高效液相色谱-串联质谱法测定猪尿中秋水仙碱

HPLC-MS/MS Determination of Colchicin in Pig Urine

提出了猪尿中秋水仙碱的液相色谱-串联质谱检测方法。采用pH8.0磷酸盐缓冲溶液碱化5mL尿样,用乙酸乙酯提取两次,合并提取液,于40℃吹氮蒸干,用2 mL流动相溶解残渣。在色谱分离中选用Sunfire C_(18)色谱柱,流动相为甲醇-2mmol·L^(-1)乙酸铵溶液(含0.1%甲酸)混合溶液(40+60),流量为0.2mL·min,进样量为10μL。质谱测定中采用电喷雾正离子模式离子化、多反应监测模式测定秋水仙碱,外标法定量。猪尿中秋水仙碱在1～50μg·L^(-1)内呈线性,测定下限(10S/N)为0.2μg·L^(-1),方法的回收率为90%～100%,相对标准偏差(n=6)小于9.0%。

Colchicin in pig urine was determined by HPLC in hyphenation with tandem MS/MS. The pig urine sample was alkalified with pH 8. 0 phosphate buffer solution and extracted twice with ethylacetate. The combined extracts were blown with nitrogen to dryness at 40 ~C. The residue was taken up with 2 mL of the mobile phase, which is a mixed solution of methanol and 2 mmol · L ^-1 NH4OAc solution containing 0. 1%（by volume） of formic acid, mixed in the ratio of 40 to 60. Sunfire C18 chromatographic column and the mobile phase mentioned above were used in the HPLC separation, with 10μL of sample solution introduced and flow-rate of mobile phase of 0. 2 mL · min^-1. The ionization mode of ESI^＋ and MRM mode of detection were adopted in MS/MS determination, with quantification by external standard method. Linear relationship between values of peak area and mass concentration of colchicin in the range of 1-50 μg · L^-1 was obtained, and value of lower limit of determination （10S/N） of 0.2 μg · L ^-1 was found. Values of recovery were found in the range of 90%-100%, with RSD＇s （n= 6） less than 9. 0%.