Determination of bradykinin and arg-bradykinin in rat muscle tissue by microdialysis and capillary column-switching liquid chromatography with mass spectrometric detection.
Quantification of bradykinin peptides in limited amounts of rat muscle tissue dialysate has been performed using a packed capillary LC-ESI-TOF-MS method. The micro dialysate samples (450 microL) with added internal standard were loaded onto a 1 mm x 5 mm loading column packed with 5 microm Kromasil C18 particles by a carrier solution of 0.1% formic acid in ACN/water (5:95, v/v) at a flow rate of 250 microL/min for online preconcentration of the analytes. Back-flushed elution onto a 150 mm x 0.5 mm Zorbax C18 column packed with 5 microm particles was conducted using a linear solvent ACN/H2O gradient containing 0.1% formic acid. (Tyr8)-bradykinin was used as an internal standard and was added to the dialysis sample prior to injection. Baseline separation of bradykinin, arg-bradykinin and (tyr8)-bradykinin was achieved within 10 min. Positive ESI was performed in the m/z range of 200-1300. The method was validated in the range 0.2-1.0 ng/mL dialysate, yielding correlation coefficients of 0.995 and 0.990 for bradykinin and arg-bradykinin, respectively. The within-assay and between-assay precisions were between 4.3-9.6% and 6.2-10.6%, respectively. Both arg-bradykinin and bradykinin were detected in dialysate from rat muscle tissue, at concentrations of 0.1 and 0.4 ng/mL for bradykinin and arg-bradykinin, respectively, confirming the presence of arg-bradykinin in rat muscles.
Wilson
Steven Ray
SR
Department of Chemistry, University of Oslo, Oslo, Norway. stevenw@kjemi.uio.no
Boix
Fernando
F
Holm
Anders
A
Molander
Pål
P
Lundanes
Elsa
E
Greibrokk
Tyge
T
D000818:Animals; D001920:Bradykinin / Q000031:analogs & derivatives* / Q000032:analysis* / Q000737:chemistry / Q000302:isolation & purification; D002197:Capillary Action; D002853:Chromatography, Liquid / Q000295:instrumentation / Q000379:methods; D017551:Microdialysis; D018482:Muscle, Skeletal / Q000737:chemistry*; D051381:Rats; D021241:Spectrometry, Mass, Electrospray Ionization / Q000295:instrumentation / Q000379:methods
D001920:Bradykinin