[Study of diphacinone in biological samples by high performance liquid chromatography/diode array detector]

An analytical approach has been developed for high performance liquid chromatographic determination of diphacinone extracted from liver, blood, urine and kidney of rabbit by solid phase extraction (SPE) cartridges (using SAX, CN or SILICA GEL) with coumarin as the internal standard. Diphacinone was separated by reversed-phase gradient chromatography with DAD detection at 286 nm. The Analytical column was Hypersil BDS C18(150 mm x 4.6 mm i.d., 5 microns) and the guard column was Phenomenex ODS(4 mm x 3.0 mm i.d.). The mobile phase was a gradient mixture of aqueous solution (A) and methanol solution (B) both containing 0.5% ion pair A. There was a good linear relationship between the concentration of diphacinone and the ratio of peak areas of diphacinone and coumarin (internal standard) (r = 0.9999). The linear range was 1 mg/L-100 mg/L, and the lower detection limit was 5 ng (S/N = 3). The average recoveries of diphacinone in urine, blood and liver were 88.4% (n = 3, RSD = 1.25%, SPE by CN column), 82.2% (n = 3, RSD = 1.67%, SPE by SAX column), 91.0% (n = 3, RSD = 2.77%, SPE by SILICA GEL column), respectively.