Synthesis of the flavonoid 3′,4′,5′-trimethoxyflavonol and its determination in plasma and tissues of mice by HPLC with fluorescence detection

3′,4′,5′-Trimethoxyflavonol (TMFol) was synthesized as a potential colorectal cancer chemopreventive agent. An HPLC method for determination for TMFol in murine plasma and tissues was developed and validated using human plasma. Analyte was separated (C18 column; fluorescence detection 330 nm excitation, 440 nm emission) using 69% methanol and 0.1 M ammonium acetate buffer (pH 5.1) as mobile phase. The method was linear for 50–2500 ng/ml plasma and 0.05–10 μg/g tissue (r > 0.99). TMFol was recovered from plasma or tissues using solid phase columns or organic solvent protein precipitation, respectively. Recovery at low, medium and high concentrations was 97.6–107.3%, with inter- and intra-day coefficients of variation of <10%. The lower limit of quantitation for plasma was 50 ng/ml. The method was applied to measure steady-state TMFol plasma and tissue levels in mice which received dietary TMFol (0.2%).