Sensitive method for the Determination of Colchicine in human plasma by gradient UPLC-ESI-MS/MS: separation of endogenous Interference

The objective of the current research work was to develop a sensitive, specific, accurate and precise method for
the determination of Colchicine in human plasma using Liquid Chromatography Electron Spray IonizationTandem Mass Spectrometry. Extraction was done using solid phase extraction technique. Ammonium Acetate
10mM with Acetonitrile was used as mobile phase with gradient programme of 6 minutes on Innoval, C18,
125mm x 4.6mm, 5µ column. Detection was performed on ESI positive mode using MRM transitions
400.400/358.300 for colchicine and 406.400/362.000 for colchicine D6 (ISTD) on API 4000 coupled with
Shimazdu Nexara UPLC. Developed method was found to be selective, specific and the Calibration Curve was
found to be linear over the range 0.075 to 10.091ng/mL. Endogenous interferences were separated from the peak
of interest. Method validation was conducted as per EMEA guideline and the developed method can be applied
to Pharmacokinetic/Bioequivalence studies.