On-line preconcentration microliquid chromatography tandem mass spectrometric method for bradykinin analysis in plasma

The practical limitations of miniaturized liquid chromatography (LC) method s for the analysis of endogenous peptides in plasma samples are studied. An automatic column switching method is used for on-line micropreconcentratio n and microLC-MS-MS (mass spectrometry) analysis of bradykinin and its meta bolites in plasma. Both preconcentration (PC) and analytical columns were f illed with C18 reversed phase due to the failure of anion exchange precolum ns to retain bradykinins. The detection limit for bradykinin in water was 1 0 pmol/L (2 fmols on column) and a good linearity was found in the range 5- 2000 fmols. In plasma, detection limit was 50 pmol/L (25 muL injected) with a linear response in the 100-2000 pmol/L range. Important rules for stable work were: (1) Cleaning the PC column after every run with pure acetonitri le (ACN) to eliminate organic matrix components; (2) to trap the more apola r organic matrix components on the analytical column to prevent their eluti on to the MS entrance; and (3) to set the electrospray needle to an off-axi s position to prevent accumulation of contaminants in the MS entrance. By u sing this setup, other procedures to reduce the organic load to the NIS ent rance such as the heart-cutting of the peaks eluting from the LC column wer e not required. (C) 2001 John Wiley & Sons, Inc.