Simultaneous Determination of Hydrocodone, and Its Two Metabolites in Human Plasma by HPLC–MS–MS

Abstract

A rapid, sensitive and specific assay method has been developed to simultaneously determine human plasma concentrations of hydrocodone and its metabolites, norhydrocodone, hydromorphone, using high-performance liquid chromatography with an electrospray ionization (ESI) tandem mass spectrometry (HPLC-MS-MS). Hydrocodone, its metabolites, and internal standard, hydrocodone-d 3 , norhydrocodone-d 3 , hydromorphone-d 3 , were separated from human plasma using solid-phase extraction (Empore MPC-SD Solid Phase Extraction Disk). The eluate was dried, reconstituted and injected into the LC-MS-MS system. Chromatographic separation was performed on a Kromasil 100-5SIL-Dimensions C 18 column (100 9 2.1 mm, 5.0 lm, Thermo Hypersil-Keystone, USA) using a gradient mobile phase with 20 mmol L -1 ammonium formate in water with 0.2% formic acid and 0.1% formic acid in acetonitrile. Detection and quantitation were performed by MS/MS using electrospray ionization and multiple reactions monitoring in the positive ion mode. The calibration curves were linear over the concentration ranges 0.05-50 ng mL -1 for hydrocodone (r 2 = 0.9991) and norhydrocodone (r 2 = 0.9990), and 0.01-10 ng mL -1 for hydromorphone (r 2 = 0.9990). The limit of quantification was 0.05 ng mL -1 for hydrocodone and norhydrocodone, and 0.01 ng mL -1 for hydromorphone. The extraction recovery was above 64.36, 68.51 and 71.78% for hydrocodone, norhydrocodone and hydromorphone. The accuracy was higher than 99.06, 97.70 and 100.07% for hydrocodone, norhydrocodone and hydromorphone. The intra-and inter-day precisions were \5.80, 5.90 and 3.02% for hydrocodone, norhydrocodone and hydromorphone. The method was accurate, sensitive and simple and was successfully applied to a pharmacokinetic study after a single oral administration of hydrocodone bitartrate at a dose of 5 mg in 12 healthy Chinese volunteers.

In Vivo Activity of Norhydrocodone: An Active Metabolite of Hydrocodone

“…To date, the in vivo effects of norhydrocodone are not well documented. If norhydrocodone is active in vivo, it may contribute to the pharmacodynamic profile of hydrocodone since plasma concentrations of norhydrocodone in humans have been reported to be greater than that of hydromorphone (Hao et al, 2011). Furthermore, norhydrocodone is detected in urine for significantly longer than hydromorphone and hydrocodone (Valtier and Bebarta, 2012).…”

“…It is possible that the unpleasant effects might be a precursor to frank neuroexcitatory actions, although this is not known. Nevertheless, following acute oral hydrocodone, the plasma and urine levels of norhydrocodone invariably exceed that of the hydromorphone and sometimes that of hydrocodone (Hao et al, 2011;Valtier and Bebarta, 2012). Chronic hydrocodone administration might result in substantial levels of the nor-metabolite that may mediate unpleasant subjective effects and perhaps incipient neuroexcitatory action.…”

“…The reported elimination half-life for hydrocodone following oral administration in humans is 3-9 hours (Otton et al, 1993;Hao et al, 2011) and for hydromorphone following intravenous and oral administration in humans, the t 1/2 is 2-4 hours (Vallner et al, 1981;Parab et al, 1988). The reported half-life for norhydrocodone following oral hydrocodone administration is 8 hours (Hao et al, 2011).…”

“…The reported half-life for norhydrocodone following oral hydrocodone administration is 8 hours (Hao et al, 2011). Studies analyzing human plasma and urine following oral hydrocodone indicated that norhydrocodone concentration exceeds that of hydromorphone, and in some cases, that of the parent compound (Hao et al, 2011;Valtier and Bebarta, 2012).…”

“…The reported half-life for norhydrocodone following oral hydrocodone administration is 8 hours (Hao et al, 2011). Studies analyzing human plasma and urine following oral hydrocodone indicated that norhydrocodone concentration exceeds that of hydromorphone, and in some cases, that of the parent compound (Hao et al, 2011;Valtier and Bebarta, 2012). In addition, the examination of urine specimens revealed that norhydrocodone is detected for a significantly longer period compared to both hydrocodone and hydromorphone (Valtier and Bebarta, 2012).…”