Liquid chromatography method for quantifying d-threo-1-phenyl-2-palmitoylamino-3-morpholino-1-propanol (d-threo-PPMP) in mouse plasma and liver

A high-performance liquid chromatography (HPLC) method was developed to measure levels of d-threo-1-phenyl-2-palmitoylamino-3-morpholino-1-propanol (d-threo-PPMP) in mouse plasma and liver. d-threo-PPMP was measured by HPLC with a Luna Pheny-Hexyl column (5 μm, 250 mm × 4.6 mm) employing UV detection at 210 nm using a mobile phase of potassium phosphate buffer (20 mM, pH 3.0)–acetonitrile in a 45:55 (v/v) ratio. d-threo-1-phenyl-2-pentadecanoylamino-3-morpholino-1-propanol (PC15MP) was employed as an internal standard (IS). The lower limit of quantitation (LLOQ) was 0.3 μg/ml. The assay was linear over a concentration range of 0.3–10 μg/ml, with acceptable precision and accuracy. Assayed in plasma, the intra- and inter-day validation for all coefficients of variation (R.S.D.%) were found less than 15%. The method was applied to samples from athymic (nu/nu) mice treated with d-threo-PPMP by intraperitoneal injection. d-threo-PPMP levels of ∼10–20 μg/ml (∼20–40 μM) in plasma and ∼45 μg/g in liver were obtained. The present method can be used to quantify d-threo-PPMP in mice for bioavailability and dose-response studies.