LC–MS/MS analysis of dextromethorphan metabolism in human saliva and urine to determine CYP2D6 phenotype and individual variability in N-demethylation and glucuronidation

In order to establish a fast screening method for the determination of the CYP2D6 metabolic phenotype a sensitive LC–MS/MS assay to quantify dextromethorphan (DEX) and its O-demethylated metabolite dextrorphan (DOR) in human saliva was developed with limits of quantitation of 1 pmol/ml. Saliva was provided by 170 medical students 2 h after oral ingestion of 30 mg (81 μmol) dextromethorphan hydrobromide. Individual ratios of the concentrations DEX/DOR (metabolic ratio, MRDEX/DOR) varied more than 25,000-fold (0.03–780). Two groups comprising 156 ‘Extensive’ and 14 ‘Poor Metabolizers’ were clearly distinguished. For the investigation of individual differences in N-demethylation and glucuronidation, four additional metabolites of DEX, 3-methoxymorphinan (MOM), 3-hydroxymorphinan (HOM), and the two O-glucuronides (DORGlu and HOMGlu) were measured by LC–MS/MS analysis of 6-h urine of 24 volunteers. The N-demethylation reactions DEX-to-MOM and DOR-to-HOM defined by the respective MR were significantly correlated. The same holds for the glucuronidation pathways (MRDOR/DORGlu versus MRHOM/HOMGlu). The three poor CYP2D6 metabolizers excreted relatively high amounts of the parent compound DEX (up to 7 μmol), but only low amounts of glucuronides (DORGlu: 0.4–1.0 μmol; HOMGlu: 0.2–0.7 μmol). For the 21 ‘Extensive Metabolizers’, the two glucuronides were the most abundant, with relatively little interindividual variation (DORGlu: 10–44 μmol; HOMGlu: 5–17 μmol). For the excretion of the glucuronides, two normal distributions provided the best fit, indicating that the determination of the glucuronides alone could allow assignment of the CYP2D6 metabolic phenotype.