The synthetic peptide [Dmt1]DALDA (Dmt-D-Arg-Phe-Lys-NH2; Dmt = 2′,6′-dimethyltyrosine; ‘super-DALDA’) is a mu opioid-receptor agonist. On-line liquid chromatography/quadrupole time-of-flight mass spectrometry and the corresponding stable isotope-incorporated synthetic peptide internal standard were used to quantify [Dmt1]DALDA that had been extracted from ovine plasma samples. The [M+2H]2+ ion was used to construct the calibration curve, and the product ion was used for verification of the peptide. The detection sensitivity for the [Dmt1]DALDA [M+2H]2+ ion was 12.5 fmol and 50 fmol for the m/z 432.3 product ion. The concentration profile of [Dmt1]DALDA was determined from a set of ovine plasma samples. The molecular specificity of the peptide quantification was confirmed by tandem mass spectrometry (MS/MS). Copyright © 2003 John Wiley & Sons, Ltd.