A simple high-performance liquid chromatography assay for simultaneous determination of plasma norepinephrine, epinephrine, dopamine and 3,4-dihydroxyphenyl acetic acid.
A reversed-phase HPLC assay coupled with electrochemical detection for simultaneously measuring plasma levels of norepinephrine, epinephrine, dopamine, and 3,4-dihydroxyphenylacetic acid (DOPAC) was developed. Separation of the catecholamines and the internal standard isoproterenol was obtained by a mobile phase consisting of 7% methanol in 0.1 M citrate buffer containing 0.3 mM sodium ethylenediaminetetraacetic acid (EDTA), and 0.5 mM 1-octanesulfonic acid, operated under isocratic condition at a flow rate of 1.2 ml/min. The potential of the guard cell was set at +650 mV, the first electrode of the analytical cell at +100 mV and the second at + 350 mV. Using a signal-to-noise ratio of > 3, the minimum detection limit assessed by direct on column injection was < 10 pg for analyte. The assays were linear from basal concentrations to 400 ng/ml. The intra- and inter-assay variations were < 10 and 15%, respectively. The assay has been applied successfully to measure plasma concentrations of these catecholamines in humans, rabbits and rats.
Wang
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Pharmacokinetics and Metabolism Laboratory, College of Pharmacy, Burbridge Building, Dalhousie University, Halifax, Canada.
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D015102:3,4-Dihydroxyphenylacetic Acid / Q000097:blood / Q000302:isolation & purification; D000818:Animals; D002395:Catecholamines / Q000097:blood* / Q000302:isolation & purification; D002851:Chromatography, High Pressure Liquid / Q000379:methods*; D004298:Dopamine / Q000097:blood / Q000302:isolation & purification; D004837:Epinephrine / Q000097:blood / Q000302:isolation & purification; D006801:Humans; D009638:Norepinephrine / Q000097:blood / Q000302:isolation & purification; D011786:Quality Control; D011817:Rabbits; D051381:Rats; D015203:Reproducibility of Results
D002395:Catecholamines; D015102:3,4-Dihydroxyphenylacetic Acid; D004298:Dopamine; D009638:Norepinephrine; D004837:Epinephrine