Method For Determination Of Methadone Research Articles

Ultrasound-assisted liquid-liquid extraction for the determination of methadone in urine samples using gas chromatography-mass spectrometry

Background: Methadone abuse and dependence has been reported as a growing concern in some countries. In present study, a rapid, simple, easy and sensitive method for determination of methadone in human urine samples for use in clinical and forensic toxicology and drug screening laboratories was developed and validated.
 Methods: We determined methadone in urine sampled by gas chromatography/mass spectrometry (GC/MS) and used ultrasound assisted liquid-liquid extraction (UALLE) method for the extraction and preconcentration of methadone before analysis.
 Results: The limit of detection was 2.1 ng/mL and the limit of quantification 7 ng/mL. Correlation coefficient was 0.9984 for the methadone calibration curve in linear range from 7 to 10,000 ng/mL. The method is accurate and precise. Recovery was in the range of 81.3 % to 97.4 % and enrichment factor was 8.7. The method was successfully applied for determining methadone in clinical and postmortem urine samples.
 Conclusions: The present method is a rapid, simple, easy and sensitive procedure and can be used in clinical and forensic toxicology laboratories as routine method for qualitative and quantitative analysis of methadone.

A Novel Method for Determination of Methadone in the Serum by High-Performance Liquid Chromatography with Electrochemical Detection.

In March 2013, the clinical use of oral methadone tablets was initiated in Japan. There are many factors responsible for the change in blood concentrations of methadone, and its pharmacokinetics is very complex. Therefore, a simple and accurate measurement method for methadone blood concentrations was developed using HPLC/electrochemical detector (ECD). An eluent of 10 mM Na2HPO4/CH3CN/CH3OH (20 : 19 : 3) was used as the mobile phase. The column was used the XTerra® RP18, and the voltage of the ECD was set at 400 to 800 mV. As a result, the calibration curve was linear in the ranges of 10 to 100 ng/mL (y=5012.7x+1041.1, r=0.999). The intra- and inter-day coefficients of variation were <5.2 and <5.8%, respectively. Therefore, this method was considered to be useful for the measurement of methadone blood levels in cancer patients. Also, using this method, blood methadone concentration was measured over time in a patient with cancer-associated pain who was treated with methadone. The estimated clearance (CL/F) and distribution volume (Vd/F) of methadone were 2.84 L/h and 502.8 L, respectively, and took about two weeks to reach steady state.

Determination of Methadone in Exhaled Breath Condensate by Liquid Chromatography-Tandem Mass Spectrometry

Within the field of toxicology exhaled breath is used as specimen only for determination of alcohol. However, it was recently discovered that when using sensitive liquid chromatography-mass spectrometry (LC-MS) technique, amphetamine, methamphetamine, and methadone are detectable in exhaled breath following intake by drug addicts. We therefore undertook to develop a method for determination of methadone in exhaled breath condensate from patients undergoing methadonemaintenance treatment. Exhaled breath condensate was collected from 14 patients after intake of the daily methadone dose. The exhaled breath condensate was collected for 10 min using an Ecoscreen instrument. After extraction of any trapped methadone from the condensate by solid-phase extraction, the final extract was analyzed by a combined LC-MS-MS method. Recovery of methadone from breath condensate in the solid-phase extraction was 104%, no significant matrix effects were observed, and the quantification using methadone-d(3) as internal standard was accurate (10% bias) and precise (coefficient of variation 6.2%). Methadone was indisputably identified by means of the MS technique in exhaled breath condensate from all 14 patients. Identification was based on monitoring two product ions in selected reaction monitoring mode with correct relative ratio (± 20%) and correct retention time. Excretion rates ranged from 23.6 to 275 pg/min. No methadone was detected in five control subjects (< 2 pg/min). This finding confirms that methadone is present in exhaled breath from patients in methadone treatment. Collection of exhaled breath specimen is likely to be complementary to other matrices presently in use in testing for drugs-of-abuse.

