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1 5 12 Vol. 5 No Journal of Food Safety and Quality Dec., 2014 王培锋 *, 王凤美, 张鸿伟, 徐彪, 邱芳, 何京澄, 周维政 (, ) 摘要 : 目的 Q-Exactive 方法, QuEChERS,, ACQUITY BEH C 18, 0.1% -, Q-Exactive, 结果 , (limit of detection, LOD) 10 μg/kg 结论, 关键词 : ; ; Q-Exative Screening of antibiotics in foodstuffs of animal origin by high performance liquid chromatography-quadrurpole/electrostatic field orbitrap high resolution mass spectrometry WANG Pei-Feng *, WANG Feng-Mei, ZHANG Hong-Wei, XU Biao, QIU Fang, HE Jing-Cheng, ZHOU Wei-Zheng (Technical Center of Shandong Entry-Exit Inspection and Quarantine Bureau, Qingdao , China) ABSTRACT: Objective To develop a method of high performance liquid chromatography (HPLC) coupled with linear ion trap/orbitrap high resolution mass spectrometry (QuEChERS) to detect antibiotics quickly in food of animal origin. Methods Antibiotics in samples were extracted using a solvent of acidic acetonitrile, following by purification with QuEChERS, dilution with acetonitrile and H 2 O. The separation was performed on an ACQUITY BEH C 18 column using gradient elution with aqueous solution (containing 0.1% formic acid)/acetonitrile as a mobile phase. The accurate mass of quasi-molecular ion were acquired by full scanning of electrostatic field orbitrap and applied for rapid screening and analysis, further confirmation was accomplished according to the retention time and qualitative fragments obtained by data dependent scan mode. Results Under the optimal conditions, 31 antibiotics were routinely detected with mass accuracy error below , and good linearity were provided in their respective linear ranges with correlation coefficients all higher than 0.99, the limit of detection (LOD) for all the 31 antibiotics were 10 μg/kg. Conclusion The method was applied to fish, milk, and honey and satisfying screening results proved that it is an efficient path 基金项目 : (2013IK176)( jh) ( ) Fund: Supported by the Scientific and Technological Project of the General Administration of Quality Supervision, Inspection and Quarantine of the People s Republic of China (2013IK176), Science and Technology Support Plan of Qingdao ( jh) and Scientific Research Projects by Quality Inspection Service Industry ( ) * 通讯作者 :,, @qq.com *Corresponding author: WANG Pei-Feng, Engineer, Technical Center of Shandong Entry-Exit Inspection & Quarantine Bureau, No.70, Qutangxia Road, Shinan District, Qingdao , Shandong Province, China @gmail.com

2 for rapid analysis of antibiotics in food of animal origin. KEY WORDS: antibiotics, food of animal origin, Q-exactive high resolution mass spectrometry 1 引言, (fluoroquinolones, FQs) 3, 6, 7, FQs, FQs,, FQs,,, (sulfonamides, SAs),,,,,, FQs SAs [1-3] [4-7] - [8-10], -,,,,, /,,,,, / [11] [12] [13-15] /, 2 材料和方法 2.1 仪器与试剂 Thermo Q-Exactvie ( Thermo Fisher ); Himac CR22GII ( Hitachi ); MS1Minishaker ( IKA ); FA1104 ( ); Milli-Q (18.2 MΩ cm) ( Millipore ) (, Fluka ); (, Merck ); (, Mallinckrodt Bake ) , Dr. Ehrenstorfer, 99% 2.2 标准品溶液的配制, 100 μg/ml, 18, 2.3 实验部分 2 g( 0.01 g) 50 ml, 4 ml, 1 min, 10 ml1%, 1 min, Quchers , 10 min, r/min 5 min, 30 min,

3 12, : ml Quchers , 1 min, r/min 5 min, 4 ml, 40, :0.1% (1:9), 1 min, r/min 10 min,, 2.4 色谱 - 质谱条件色谱条件 : Waters ACQUITY UPLC BEH C 18 (2.1 mm 100 mm, 1.7 μm, Waters ); : 300 μl/min; : 10 μl; : 30 ; A 0.1%, B ; 质谱条件 : Q-Exactive; : (electron spray ionization, ESI); : 350 ; : 3.0 kv; : 30 L/min; : 3.3 L/min; ; (m/z): 100~1000 R= Q-Exactive MS 1 Table Analysis parameters of the 31 antibiotics No. Compounds Retention time(min) Ion formation Theoretical (m/z) Measured (m/z) Mass accuracy error/10 6 (ppm) [M+H] [M+H] [M+H] [M+H] [M+H] [M+H] [M+H] [M+H] [M+H] [M+H] [M+H] [M+H] [M+H] [M+H] [M+H] [M+H] [M+H] [M+H] [M+H] [M+H] [M+H] [M+H] [M+H] [M+H] [M+H] [M+H] [M+H] [M+H] [M+H] [M+H] [M+H]

4 结果与讨论 3.1 样品的提取,,, ( 0.1% 0.1%), 0.1%,, 3.2 色谱 / 质谱条件的优化 Waters ACQUITY UPLC BEH C 18, 31 ACQUITY UPLC BEH C μm,,, ( -0.1% -0.1% ), 0.1%,, -0.1%, 0.1% - 31,,,, ESI + ESI - 31,, 31 ESI,, [M+H] +, 31,, ~3 3.3 回收率 3, 1 (limit of quantity, LOQ) 2 (2LOQ) 5 (5LOQ) 3, 6, (relative standard deviation, RSD), %~110.90%,RSD 1.56%~9.83%( 2) Fig Extracted ion chromatograms of the 9 kinds of representative antibiotics in fish muscle

