Measurement of naloxone in plasma using high-performance liquid chromatography with electrochemical detection

A new sample preparation technique, solid-phase extraction employing the ion-pair reagent octyl sodium sulfate (OSS), was developed for the selective isolation of nanogram quantities of naloxone from human plasma. Plasma samples containing naloxone, OSS, buffer, and naltrexone (internal standard) were applied to octyldecylsilane (C18) cartridges, and the opiates were eluted with 100% methanol. An extraction efficiency of 90% was achieved. Extracts were then examined by reversed-phase high-performance liquid chromatography with electrochemical detection, with mobile phase composition adjusted to optimize separation and quantitation. An oxidizing potential of +0.93 V resulted in an assay sensitivity of 1.0 ng. Thus, plasma naloxone levels of ca. 20 ng/ml could be readily detected and quantified. The intra-assay coefficient of variation at plasma levels of 80 ng/ml was 7.2%. Plasma naloxone values obtained during a 5-h infusion of the drug (0.24 mg/min) in two normal human subjects were stable and reproducible. Thus, the method herein described is applicable to human studies employing naloxone infusion.