HPLC-MS/MS Determination of Colchicin in Pig Urine

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 ℃. 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 10 μL 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%.