5 第 12 期王培锋, 等: 高效液相色谱-四极杆轨道阱高分辨质谱对动物源食品中抗生素类成分的快速筛查 Fig. 2 Fig. 3 图 2 蜂蜜添加回收中 9 种代表抗生素提取离子色谱图 Extracted ion chromatograms of the 9 kinds of representative antibiotics in honey 图 3 牛奶添加回收中 9 种代表抗生素提取离子色谱图 Extracted ion chromatograms of the 9 kinds of representative antibiotics in milk 3773

6 Fig. 4 TIC of 31 kinds of antibiotics in standard solution Compounds 表 2 鱼肉, 牛奶, 蜂蜜中 31 种抗生素化合物的添加回收率 Table 2 Recoveries of the 31 antibiotics spiked in cosmetic samples (n=6) Add Level (μg/l) Average(%) RSD(%) Average(%) RSD(%) Average(%) RSD(%)

7 12, : 续表 2 Compounds Add Level (μg/l) Average(%) RSD(%) Average(%) RSD(%) Average(%) RSD(%)

8 续表 2 Compounds 5 Add Level (μg/l) Average(%) RSD(%) Average(%) RSD(%) Average(%) RSD(%) 实际样品的检测 Q-Exactvie,, 200, 4 结论 Q-Exactvie, 12 min 31,, 参考文献 [1]. [M]. :, Pang GF. Modern analytical technology for pesticide residue and veterinary drug residue [M]. Beijing: Science Press, [2] Regulation (EC) No 1223/2009 of The European Parliament and of the Council of 30 November 2009 on cosmetic products [Z]. [3]. [M]. :, Zhu BL. Animal drug residues in food [M]. Shanghai: Shanghai Science and Technology Press, 1994 [4] Wu ZJ, Peng Y. Analysis of penicillin antibiotics in cosmetics using ultra performance liquid chromatography [J]. Adv Mater Res, 2011, : [5] Jin W, Yang YJ, Wang WY, et al. Simultaneous determination of antibiotics in anti-acne cosmetics by rapid LC with DAD [J]. Chromatographia, 2009, 69: [6],,. 12 [J]., 2007, 25(2): Zheng HH, Wang P, Li J. Determination of 12 sulfonamides in cosmetics by ultra performance liquid chromatography [J]. Chin

12, : - 3777 J Chromatogr, 2007, 25(2): 238 240. [7],,,. 19 [J]., 2013, 41(6): 931 935. Chen J, Zheng R, Ji S, et al.

9 12, : J Chromatogr, 2007, 25(2): [7],,,. 19 [J]., 2013, 41(6): Chen J, Zheng R, Ji S, et al. Determination of 19 quinolone antibiotics in cosmetics by ultra performance liquid chromatography [J]. Chin J Anal Chem, 2013, 41(6): [8] Wu CS, Zhang JL, Qiao YL, et al. Simultaneous determination of 14 β-lactam antibiotics in cosmetic products by liquid chromatography tandem mass spectrometry method [J]. Chin Chem Lett, 2011, 22: [9],,,. - [J]., 2009, 27(1): Liu HL, Li F, Yang R, et al. Determination of common antibiotics and metronidazole in cosmetics by ultra performance liquid chromatography tandem mass spectrometry[j]. Chin J Chromatogr, 2009, 27(1): [10],,, [J]., 2008, 36(12): Ma Q, Wang C, Wang X, et al. Simultaneous determination of 22 sulfonamides in cosmetics by ultra performance liquid chromatography tandem mass spectrometry [J]. Chin J Anal Chem, 2008, 36(12): [11],,,. - / [J]., 2013, 31(10): Xiu XL, Luo X, Niu ZY, et al. Screening and confirmation of carcinogenic dyes in textiles by high performance liquid chromatography linear ion trap/orbitrap high resolution mass spectrometry [J]. Chin J Chromatogr, 2013, 31(10): [12] Hurtaud-Pessel D, Jagadeshwar-Reddy T, Verdon E. Development of a new screening method for the detection of antibiotic residues in muscle tissues using liquid chromatography and high resolution mass spectrometry with a LC-LTQ-Orbitrap instrument [J]. Food Addit Contam, 2011, 28(10): [13],,,. 15 [J]., 2007, 25(4): Ma Q, Wang C, Wang X, et al. Simultaneous determination of 15 hormones in cosmetics using ultra performance liquid chromatography [J]. Chin J Chromatogr, 2007, 25(4): [14] Pichler P, Köcher T, Holzmann J, et al. Peptide labeling with isobaric tags yields higher identification rates using itraq 4-plex compared to TMT 6-Plex and itraq 8-Plex on LTQ Orbitrap [J]. Anal Chem, 2010, 82(15): [15] US EPA. Title 40: Part 136, Appendix B to Part 136. [EB/ OL]. [ ]. http: // gov/fdsys/pkg/cfr title40- vil23/pdf/cfr-2011-title40-vol23- part136-appb. pdf 作者简介 ( 责任编辑 : 张宏梁 ) 王培锋, 中级职称, 主要研究方向为兽药残留分析 @qq.com

DOI /j.issn Food Research And Development UPLC-MS/MS 5 0.

DOI /j.issn Food Research And Development UPLC-MS/MS 5 0. 118 217 5 38 1 DOI 1.3969/j.issn.5-6521.217.1.28 Food Research And Development 1 2 1 2 1 2 1 2 1 2 1 2 1. 731 2. 731 - UPLC-MS/MS 5. 1 mol/l ProElut PXC 5 % - Thermo Hypersil Gold mm 2.1 mm 1.9 μm ESI