Determination of naltrexone and 6-beta-naltrexol in plasma by solid-phase extraction and gas chromatography-negative ion chemical ionization mass spectrometry

Solid-phase extraction (SPE) and a one-step derivatization are combined with gas chromatography-negative ion chemical ionization-mass spectrometry to simplify a previously reported method for the determination of naltrexone and its metabolite, 6-β-naltrexol, in human plasma. Deuterated isotopomers of naltrexone and 6-β-naltrexol are used as internal standards. After SPE, the extracts are derivatized with pentafluoropropionic anhydride at room temperature to form predominantly the bis-pentafluoropropionyl derivative of naltrexone and the tris-pentafluoropropionyl derivative of 6-β-naltrexol. The derivatized extracts are analyzed by monitoring ion currents at m/z 633 (naltrexone), m/z 636 (naltrexone-2H3), m/z 633 6-β-naltrexol), and m/z 640 (6-β-naltrexol-2H7). Control plasma samples containing 0.3, 3, or 30 ng/mL of each analyte were analyzed for precision and accuracy with the following results: intra-assay, the percentage of target concentrations were 107-113% for naltrexone and 107-120% for 6-β-naltrexol, and the coefficients of variation (CVs) were 3.1-6.3% for naltrexone and 3.1-5.7% for 6-β- naltrexol; inter-assay, the percentage of target concentrations were 103- 110% for naltrexone and 110-113% for 6-β-naltrexol, and the CVs were 6.1- 9.1% for naltrexone and 5.9-9.1% for 6-β-naltrexol. At the limit of quantitation (LOQ) of 0.1 ng/mL, both analytes quantitated within 20% of the target concentration with CVs less than 17%. The extraction recoveries determined at 0.3 and 30 ng/mL were 79 and 80% for naltrexone and 76 and 75% for 6-β-naltrexol. Bench-top stability tested with concentrations of 0.3 and 3.0 ng/mL did not decrease more than 10% from the zero-hour controls at 3, 6, and 24 h. Selectivity was determined using plasma from six donors and none showed interfering peaks greater than 22% of the LOQ for naltrexone and 53% of the LOQ for 6-β-naltrexol. Using this method, naltrexone and 6-β- naltrexol were readily detected in plasma specimens collected 5.5 h after oral doses of 25 or 100 mg naltrexone. Following discontinuation of treatment, naltrexone was detected 30 h after the 100-mg dose, whereas 6-β- naltrexol was detected 125 h after both the 25- and 100-mg doses.