    1 重庆化工职业学院环境与质量检测系, 重庆 401228
    2 武汉轻工大学食品科学与工程学院, 武汉 430023

通讯作者:

*陈季旺(1970—), 男, 博士, 教授, 主要研究方向为肉制品加工与质量控制。E-mail: ;
段烁(1988—), 男, 博士后, 副教授, 主要研究方向为电化学在食品安全中的应用。E-mail:
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2种不同金属材料的力学参数
科
Family		 属数
Number of
genus		 种数
Number of
species		 占总种数比例
Percentage of
total species (%)		 属
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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