Quantitative determination of peptide drug in human plasma samples at low pg/ml levels using coupled column liquid...

Plasma concentrations after administration of peptide drugs are often low due to the high potency often seen with this class of compounds. In this work a bioanalytical method based on coupled column liquid chromatography-tandem mass spectrometry (LC-MS/MS) is presented for quantification of a peptide drug, FE 202158, under clinical development. A volume of 0.5 ml human plasma is solid phase extracted on a weak cationic exchanger. After evaporation of the solvent to dryness, the reconstituted sample is injected into a coupled column liquid chromatography system. A heart-cut from the initial column, a cyano column, is trapped on a C(4) column and thereafter injected into a microbore C(18) column. For the detection a triple quadrupole mass spectrometer, equipped with a TurboIonSpray interface working in positive ion mode, is used. The design of the system is described and the gain in sensitivity and selectivity, compared to a conventional system, is discussed. Data from validation of the bioanalytical method are presented. For human plasma samples a lower limit of quantification (LLOQ) of 5.00 pg/ml (=4.77 pmol/l) was achieved. The inter-assay precision was less than 11% and bias was within +/-4% over the whole validated range of 5.00-860 pg/ml.