高效液相色谱-串联质谱法测定猪组织中的土拉霉素

Determination of Tulathromycin in Pork Tissues by Liquid Chromatography-Tandem Mass Spectrometry

建立了高效液相色谱-串联质谱法测定土拉霉素在猪肝脏、肾脏、肌肉、皮脂和肺脏中的残留检测方法，并初步探讨了土拉霉素的基质效应。样品用乙腈提取，涡旋振荡离心后经正己烷脱脂，C18固相萃取柱净化后，40℃氮气吹干，用50％乙腈定容。以Penomenex Luna C18柱为色谱柱，乙腈-2mmol／L乙酸铵水溶液（含0．1％甲酸）为流动相，梯度洗脱；流速0．25mL／min；柱温35℃。质谱条件为电喷雾离子源，检测方式为正离子多离子反应监测（MRM），外标法定量。在10．0～500μg／kg范围内，方法的线性关系良好（r〉0．99）。5种基质中土拉霉素的检出限为3～5μg/kg，定量下限均为10μg/kg。在高、中、低3个加标水平下的平均回收率为74％～96％，批内相对标准偏差（RSD）为4．0％～16％，批间RSD为7．0％-15％。该法准确、灵敏，适用于猪组织中土拉霉素的测定。

A high performance liquid chromatography - tandem mass spectrometric （ HPLC - MS/MS） method was developed for the determination of tulathromycin in pork tissues, such as liver, kidney, muscle, skin, fat and lung samples. The matrix effects of tulathromycin were also discussed in this paper. The samples were extracted with acetonitrile, and defatted with n-hexane. The extract was cleaned up on a Cis solid phase extraction（SPE） cartridge. After washed with 4% amonium hydroxide in methanol and then condensed to nearly dryness at 40℃ water bath, the extract was redissolved with 50% acetonitrile. Tulathromycin was separated on a Penomenex Luna Cls column with a mobile phase of acetonitrile - 2 mmol/L ammonium acetate （ containing 1% fomic acid） at a flow rate of 0. 25 mL/min. The column temperature was set at 35 ~C. The identification was carded out by MS/MS with electrospray ionization under positive scan and multiple reaction monitoring（MRM） mode. The quantitation was performed by the external standard method. The results showed that the calibration curves for all the tissues were linear（r 〉 0. 99） in the range of 10. 0 -500 μg/kg. The limits of detection （LODs） of tulathromycin in five tissues were in the range of 3 - 5 μg/kg and the limits of quantita- tion（LOQs） for all the tissues were 10 μg/kg. The spiked recoveries at three spiked concentration levels were in the range of 74% - 96%. The RSDs of intra-day and inter-day were in the range of 4.0% - 16% and 7.0% - 15% , respectively. The method was proved to be simple and accurate, and could be applied in the determination of tulathromycin in real samples.