Direct analysis of crude plasma samples by turbulent flow chromatography/tandem mass spectrometry

Turbulent flow chromatography coupled to tandem mass spectrometry (TFC-MS-M S) has recently emerged as a potentially fast, sensitive and specific techn ique for the direct analysis of pharmaceutical compounds from crude plasma. TFC-MS-MS removes the need for time-consuming sample preparation procedure s such as solid-phase extraction (SPE) or liquid-liquid extraction (LLE). A relatively high flow rate combined with the use of an HPLC column with lar ge porous particles allows the on-line clean up and quantification of compo unds in plasma samples. Until now, the amount of plasma directly injected i nto TFC systems has rarely exceeded 30 muL in order to prevent rapid column degradation. Increasing the injection volume also induces high carry-over levels, particularly for drugs with basic and/or lipophilic properties. This paper describes the first generic TFC-MS-MS method developed in a 96-w ell format, which allows the direct injection of 200 muL of 1:1 diluted pla sma (equivalent to 100 muL neat plasma). An average of 390 injections was c arried outwith each extraction column. More than 2000 dog plasma samples we re injected into the system without any sign of carryover: The method was f ully validated over a 5 - 500 ng mL(-1) range for three basic compounds: do xazosin, CP122,288 and dofetilide. The imprecision was 1.2 to 8.3% for doxa zosin, 1.5 to 4% for CP122,288 and 1.6 to 9.2% for dofetilide. The inaccura cy ranged from 6% to 7.9%. This generic methodology was then used to assay two structurally unrelated development compounds, showing that the method a ccuracy and sensitivity were adequate for the early pharmacokinetic (PK) st udies in animals.