Method for determination of methadone in exhaled breath collected from subjects undergoing methadone maintenance treatment

At present drugs of abuse testing using exhaled breath as specimen is only possible for alcohol. However, we recently discovered that using modern liquid chromatography–mass spectrometry technique amphetamine and methamphetamine is detectable in exhaled breath following intake in drug addicts. We therefore undertook to develop a method for determination of methadone in exhaled breath from patients undergoing methadone maintenance treatment. Exhaled breath was collected from 13 patients after intake of the daily methadone dose. The compounds were trapped by filtering the air through a C18 modified silica surface. After elution of any trapped methadone the extract was analysed by a combined liquid chromatography–tandem mass spectrometry method. Recovery of trapped methadone from the filter surface was 96%, no significant matrix effect was observed, and the quantification using methadone-d3 as an internal standard was accurate (<10% bias) and precise (coefficient of variation 1.6–2.0%). Methadone was indisputably identified by means of the mass spectrometry technique in exhaled breath samples from all 13 patients. Identification was based on monitoring two product ions in selected reaction monitoring mode with correct relative ratio (±20%) and correct retention time. Excretion rates ranged from 0.39 to 78 ng/min. No methadone was detected in 10 control subjects. This finding confirms that breath testing is a new possibility for drugs of abuse testing. Collection of exhaled breath specimen is likely to be more convenient and safe as compared to other matrices presently in use.

Validated method for the simultaneous determination of methadone and its main metabolites (EDDP and EMDP) in plasma of umbilical cord blood by gas chromatography–mass spectrometry

A sensitive and specific GC/MS method for the determination of methadone (MDN) and its two main metabolites, 2-ethylidene-1,5-dimethyl-3,3-diphenylpyrrolidine (EDDP) and 2-ethyl-5-methyl-3,3-diphenylpyrroline (EMDP), in plasma samples obtained from venous and arterial umbilical cord blood and maternal blood has been developed, optimized and validated. Specimen preparation includes protein precipitation with acetonitrile and simultaneous solid-phase extraction of the three analytes. Methadone-d9 was used as internal standard for the determination of MDN and EMDP, while EDDP-d3 for EDDP. Limits of detection were 0.6 μg/L for MDN and 0.3 μg/L for EDDP and EMDP, while limits of quantification were 2.0 μg/L for MDN and 1.0 μg/L for EDDP and EMDP. The calibration curves were linear up to 2000 μg/L for MDN and up to 1000 μg/L for EDDP and EMDP. Absolute recovery ranged from 94.8 to 99.7% for all three analytes. Intra- and interday accuracy was less than 5.3 and 5.5%, respectively, while intra- and interday precision was less than 3.5 and 5.0%, correspondingly, for all analytes. The method proved suitable for the determination of MDN and its two main metabolites in plasma samples obtained from umbilical cord and maternal blood of a woman participating in a MDN maintenance program, during the prenatal and postpartum period.

Enantioselective high-performance liquid chromatographic method for the determination of methadone and its main metabolite in urine using an AGP and a C 8 column coupled serially

A simple and sensitive method for the enantioselective high-performance liquid chromatographic determination of methadone and its main metabolite, EDDP, in human urine is described. (−)-( R)-Methadone, (+)-( S)-methadone, (+)-( R)-EDDP, (−)-( S)-EDDP and imipramine as an internal standard are detected by ultraviolet detection at 200 nm. The enantiomers of methadone and EDDP were extracted from human urine by a simple liquid–liquid extraction procedure. The extracted sample was reconstructed in mobile phase and the enantiomers of methadone and EDDP were quantitatively separated by HPLC on a short analytical LiChrospher RP8 column coupled in series with a chiral AGP column. Determination of all four enantiomers was possible in the range of 0.03 to 2.5 μ M. The recoveries of methadone enantiomers and EDDP enantiomers added to human urine were about 90% and 80%, respectively. The method was applicable for determination of methadone enantiomers and the enantiomers of its main metabolite in urine samples from methadone maintenance patients and patients suffering from severe chronic pain.

Stereospecific gas chromatographic method for determination of methadone in serum.

rac-Methadone is used clinically for the chronic maintenance treatment of heroin addiction and for the relief of pain. As the pharmacological activity of methadone is due primarily to the (-)-(R)-enantiomer, stereospecific measurements of methadone serum concentrations in methadone-treated patients are expected to be more relevant for clinical studies than earlier described total drug measurements. This study describes a stereospecific gas chromatographic (GC) method for the determination of methadone in serum. The extracted methadone was derivatizised with (-)-menthyl chloroformate. The diastereometric derivatives were analysed by GC on a capillary column and detected with a nitrogen-phosphorus detector. The resolution factor obtained for the methadone enantiomers was 1.1 with a relatively short time of analysis (30 min). By analysing the pure (-)-(R)-enantiomer, no racemization was seen during the analysis. The lower limit of quantitation was 75 nmol/l for each enantiomer. Measurements of the ratio between (-)-(R)- and (+)-(S)-methadone concentrations in serum from five methadone-treated patients showed interindividual differences (range 0.5-1.1). The patient results correlated well with those from another GC method measuring total methadone.