Application of column-switching liquid chromatography-tandem mass spectrometry for the determination of pharmaceutical compounds in tissue samples.
Information on plasma-tissue distribution which is important for drug development may be obtained by "in silico" prediction tools. To support the validation of computer models, drug concentrations in rat plasma and tissues (brain, liver, kidney, testes, spleen, gut, lung. heart, muscle, skin and fat) had to be determined. In our work, we established analytical assays for a variety of substances including nicardipine, nitrendipine, felodipine and benzodiazepines. Sample preparation had to be simple and method development as well as analytical run time short to allow a high sample throughput and to minimize resources. Column-switching HPLC after homogenization and protein precipitation served as an efficient, easy and rapid sample preparation method, followed by selective MS-MS detection. Optimization of the trapping procedure was performed in order to reduce the influence of endogenous interferences and to obtain good recovery. Chromatographic separation was necessary to increase the selectivity. The use of small analytical column dimensions (2.1 x 10 mm) was investigated to achieve higher sample throughput without compromising the assay quality. Mass spectrometric parameters, such as ionization modes (positive vs. negative) and ion source types (TurbolonSpray vs. APCI) were screened to find suitable conditions for sensitive analysis of the compounds. Matrix suppression effects were taken into consideration. Calibration samples were prepared in plasma only, whereas quality control samples were prepared in both plasma and tissues to save animals and time. Accuracy and precision were in the range of 84.4-119.1% and 1-16.5%, respectively. Limits of quantification were in the range of 0.5-2.5 ng/ml for plasma and 2-10 ng/ml for tissues. Run times as short as 2.2 min could be achieved.
Heinig
Katja
K
F. Hoffmann-La Roche Ltd, Pharmaceuticals Division, Non-clinical Drug Safety, Basle, Switzerland. katja.heinig@roche.com
Bucheli
Franz
F
D000818:Animals; D002851:Chromatography, High Pressure Liquid / Q000379:methods*; D013058:Mass Spectrometry / Q000379:methods*; D004364:Pharmaceutical Preparations / Q000378:metabolism*; D010599:Pharmacokinetics; D051381:Rats; D012015:Reference Standards; D015203:Reproducibility of Results; D012680:Sensitivity and Specificity; D014018:Tissue Distribution
D004364:Pharmaceutical Preparations