TY - JOUR
T1 - Quantitative determination of the enantiomers of methadone and its metabolite (EDDP) in human saliva by enantioselective liquid chromatography with mass spectrometric detection
AU - Rodriguez Rosas, Maria Esther
AU - Preston, Kenzie L.
AU - Epstein, David H.
AU - Moolchan, Eric T.
AU - Wainer, Irving W.
PY - 2003/11/5
Y1 - 2003/11/5
N2 - A sensitive enantioselective liquid chromatographic assay with mass spectrometric detection (LC-MS) has been developed and validated for the simultaneous determination of saliva concentrations of (R)- and (S)-methadone (Met) and (R)- and (S)-2-ethylidene-1,5-dimethyl-3,3-diphenyl-pyrrolidine (EDDP, a primary metabolite of Met). Saliva specimens were collected using Salivette devices (Sarsedt), and centrifuged; collected saliva was then spiked with deuterated internal standards, D3-Met and D3-EDDP, and directly injected into the LC-MS. Enantioselective separations were achieved on a liquid chromatographic chiral stationary phase (CSP) based upon immobilized α1-acid glycoprotein (AGP) using a mobile phase composed of acetonitrile: ammonium acetate buffer (10mM, pH 7.0) in a ratio of 18:82 (v/v), a flow rate of 0.9ml/min and a temperature of 25°C. Under these conditions, enantioselective separations were observed for methadone (α=1.30) and EDDP (α=1.17) within 15min. Met, EDDP, D3-Met and D3-EDDP were detected using selected ion monitoring at m/z 310.20, 278.20, 313.20 and 281.20, respectively. Linear relationships between peak height ratio and drug-enantiomer concentrations were obtained for methadone in the range of 5.0-600.0ng/ml, and for EDDP from 0.5 to 15.0ng/ml per enantiomer with correlation coefficients better than 0.9994, where lower limit of quantification (LLOQ) for Met was 5ng/ml and for EDDP 0.5ng/ml. Acceptable intra- and inter-day precision of the method (CVs<4.0%) and accuracy (CVs<4.0%) were obtained. These findings demonstrate the accuracy and precision of the method used to successfully analyze saliva obtained from patients enrolled in a methadone-maintenance program.
AB - A sensitive enantioselective liquid chromatographic assay with mass spectrometric detection (LC-MS) has been developed and validated for the simultaneous determination of saliva concentrations of (R)- and (S)-methadone (Met) and (R)- and (S)-2-ethylidene-1,5-dimethyl-3,3-diphenyl-pyrrolidine (EDDP, a primary metabolite of Met). Saliva specimens were collected using Salivette devices (Sarsedt), and centrifuged; collected saliva was then spiked with deuterated internal standards, D3-Met and D3-EDDP, and directly injected into the LC-MS. Enantioselective separations were achieved on a liquid chromatographic chiral stationary phase (CSP) based upon immobilized α1-acid glycoprotein (AGP) using a mobile phase composed of acetonitrile: ammonium acetate buffer (10mM, pH 7.0) in a ratio of 18:82 (v/v), a flow rate of 0.9ml/min and a temperature of 25°C. Under these conditions, enantioselective separations were observed for methadone (α=1.30) and EDDP (α=1.17) within 15min. Met, EDDP, D3-Met and D3-EDDP were detected using selected ion monitoring at m/z 310.20, 278.20, 313.20 and 281.20, respectively. Linear relationships between peak height ratio and drug-enantiomer concentrations were obtained for methadone in the range of 5.0-600.0ng/ml, and for EDDP from 0.5 to 15.0ng/ml per enantiomer with correlation coefficients better than 0.9994, where lower limit of quantification (LLOQ) for Met was 5ng/ml and for EDDP 0.5ng/ml. Acceptable intra- and inter-day precision of the method (CVs<4.0%) and accuracy (CVs<4.0%) were obtained. These findings demonstrate the accuracy and precision of the method used to successfully analyze saliva obtained from patients enrolled in a methadone-maintenance program.
KW - 2-Ethylene-1,5-dimethyl-3,3-diphenyl-pyrrolidine
KW - Chiral stationary phases, LC
KW - Enantiomer separation
KW - Methadone
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UR - http://www.scopus.com/inward/citedby.url?scp=0142227094&partnerID=8YFLogxK
U2 - 10.1016/j.jchromb.2003.08.017
DO - 10.1016/j.jchromb.2003.08.017
M3 - Article
C2 - 14581075
AN - SCOPUS:0142227094
SN - 1570-0232
VL - 796
SP - 355
EP - 370
JO - Journal of Chromatography B: Analytical Technologies in the Biomedical and Life Sciences
JF - Journal of Chromatography B: Analytical Technologies in the Biomedical and Life Sciences
IS - 2
ER -