5 12 Vol. 5 No. 12 2014 12 Journal of Food Safety and Quality Dec., 2014 何京澄 *, 王培锋, 宿立强, 温晓敬, 邱芳, 蔡雪, 周维政 (, 266001) 摘要 : 目的, 方法 GB/T21311-2007,, - (HPLC-MS/MS) (SEM) (AHD) (AOZ) (AMOZ) 4 55, ph, ph, 结果, GB/T21311-2007,, 结论 关键词 : ; ; ; Rapid determination of nitrofuran metabolite residues in collagen powder by high performance liquid chromatography-tandem mass spectrometry HE Jing-Cheng *, WANG Pei-Feng, SU Li-Qiang, WEN Xiao-Jing, QIU Fang, CAI Xue, ZHOU Wei-Zheng (Technical Center of Shandong Entry-Exit Inspection and Quarantine Bureau, Qingdao 266001, China) ABSTRACT: Objective To simplify the experimental process for determination of furan metabolite residues in collagen powder, and shorten the detection time of the experiment. Methods Based on GB/T21311-2007, the method was improved, and a high performance liquid chromatography-tandem mass spectrometry (HPLC-MS/MS) method was established to rapidly determine 4 kinds of nitrofuran metabolite residues in collagen powder, including nitrofurazone (SEM), nitrofurantoin (AHD), furaltadone (AOZ) and furazolidone (AMOZ). After derivation at 55, the derivatives were processed following the steps of centrifugation, ph adjustment, and centrifugation again, making it easier to adjust the ph value, while avoiding the problem found in the conventional test method using a separatory funnel where the sample emulsified into colloid and could not achieve the purpose of liquid-liquid separation. Results The experimental results showed no significant differences between the proposed method and the national standard GB/T21311-2007, while the detection time of the experiment was greatly reduced, and the experimental process was simplified. Conclusion This method is more suitable for experimental detection of large quantities of samples. KEY WORDS: nitrofuran metabolite residues of veterinary drugs; liquid chromatography-tandem mass spectrometry; collagen powder; emulsification * 通讯作者 :,, E-mail: 277467854@qq.com *Corresponding author: HE Jing-Cheng, Assistant Engineer, Technical Center of Shandong Entry-exit Inspection and Quarantine Bureau, Qingdao 266001, China. E-mail: 277467854@qq.com
12, : 3803 1 引言, [1-3],, (nitrofurazone, SEM) (nitrofurantoin, AHD) (furaltadone, AOZ) (furazolidone, AMOZ),,, [4-9],,, (LC-MS/MS) [10-13] 2 材料与方法 2.1 仪器与试剂 : Agilent2600 ( Agilent ); API4000Q-trap ( AB ), ; (CR22GII/CR21GII), (Caliper TurboVap LV), (IKA MS2), (SHA-B), ph (SARTORIUS TB10), : ( ), ( ), ( ), ( ), ( ), ( ), ( ), ( ), ( ), 2- (HPLC), ( ), : SEM AHD AMOZ AOZ Sigma-Aldrich, 99%; AMOZ-D 5, AHD -13 C 3, SCA 15 N 2-13 C HCL, AOZ-D 4 Witega, 99% :, 10 ml 1000 μg/ml 2.2 色谱条件 : Intersil ODS-3(150 mm 4.6 mm, 5 μm) ; : 20 ; : A: ; B: 0.1 % ; : 0~10 min: 10%A+90%B, 0.6 ml/min; 10~11 min: 95%A+5%B, 0.6 ml/min; 11.00~ 11.05 min: 95%A+5%B, 0.8 ml/min; 11.05~16 min: 10%A+90%B, 0.8 ml/min; 16.00 min: 10%A+90% B, 0.8 ml/min; : 30 µl; 2.3 质谱条件 : ; : ; : ; : 5500 V; : 0.448 MPa; : 0.193 MPa; : 0.344 MPa; : 500 ;, 1 2.4 样品的前处理方法 1.5 g ( 0.01 g) 50 ml, 1 g, 10 ml 0.125 mol/l, 60 µl 0.1 µg/ml 300 µl 0.015 g/ml2-, 2 min, 55 2 h, 60 Hz, (8000 r/min, 8 min, 15 ), 表 1 硝基呋喃代谢物的质谱条件 Table 1 MS parameters of nitrofuran metabolite /(m/z) /(m/z) /ev /V SEM 209.0/192.0, 209.0/166.0 209.0/166.0 18, 15 60, 60 AHD 249.0/133.9, 249.0/178.0, 249.0/103.9 249.0/133.9 18, 17, 32 60, 60, 60 AOZ 236.0/133.9, 236.0/103.9 236.0/133.9 19, 31 60, 60 AMOZ 335.1/291.0, 335.1/262.0 335.1/291.0 18, 25 60, 60 SCA 13 C 15 N 2 212.0/168.0 212.0/168.0 22 60 AHD 13 C 3 252.0/133.9 252.0/133.9 21 60 AOZ-D 4 240.0/133.9 240.0/133.9 20 60 AMOZ-D 5 340.0/296.9 340.0/296.9 15 60
