ASE/UHPLC-FLD Determination of Florfenicol and Florfenicol Amine Residues in Pigeon and Quail Eggs
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摘要:
目的 建立鸽蛋和鹌鹑蛋中尼考(florfenicol,FF)及其主要代谢产物氟苯尼考胺(florfenicol amine,FFA)残留检测的加速溶剂萃取/超高效液相色谱-荧光检测(ASE/UPLC-FLD)法,为动物源性食品中FF和FFA残留检测提供新的安全可靠的方法。 方法 采用加速溶剂萃取(ASE)法,用乙腈﹕氨水(98﹕2,V/V)为提取剂萃取样品,用乙腈饱和的正己烷去脂进行纯化,使用由0.005 mol·L-1 NaH2PO4、0.003 mol·L−1十二烷基硫酸钠和0.05%三甲胺组成的流动相供UPLC-FLD检测。 结果 鸽蛋和鹌鹑蛋中FF和FFA在定量限(LOQ)~400 μg·kg−1范围内,目标物的峰面积与其浓度均呈现良好的线性关系,相关系数(R2)均大于0.999 2。样品在LOQ、50.0、100.0、200.0 μg·kg−1的加标水平下的平均回收率为84.69%~98.04%,相对标准偏差(RSD)均低于3.7%。鸽蛋中FF的检测限(LOD)和LOQ为4.8 μg·kg−1和11.2 μg·kg−1,FFA的LOD和LOQ为1.9 μg·kg−1和4.8 μg·kg−1;鹌鹑蛋中FF的LOD和LOQ为4.7 μg·kg−1和10.6 μg·kg−1,FFA的LOD和LOQ为1.8 μg·kg−1和4.6 μg·kg−1。 结论 该方法快速、简单、灵敏度高,适合禽蛋中FF及其代谢产物FFA的检测。 Abstract:Objective To establish an analytical method for simultaneous detections of florfenicol (FF) and its metabolite, florfenicol amine (FFA), in animal-derived foods. Method Samples of pigeon and quail eggs were extracted by the accelerated solvent extraction (ASE) method with acetonitrile-ammonia (98:2, V/V) as the solvent. The extract was degreased and purified with acetonitrile-saturated n-hexane. A mobile phase consisting of 0.005 mol·L−1 NaH2PO4, 0.003 mol·L−1 sodium lauryl sulfate and 0.05% trimethylamine was used in the ultra-high performance liquid chromatography-fluorescence (ASE/UPLC-FLD) determination of FF and FFA contents. Result Highly correlated linear relationships (R2>0.999 2) within the limit of quantitation (LOQ) were achieved between the peak area and concentration on the target compounds. The average recoveries of the method at the spiked levels of 50.0, 100.0, 200.0 μg·kg−1 and LOQ ranged from 84.69% to 98.04% with RSDs less than 3.7%. The method delivered the limit of detection (LOD) and LOQ on FF in the pigeon eggs at 4.8 µg·kg−1 and 11.2 µg·kg−1, respectively, and those on FFA, at 1.9 µg·kg−1 and 4.8 µg·kg−1, respectively; whereas, those on FF in the quail eggs, at 4.7 µg·kg−1 and 10.6 µg·kg−1, respectively, and those on FFA, at 1.8 µg·kg−1 and 4.6 µg·kg−1, respectively. Conclusion The newly developed analytical methodology was rapid, simple, sensitive, and considered suitable for FF and FFA determinations in pigeon and quail eggs. -
图 1 标准品、空白鸽蛋/鹌鹑蛋、空白鸽蛋/鹌鹑蛋样品添加标准品(FF和FFA 50 μg·kg−1)色谱
注:A,氟苯尼考和氟苯尼考胺标准品(50.0 μg·kg−1)色谱图;B,空白鸽蛋色谱图;C,加标鸽蛋样品色谱图;D,空白鹌鹑蛋色谱图;E,加标鹌鹑蛋样品色谱图。
Figure 1. Chromatograms of standards, pigeon egg/quail egg blank, pigeon egg/quail egg spiked with 50.0 µg·kg−1 FF and FFA
Note:A=Chromatogram of FF and FFA standards (50.0 µg·kg−1); B=Chromatogram of pigeon egg blank; C=Chromatogram of spiked pigeon egg samples; D=Chromatogram of quail egg blank; E=Chromatogram of spiked quail egg samples.
表 1 不同萃取剂对氟苯尼考和氟苯尼考胺提取回收率的影响
Table 1. FF and FFA recoveries by different solvents
(单位:%) 分析物
Analyte基质
Matrix萃取剂乙腈∶氨水 Extraction reagent acetonitrile-ammonia 99∶1 98∶2 97∶3 96∶4 95∶5 氟苯尼考
FF鸽蛋
Pigeon egg92.28±2.41 93.12±1.95 83.82±1.88 76.12±3.01 61.00±2.19 鹌鹑蛋
Quail egg92.73±2.23 94.24±2.47 86.24±2.49 74.48±2.41 68.53±2.48 氟苯尼考胺
FFA鸽蛋
Pigeon egg88.23±2.24 96.32±2.14 86.53±2.70 73.09±2.13 64.82±2.27 鹌鹑蛋
Quail egg86.46±1.79 92.96±1.92 84.65±2.51 76.20±1.92 70.38±1.84 表 2 空白鸽蛋中FF和FFA添加回收率和精密度(n=6)
Table 2. Recovery and precision of determinations on pigeon egg spiked with FF and FFA (n=6)
分析物
Analyte添加质量比
Adding level/(μg·kg−1)回收率
Recovery/%相对标准偏差
RSD/%日内相对标准偏差
Intra-day RSD/%日间相对标准偏差
Inter-day RSD/%氟苯尼考
FF11.2a 85.33±2.39 2.80 2.96 3.15 50 96.42±2.10 2.18 3.84 4.52 100b 98.04±3.63 3.70 3.25 4.97 200 97.93±2.56 2.61 2.94 3.31 氟苯尼考胺 FFA 4.8a 84.69±1.84 2.17 3.42 4.43 50 94.08±2.43 2.58 2.70 3.06 100.0b 95.14±2.90 3.05 3.86 5.18 200 96.82±1.73 1.79 2.21 3.60 注:a.定量限;b. 最高残留限量,表3同。
Note:a=LOQ; b=Maximum residue limit. The same for Table 3.表 3 空白鹌鹑蛋中FF和FFA添加回收率和精密度(n=6)
Table 3. Recovery and precision of determinations on quail egg spiked with FF and FFA (n=6)
分析物
Analyte添加质量比
Adding level/(μg·kg−1)回收率
Recovery/%相对标准偏差
RSD/%日内相对标准偏差
Intra-day RSD/%日间相对标准偏差
Inter-day RSD/%氟苯尼考
FF10.6a 87.26±3.23 3.70 4.98 5.33 50 95.73±2.14 2.24 2.65 3.58 100b 95.36±2.91 3.05 3.73 4.49 200 96.95±2.42 2.50 2.91 3.52 氟苯尼考胺
FFA4.6a 86.32±2.49 2.88 3.60 3.92 50 96.73±2.75 2.84 4.13 5.43 100.0b 95.62±3.21 3.36 3.54 5.62 200 96.84±3.36 3.47 4.86 5.84 -
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