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1 5 5 Vol. 5 No Journal of Food Safety and Quality May., 2014 万德慧 1, 王晓朋 1, 吴中波 1, 唐晗 1, 刘晓宇 (1., ; 2., ) 1, 2* 摘要 : 目的 (ractopamine, RCT) (molecularly imprinted polymers, MIPs),, 方法, ;,,, 结果, ng/ml, I=4.8777C , r=0.9907, 0.83 ng/ml, 10 ng/ml 11, RSD( ) 7.1% RCT, 90.3%~99.8% 结论, 关键词 : ; ; A microfluidic chip-chemiluminescence sensor based on molecular imprinted recognition for determination of ractopamine WAN De-Hui 1, WANG Xiao-Peng 1, WU Zhong-Bo 1, TANG Han 1 1, 2*, LIU Xiao-Yu (1. College of Food Science and Technology, Huazhong Agricultural University, Wuhan , China; 2. Key laboratory of Environment Correlative Dietology, Ministry of Education, Wuhan , China) ABSTRACT: Objective A ractopamine (RCT) imprinted polymer was prepared as a recognition element, which had the ability of specific recognition. Based on the recognition element, a novel microfluidic chip-chemiluminescence(cl) sensor was designed and assembled to monitor the RCT content in food. Methods The RCT imprinted polymer was synthesized by precipitation polymerization. SEM, FT-IR spectra and absorption experiment were applied as characterization methods. With MIPs working as the recognition element, microfluidic chip as flow reaction cell, a novel flow analysis CL sensor for the determination of RCT was developed and optimized. Results Under the optimal conditions, the relative CL intensity showed a linear relationship in the range of ng/ml RCT with detection limit of 0.83 ng/ml. The linear regression 基金项目 : ( ) (2010BBB022) (2011PY094) ( ) Fund: Supported by the National Natural Science Foundation of China( ), Project of Research and Development of Hubei Province of China(2010BBB022), Fundamental Research Funds for the Central Universities(2011PY094)and National Undergraduate Training Programs for Innovation and Entrepreneurship of Huazhong Agricultural University( ) * 通讯作者 :,,,, liuxiaoyu@mail.hzau.edu.cn *Corresponding author: LIU Xiao-Yu, Professor, College of Food Science and Technology, Huazhong Agricultural University, Wuhan , China. liuxiaoyu@mail.hzau.edu.cn

2 equation was ΔI=4.8777C , r= The relative standard deviation was 7.1% for 10 ng/ ml RCT (n=11). The method was successfully used to analyze the RCT content in pig liver and beef. The recovery was between 90.3% and 99.8%. Conclusion The method is rapid, accurate, and simple pre-treatment, which can be well used in the determination of ractopamine residues in food. KEY WORDS: ractopamine; molecularly imprinted polymers; microfluidic chip chemiluminescence sensor 1 引言 (ractopamine, RCT) β-, RCT,, RCT [1] RCT,,, [2] RCT RCT [3-7] [8-12] [13, 14],,,, (molecular imprinting technology, MIT), [15], [16],,,, -, RCT 2 材料与方法 2.1 试剂与仪器 ( 98%, Dr.Ehrenstorfer GmbH); (MAA, 98%, ); (EGDMA, 98%, Sigma-Aldrich ); (AIBN, 98%, ); ( Sigma ); ( ); UV ( ); NEXUS 470 FT-IR ( Thermo Nicolet ); ( ); DHG-90A ( ); ( Beckman Coulter ); FA1204B (0.1 mg, ); ( ); HL-2 ( ); ( ); SB-3200DT( ) 2.2 实验方法分子印迹柱的制备 (1) MIPs g(0.25 mmol), 25 ml - (1:1, v/v), MAA 85 μl(1 mmol)(, ), 7~10 min 1 h,,,, ml(5 mmol)egdma, 40 mg AIBN,, 3~4 min, 10 min, h,, ( : 9: 1), 24 h, 12 h,, (NIPs) (2) MIPs - (1:1, v/v) mg/ml,, mg/ml A, - A 9 20 mg MIPs, 10.0 ml

