4 4 Vol. 4 No. 4 13 8 Journal of Food Safety and Quality Aug., 13-9 李 学 民 *, 曹 彦 忠, 张 进 杰, 母 健, 王 蕾, 王 贺 琴 (, 066004) 摘 要 : 目 的 - (HPLC-MS-MS) 9 方 法 -, C 18, -, - 结 果 1.0~ μg/kg(loq~10loq),, 0.999 LOQ 2 LOQ 4 LOQ 10 LOQ 81.4%~109.7%, 2.81%~7.12%, : G V 1.0 μg/kg; 2.0 μg/kg 结 论,, 关 键 词 : - ; ; ; Determination of 9 penicillins in Fugu and eel by high pressure liquid chromatography-electrospray tandem mass spectrometry LI Xue-Min *, CAO Yan-Zhong, ZHANG Jin-Jie, MU Jian, WANG Lei, WANG He-Qin (Qinhuangdao Entry-Exit Inspection and Quarantine Bureau, Qinhuangdao 066004, China) ABSTRACT: Objective A sensitive method was developed for the determination of 9 penicillins in Fugu and eel by high pressure liquid chromatography-electrospray tandem mass spectrometry (HPLC-MS/MS). Methods Samples were extracted with acetonitrile-ammonia water, separated on a C 18 column by gradient elution with acetonitrile and 0.3% acetic acid solution as mobile phase. Identification was achieved by electrospray ionization(esi) in positive mode. Results The method showed a good linearity over the range of 1.0~ μg/kg for 9 compounds of penicillins with r 0.999. Recoveries of 9 penicillins were between 81.4% and 109.7% (n=6) at spiked levels of LOQ, 2LOQ, 4LOQ and 10LOQ with relative standard deviations (RSD of 2.81% 7.12%). The limit of quantitation(loq) for nafcillin, penicillin G, piperacillin, penicillin V and oxacillin were 1.0 μg/kg and the LOQ for amoxicillin, ampicillin, cloxacillin and dicloxacillin were 2.0 μg/kg. Conclusion This method is sensitive and accurate, and it is suitable for the analysis of 9 penicillins in aquatic products. KEY WORDS: high pressure liquid chromatography-electrospray tandem mass spectrometry (HPLC-MS-MS); penicillins; Fugu; eel * 通 讯 作 者 :,, E-mail: ciqlixm@163.com *Corresponding author: LI Xue-Min, Senior Engineer, Qinhuangdao Entry-Exit Inspection and Quarntine Bureau of P.R.China Inspection and Quarntine Technique Centre, No.1, Liupanshan Road, Qinhuangdao 066004, China. E-mail:ciqlixm@163.com
1166 4,,, [1],,,,, [2,3], FDA : (MRL) 10 μg/kg, G (MRL) 0, (MRL) 10 μg/kg, (MRL) 4~30 μg/kg,, [4], ; [5],, ; [6],,, ; [7], ; [8],,, ; - [9],, ; [10-11],, ; - [12-14],, (0.4~5 μg/kg),,, - 1 材 料 与 方 法 1.1 试 剂 与 材 料 ( ); ( ); ( ); ( ); ( ); (CAS26787-78-0) (CAS69-53-4) (CAS59703-84-3) G(CAS69-57-8) V(CAS132-98-9) (CAS7177-50-6) (CAS7240-38-2) (CAS642-78-4) (CAS13412-64-1) Sigma 9 :, 1.0 mg/ml, 25 9 :, 1.2 仪 器 与 设 备 - (API 3000, ); (Angilent 1100); ; ; ; ; ; ph, ±0.02( Precisa ph 900 ) 1.3 测 定 条 件 : ZORBAX SB-C 18 3.5 μm, 150 mm 2.1 mm ( ); 0 μl/min; : 30 ; : μl: (A) 0.3 - (B), 5 A 3 min, 10 min A 50%, 5 min 75%, 7 min A 5% : : (ESI); : ; : (MRM); (IS): 5500 V; (NEB): 0.055 MPa; (CUR): 0.079 MPa; (AUIX): 6 L/min; 400 (DP) (FP) (CE) (CXP) 1 1.4 操 作 步 骤 5 g 50 ml, 2 g, 25 ml 0.15 ml 1 min, 1000 r/min 10 min,,,, 10 ml, 1 min, 100 ml, 40, 2 ml,,, -
