Adenosine and dopamine simultaneously determined in urine by reversed-phase HPLC, with on-line measurement of ultraviolet absorbance and electrochemical detection

Abstract

This assay method allows the simultaneous determination of adenosine and dopamine in 0.5 mL of human urine within ~35 min, by using isocratic, reversed-phase, high-performance liquid chromatography coupled with an on-line detector for ultraviolet absorbance and electrochemical activity. The analytes were rapidly separated on an affinity column packed with phenylboronate-bonded silica. Mean recoveries for adenosine, 2-chloroadenosine (internal standard), dopamine, and (±)-isoproterenol (also an internal standard) were 91%, 99%, 88%, and 104%, respectively. The calibration curves for adenosine and dopamine were linear (r = 0.999, P<0.01) over the respective concentration ranges of 0.10 to 2.00 mg/L and 0.05 to 1.00 mg/L. Analytical precisions, assessed by the within-day and the day-to-day CVs, were 4.5% and 6.5% for adenosine, and 5.1% and 6.6% for dopamine, respectively. The mean (± SD) concentrations of adenosine (1.27 ± 0.73 mg/L) and dopamine concentrations (0.38 ± 0.25 mg/L) measured in urine from 19 healthy subjects agreed well with those previously reported.