3 5, : mg/ml, 12 h, 0.45 μm,, NIPs, Q : Q=(C 0 C)V/m : Q /(mg/g); C 0 /(mg/ml); C /(mg/ml); V /ml; m /g (3) 1000 μl, 300,,, 1 mm, 50 mg MIP, 样品制备,, 5.0 g,, 10 ml, min, r/min 5 min,,, 10.0 ml 50,, 传感器的组装及莱克多巴胺的检测 P 2, c, MIPs ; G, d c, ;, P 2,, P 1, 4 mmol/l b 0.5 mmol/l a,, I 0 ; G, e, MIP RCT, -, Is; 1 - Fig. 1 The flow analysis system with MIPs-CL sensor : a: ; b: ; c: ; d: ; e: ; G: ; P1 P2: 1 2 ; MIP: ; FC: ; NH: ; PMT: ; W: ; PC: (1.0 mm i.d., 1.4 mm o.d.),, -400 V 3 结果与讨论 3.1 印记聚合物的形貌结构分析 2 MIPs NIPs, NIPs, ; MIPs, Fig. 2 2 MIPs Scanning electron micrographs of MIPs

4 红外光谱分析 MAA 3, 1640 cm -1 C=C, 947 cm cm -1 C-H ; C=O 1700 cm -1, MAA EGDMA MIPs, 3, 1730 cm -1 MIPs C=O 3 MIPs 3540 cm -1 MAA O-H, 1160 cm -1 EGDMA C-O, 1450 cm cm -1 -CH 2 -CH 3 MIPs MAA, 1640 cm cm cm -1, C=C C-H, MAA C=C MIPs NIPs, IR MIPs Q NIPs, MIPs NIPs 2.56 MIPs 4 MIPs NIPs Fig. 4 Adsorption isotherms of MIPs and NIPs 3.4 条件优化鲁米诺浓度的选择 mol/l, mol/l, ~ mol/l 5,,,, mol/l 3 MIPs NIPs MAA Fig. 3 IR spectra of MIPs NIPs and MAA 3.3 分子印迹聚合物的吸附特性分析, 276 nm, 276 nm A Y=0.0100X , R 2 = MIPs NIPs 4,,, MIPs NIPs Q, 5 (n=3) Fig. 5 Effect of luminol concentration on relative CL intensity (n=3)

5 5, : 铁氰化钾浓度的选择 K 3 Fe(CN) 6, ph, mol/l 6,,, mol/l,, mol/l, mol/l ( I), I=4.8777C , r=0.9907, 0.83 ng/ml, 10 ng/ml 11, RSD( ) 7.1% 3.6 干扰实验, 10 ng/ml, ±5%, 实际样品的测定, HPLC,, (n=3) Fig. 6 Effect of potassium ferricyanide concentration on relative CL intensity(n=3) 富集及清洗时间的选择,,,, min 7, 1~8 min,, 5 min, 5 min 1~5 min,, 8, 2.5 min, 2.5 min 7 (n=3) Fig. 7 The effect of preconcentration time on relative CL intensity (n=3) 3.5 标准曲线精密度和检出限,, 6~ 960 ng/ ml 8 (n=3) Fig. 8 The effect of cleaning time on relative CL intensity (n=3)

6 Table 1 表 1 共存物的最大允许量 Interference resistance of this method to coexisting substances * K + NH 4 + Cl Na + Ca 2+ Mg 2+ C 2 O 4 2- SO 4 2- CO EDTA Ni : *1000 Table 2 表 2 猪肝和牛肉样品中 RCT 含量 The results of RCT in pork liver and beef samples * RSD(%) HPLC RSD(%) (ng/g) (ng/g) : * Table 3 表 3 加标回收率结果 The results of sample recovery * (%) RSD(%) (ng/g) % % % 8.5 (ng/g) % 7.4 : * 4 总结,,,, 参考文献 [1]. [J]., 2010, (4): Zhang W. The Residue and Assay of Clenbuterol Hydrochloride in Meats [J]. Meat Res, 2010, (4): [2],,. [J]., 2010, 26(9): Wang XX, Liu XY, Peng YP. Reviews of Detection Methods and Devolvement of Ractopamine [J]. Mod Food Sci Technol, 2010, 26(9): [3],. [J]., 2005, 15(6): Gao YH, Wu XM. Rapid determination of ractopamine and clenbuterol with liquid chromatography mass spectrometry in feeds [J]. Chin J Health Lab Technol, 2005, 15(6): [4],,,. - [J]., 2007, (1): Zang YJ, Zhang CH, Zhang H, et al. Simultaneous determination of ractopamine and clenbuterol residues in meat products by liquid chromatography-electrospray-ionization/mass spectrometry [J].