3804 5 Fig. 1 1 4 Extracted ion chromatograms of 4 kinds of nitrofuran metabolite in standard solution 50 ml, 3 mol/l NaOH 1 mol/l KH 2 PO 4, 1 mol/l HCL PH 7.0~7.5, 10 ml, NaCl, 10 min, 10000 r/min 10 min, 15 ml, 40, 1mL 0.2%, 2.5 ml 1:1, 1 min, 8000 r/min 5 min, 0.22 µm, - 3 结果与讨论 3.1 提取与净化 40 50 55 60 2 h [14], : 55,,, 55 GB/T21311-2007 37 16 h,,,, GB/ T21311-2007 t, 0.50 µg/kg. SEM AHD AMOZ AOZ t t 0.05, 95% GB/T21311-2007 PH, PH,
12, : 3805 ; PH,,, GB/T21311-2007, ph GB/T21311-2007,, ;,,, GB/T21311-2007,,, ;,,, 3.2 实际样品的测定,, SEM: 16.3 µg/kg 2 表 2 硝基呋喃代谢物 GB/T21311-2007 与本实验的 t 检验对比 Table 2 t-test comparison of GB/T21311-2007 and this method of nitrofuran metabolite determination SEM AHD AOZ AMOZ GB/T21311-2007 GB/T21311 2007 GB/T21311 2007 GB/T21311 2007 0.495 0.489 0.533 0.512 0.455 0.494 0.495 0.516 0.478 0.489 0.479 0.522 0.509 0.472 0.518 5.529 0.501 0.479 0.45 0.488 0.502 0.534 0.463 0.490 0.499 0.455 0.488 0.511 0.508 0.536 0.452 0.476 0.523 0.512 0.533 0.501 0.477 0.491 0.461 0.482 0.467 0.475 0.479 0.521 0.498 0.534 0.541 0.537 t 0.96 1.07 1.30 1.01 v 10 10 10 10 t 0.05 2.228 2.228 2.228 2.228 Fig. 2 2 Extracted ion chromatograms of nitrofuran metabolite in positive solution
3806 5 Table 3 表 3 4 种呋喃代谢物的线性方程 相关系数及检出限 Linear equations, correlation coefficient and limits of quantitation of 4 kinds of nitrofuran metabolite µg/kg SEM Y=0.82X 3.22e 7 0.9998 0.5 AHD Y=0.673X-5.23e 7 0.9990 0.5 AOZ Y=0.619X-1.32e 8 0.9999 0.5 AMOZ Y=0.835X+1.21e 6 0.9994 0.5 Table 4 表 4 4 种呋喃代谢物回收率及精密度 (n=6) Recoveries and relative standard deviations of 4 kinds of nitrofuran metabolite (µg/kg) (µg/kg) (µg/kg) % SEM 0.50, 1.00, 3.00 102.07, 99.70, 101.18 6.06, 6.41, 6.00 5.9, 6.4, 5.9 AHD 0.50, 1.00, 3.00 96.35, 103.18, 103.63 8.95, 5.89, 8.53 9.3, 5.7, 8.2 AOZ 0.50, 1.00, 3.00 102.1, 98.52, 99.23 7.92, 5.83, 6.97 7.8, 5.9, 7.0 AMOZ 0.50, 1.00, 3.00 92.98, 102.23, 98.73 7.10, 7.16, 6.15 7.6, 7.0, 6.2 3.3 线性关系及检出限 4 0~20.0 ng/ml, >0.99. 4 0.5 µg/kg( ) [15] 3.4 样品的添加回收率和精密度, 0.5 µg/kg( ) 1.0 µg/kg 3.0 µg/kg, 6, 4 4 结论 80%~120%, 10%, 2002/657/EC,,, 4 参考文献 [1],,,. [J]., 2004, 20(4): 382 384. Yu JX, Hu XZ, Lin YF, et al. Residues in tandem mass Spectrometry in honey and fish nitrofuran antibiotic metabolites [J]. J Anal Sci, 2004, 20(4): 382 384. [2]. - 4 [J]., 2007, 22(1): 68 72. Chen RQ. Liquid chromatography-electro spray tandem mass spectrometry detection eel muscle tissue in four kinds of residues of nitro furan metabolites [J]. Fujian J Agric Sci, 2007, 22(1): 68 72. [3],,,. [J]., 2008, 27(12): 76 79. Li YP, Lin YH, Jia DF, et al. Ultra-high performance liquid chromatography tandem mass spectrometry detection eel shrimp nitrofuran metabolite residues in new technology [J]. Chin J Anal Lab, 2008, 27(12): 76 79. [4],. [M]. :, 2000. Zhang ZQ, Zheng M. Animal Drug User Manual [M]. Beijing: China Agricultural University Press, 2000. [5],,,. - [J]., 2008, 18(1): 19 21. Zhang JL, Gao HP, Shen WJ, et al. Ultra-high performance liquid chromatography-tandem mass spectrometry eel nitrofuran metabolites residues of [J]. Chin J Health Lab Technol, 2008, 18(1): 19 21. [6],,,. - [J]., 2008, 18(6): 978 980. Yang N, Zhao J, Quan WY, et al. Liquid chromatography - tandem mass spectrometry nitrofuran metabolites in aquatic products [J]. Chin J Health Lab Technol, 2008, 18(6): 978 980.
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