SummaryA high-performance liquid chromatography method has been developed for simultaneous determination of doxapram and its metabolites including keto-doxapram, the main and only active metabolite. The aim of the study was to evaluate this microtechnique and to report the cases involving severe adverse effects to determine toxic plasma levels in neonates. The method was found to be selective, and showed a good baseline separation of doxapram and metabolites. Recovery, linearity, intraday/interday precision, and limit of detection determined in aqueous solutions and in spiked plasma were satisfactory. The assay is simple, rapid, and plasma-sparing, which represents a true advantage in managing neonates. Case analysis was performed in two consecutive periods: 124 preterm infants in the first period and 173 in the second period. Severe toxic effects were observed in 4 cases in the first period, with doxapram plus keto-doxapram levels 9 mg/L. In the second period, only one case was observed. High-range plasma concentrations were significantly less frequent in the second period than in the first one. The authors conclude that measuring doxapram plus keto-doxapram in plasma may be of interest to avoid severe toxic effects in preterm neonates treated with doxapram.