Quantitative determination of tolazoline in human serum by high performance liquid chromatography

A micro high performance liquid chromatography assay is reported for the measurement of tolazoline in newborn infants. Pharmacokinetic data are presented for a single infant receiving tolazoline therapy. Tolazoline and the internal standard, clonidine, are extracted from alkaline serum into butylchloride/isopropanol (95/5). The organic layer is then back extracted with 50 mM phosphoric acid. A portion of the phosphoric acid layer is injected onto a 15-cm CN-bonded phase column. A mobile phase consisting of acetonitrile and phosphate buffer (pH 3) is used to elute tolazoline and the internal standard in less than 5 min. The effluent is monitored with a fixed wavelength detector at 214 nm. Using 50 microliters of serum, concentrations as low as 0.25 mg/L can be routinely determined with a coefficient of variation (CV) of 7.2%. However, when a 100-microliters sample is used, and the injection volume increased, a concentration of 0.05 mg/L can be routinely monitored with a CV of 1.2%. Interference from other drugs that are often used concurrently with tolazoline therapy was not observed.