高效液相色谱-串联质谱法检测奶中克拉维酸残留

Determination of clavulanic acid residue in milk by high performance liquid chromatography-tandem mass spectrometry

采用高效液相色谱-串联质谱(HPLC-MS/MS)建立了克拉维酸在奶中的残留检测方法。2 g样品经乙醇沉淀蛋白质后,转入鸡心瓶中旋转蒸发浓缩至0.5 mL左右,用乙酸铵定容,净化后检测。流动相为乙腈和0.1%甲酸水,梯度洗脱,经Luna 5u C8色谱柱分离,采用电喷雾电离,多反应监测负离子模式对克拉维酸进行定量分析。采用基质匹配法对奶中克拉维酸的含量进行标准校正,在克拉维酸含量为10～400μg/kg范围内呈现良好的线性关系,相关系数大于0.999;奶中加标样品的检出限(LOD,按信噪比(S/N)≥3计)为10μg/kg,定量限(LOQ,S/N≥10)为20μg/kg。在定量限、1/2最高残留限量、最高残留限量、2倍最高残留限量添加水平下,奶中克拉维酸的平均回收率为80.00%～91.25%,相对标准偏差为5.60%～8.77%。该方法可用于奶中克拉维酸残留的分析检测。

An analytical method was developed for the determination of clavulanic acid （CLAV） in milk by high performance liquid chromatography-tandem mass spectrometry（ HPLC- MS/MS）. A 2 g milk sample was deproteinized by ethanol. The supernatant was transferred into a pear-shaped bottle to be evaporated to about 0.5 mL, and the residue was dissolved with ammonium acetate solution. The sample was determined by HPLC-MS/MS after the purifica- tion. The chromatographic separation was achieved on a Luna 5u C8 column using 0. 1% formic acid in water and acetonitrile as mobile phases with gradient elution. The identification of CLAV was carried out by MS/MS equipped with electrospray ionization in negative scanning and multi- ple reaction monitoring （MRM） modes. Matrix-matched calibration standard was used for the quantification. The calibration curve showed perfect linear in the range of l0 -400μg/kg with the correlation coefficient of 0. 999. The limit of detection （LOD, S/N≥3 ） was l0 μg/kg in milk, and the limit of quantification （LOQ, S/N≥ 10） was 20μg/kg. The mean recoveries var- ied from 80.00% to 91.25% at the four spiked levels of LOQ, I/2MRL （the maximum residue limit）, MRL, and 2MRL with the relative standard deviations of 5.50% -8.77%. In conclusion, the established method can be applied for the determination of CLAV residues in milk.