4, : - 9 1167 Table 1 表 1 质 谱 参 数 Parameters of mass spectrometry (m/z) (m/z) (ev) (V) (V) (V) 阿 莫 西 林 366/114 366/8 336/8 30 19 21 90 10 氨 苄 西 林 350/192 350/160 350/160 23 90 10 哌 拉 西 林 518/160 518/143 518/143 35 35 27 25 90 10 青 霉 素 G 335/160 335/176 335/160 23 90 10 V 351/160 351/192 351/160 15 40 90 10 402/160 402/243 402/160 23 90 10 436/160 436/277 436/160 21 22 90 10 415/199 415/171 415/199 23 52 23 90 10 470/160 470/311 470/160 22 90 10 2 结 果 与 分 析 2.1 提 取 溶 液 的 选 择,, [8],, 9 9, 2 3, [8], ph,, 10.3%~35.5%, 7 89.7%~98.4%;, - 9, 2 3 + 0.6%, 9 94.7%~102.5%,,, + 0.6% 2.2 液 相 色 谱 分 析 柱 的 选 择 μbondapak C 18 300 mm 4.0 mm ZORBAX SB-C 18 (150 mm 2.1 mm, 3.5 μm) 9,, μbondapak C 18 1.5 ml/min,, 9, ZORBAX SB-C 18, μbondapak C 18,, ZORBAX SB-C 18 9 2.3 质 谱 条 件 的 确 定, 5 μl/min G V, 9 (Q1 ), 366 350 518 335 351 402 436 415 470, ( ), (DP), (CE),,,,, 9, 1 2.4 线 性 范 围 和 方 法 检 出 限 G V
1168 4 1.25~100 ng/ml(0.5 LOQ~40 LOQ); 2.5~0 ng/ml(0.5 LOQ~40 LOQ),, μl,, G V 0.025~2 ng ; 0.05~4 ng,, 4, 9 1.0~ μg/kg(loq~10 LOQ),, G V 1.0 μg/kg; 2.0 μg/kg, 10,, G V 1.0 μg/kg; 2.0 μg/kg Table 2 表 2 不 同 浓 度 乙 腈 - 氨 水 对 河 豚 鱼 样 品 中 9 种 青 霉 素 回 收 率 的 影 响 Effects of different extracts of acetonitrile and ammonia solution on the recoveries(%) of 9 penicillins in Fugu + 0.2% + 0.4% + 0.6% + 0.8% 阿 莫 西 林 15.1 76.2 88.6 94.7 93.2 氨 苄 西 林 35.5 79.7 89.5 96.4 92.9 哌 拉 西 林 95.2 96.6 97.1 96.7 98.2 青 霉 素 G 94.7 96.4 94.5 94.8 95.1 V 96.8 97.2 96.2 98.4 97.8 94.1 95.4 97.1 94.9 96.1 97.5 96.1 95.7 102.5 97.4 95.7 94.3 96.9 97.4 97.5 98.4 95.8 97.3 101.9 95.9 Table 3 表 3 不 同 浓 度 乙 腈 - 氨 水 对 鳗 鱼 样 品 中 9 种 青 霉 素 回 收 率 的 影 响 Effects of concentration of acetonitrile and ammonia solution on the recoveries(%) of 9 penicillins in eel + 0.2% + 0.4% + 0.6% + 0.8% 阿 莫 西 林 10.3 74.6 89.1 94.7 92.8 氨 苄 西 林 29.6 78.9 87.7 95.6 95.7 哌 拉 西 林 94.8 93.8 94.8 96.7 95.1 青 霉 素 G 96.2 97.2 95.4 97.2 94.2 V 97.1 96.9 94.2 95.7 97.8 89.7 95.1 93.8 97.1 94.3 93.5 94.7 97.5 96.8 97.4 94.7 96.3 98.2 102.3 95.3 96.3 97.2 95.4 100.9 102.3
4, : - 9 1169 2.5 方 法 的 回 收 率 和 精 密 度, 2~100 μg/kg(loq~10 LOQ),,,, -, 5 5, 81.4%~109.7%, 2.81%~7.12% LC-MS 1 Table 4 表 4 9 种 青 霉 素 线 性 方 程 和 相 关 系 数 Linear equation and correction coefficient of 9 penicillins (ng) (R) 阿 莫 西 林 0.05~4 Y=6670X+1480 0.9992 氨 苄 西 林 0.05~4 Y=13800X+2170 0.9992 哌 拉 西 林 0.025~2 Y=4450X+ 0.9999 青 霉 素 G 0.025~2 Y=10500X+1550 0.9999 V 0.025~2 Y=14400X+11 0.9995 0.025~2 Y=16600X+936 0.9994 0.05~4 Y=10000X+1070 0.9997 0.025~2 Y=30800X+852 0.9991 0.05~4 Y=6410X+703 0.9995 表 5 Table 5 河 豚 鱼 和 鳗 鱼 中 九 种 青 霉 素 添 加 回 收 率 (n=6) Recoveries of 9 penicillins in Fugu and eel (n=6) LOQ 2LOQ 4LOQ 10LOQ LOQ 2LOQ 4LOQ 10LOQ 阿 莫 西 林 (%) 84.3 97.8 98.7 98.3 82.4 87.5 89.8 94 氨 苄 西 林 (%) 86.1 96.2 91.3 99.7 86.3 89.4 91.6 89.6 哌 拉 西 林 (%) 87.6 91.5 94 89.9 89.6 91.2 93.4 92.7 青 霉 素 G(%) 90.2 97 89.7 96.4 91 86.3 94.5 93.6 V(%) 86.2 102.6 86.9 92.4 94.5 85.1 89.9 95.1 (%) 92.3 91.8 92.1 93.5 93.7 89.7 87.3 94.8 (%) 87.1 89.6 95.6 95.7 90.8 92.5 90.4 88.7 (%) 89.2 89.6 90.8 94.8 87.6 94 93.2 86.8 (%) 86.2 92.3 109.7 91.2 81.4 96.1 91.5 91.3 (%) 87.7 94.2 94.3 94.7 88.6 90.2 91.3 91.8 SD 2.47 4.39 6.71 3.24 4.6 3.62 2.22 2.93 RSD(%) 2.81 4.66 7.12 3.42 5.18 4.02 2.43 3.19
1170 4 : (a)(b)(c) 1 Fig. 1 MRM of 9 penicillins in Fugu
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