HPLC determination of dextromethorphan and its metabolites in urine

An HPLC method for determination of dextromethorpan (DM), dextrorphan (DEX), methoxymorphinan (MM) and hy droxymorphinan (HM) in urine with fluorescence detection (excitation and emission wavelength 280 and 310 nm) was elaborated. The mobile phase was acetonitrile - 0.01 M-KH2PO4 3:2, triethylamine 350 mu l l(-1), pH 3.6; flow rate 0.7 ml.min(-1) internal standard betaxolol (BX). For isolation of the substances from urine, liquid-liquid extraction and re-extraction with freezing out the aqueous phase in all extraction steps was used. The extraction yields were 97, 99, 83, 92 and 101 % for DM, DEX, HM, MM and EX, respectively. The method was used for determination of phenotype of cytochrome P 450 of isoenzyme 2D6 in a set of 102 healthy, unrelated volunteers from Czech population. In dependence on molar concentration ratios of DM and DEX in urine (metabolic ratio, MP), seven volunteers (6.9 %) were classified as slow and 95 (93.1 %) as rapid metabolizers of DM (MP < 0.3). The median in the former group was 1.3 (0.37-4.6) and 0.007 (0.0001-0.18) in the latter.