7 5, : 1397 Meat Ind, 2007, (1): [5],,,. - [J]., 2006, 34(1): Ying YF, Pi YE, Wu PG, et al. The determination of ractopamine in animal tissue by solid phase extraction-high performance liquid chromatography [J]. Anal Chem, 2006, 34(1): [6] Du W, Zhao G, Fu Q, et al. Combined microextraction by packed sorbent and high-performance liquid chromatography ultraviolet detection for rapid analysis of ractopamine in porcine muscle and urine samples [J]. Food Chem, 2014, 145: [7] Freire EF, Borges KB, Tanimoto H, et al. Monitoring of ractopamine concentration in the mixture of this feed additive with vitamin mineral complex and with swine feed by HPLC [J]. Food Addit & Contam: Part A, 2013, 30(5): [8],,,. [J]., 2009, (2): Guo HM, Du HJ, Liu WS, et al. Developing gold immunochromatography assay for rapid detection of ractopamine [J]. Guangdong Agr Sci, 2009, (2): [9] Cheng F, Zhang K, Zhao SQ, et al. Study on spectral and immune identification of artificial antigen of ractopamine [J]. Sci Technol Food Ind, 2009, 30(11): [10],,,. [J]., 2006, 42(12): Wang JP, Shi WM, He FY, et al. The determination of ractopamine in pig urine by Enzyme-linked immunosorbent method [J]. Chin J Vet Med, 2006, 42(12): [11] Michael P. Recent applications of surface plasmon resonance biosensors for analyzing residues and contaminants in food [J]. Chem Mon, 2009, 395(4): [12],,,. [J]., 2012, 28(1): Liu DF, Deng SL, Lai WH, et al. Development of lateral flow assay of ractopamine based on fluorescent microspheres [J]. Food Mach, 2012, 28(1): [13],,,. [J]., 2010, 38(3): Li H, Cai HY, Chen X, et al. Continuous Detection Method for Ractopamine with Surface Plasmon Resonance Biosenso [J]. Chin J Anal Chem, 2010, 38(3): [14],,,. [J]., 2011, 69(23): Chen CY, Zhang HL, Liu MS, et al. An Electrochemical Immunosensor for the Determination of Ractopamine Based on Multi-walled Carbon Nanotubes and Ionic Liquid Composite [J]. Acta Chim Sin, 2011, 69(23): [15] Alexander C, Andersson HS, Andersson LI, et al. Molecular imprinting science and technology : a survey of the literature for the years up to and including 2003 [J]. J Mol Recognit: JMR, 2006, 19 (2) : [16] Brüggemann O, Visnjevski A, Patel P. Selective extraction of antioxidants with molecularly imprinted polymers [J]. Anal Chim Acta, 2004, 504(1): 作者简介 ( 责任编辑 : 赵静 ) 万德慧, 硕士, 主要研究方向为食品安全 @qq.com 刘晓宇, 博士, 教授, 博士生导师, 主要研究方向为食品安全与农业资源综合利用 liuxiaoyu@mail.hzau.edu.cn

Microsoft Word - 3098-3102 20150611007-蔡伟江-二校

Microsoft Word - 3098-3102 20150611007-蔡伟江-二校 6 8 Vol. 6 No. 8 2015 8 Journal of Food Safety and Quality Aug., 2015 蔡伟江 *, 邓鹏飞, 刘光兰, 李丹 (, 519040) 摘要 : 目的, 方法 (4.6 mm 250 mm, 5.0 μm) ; : : : =70: 16: 14(V:V:V),, : 1.0 ml/min; : 40 ; : 10