Methadone Enantiomers Research Articles

In vitro–in silico-based prediction of inter-individual and inter-ethnic variations in the dose-dependent cardiotoxicity of R- and S-methadone in humans

New approach methodologies predicting human cardiotoxicity are of interest to support or even replace in vivo-based drug safety testing. The present study presents an in vitro–in silico approach to predict the effect of inter-individual and inter-ethnic kinetic variations in the cardiotoxicity of R- and S-methadone in the Caucasian and the Chinese population. In vitro cardiotoxicity data, and metabolic data obtained from two approaches, using either individual human liver microsomes or recombinant cytochrome P450 enzymes (rCYPs), were integrated with physiologically based kinetic (PBK) models and Monte Carlo simulations to predict inter-individual and inter-ethnic variations in methadone-induced cardiotoxicity. Chemical specific adjustment factors were defined and used to derive dose–response curves for the sensitive individuals. Our simulations indicated that Chinese are more sensitive towards methadone-induced cardiotoxicity with Margin of Safety values being generally two-fold lower than those for Caucasians for both methadone enantiomers. Individual PBK models using microsomes and PBK models using rCYPs combined with Monte Carlo simulations predicted similar inter-individual and inter-ethnic variations in methadone-induced cardiotoxicity. The present study illustrates how inter-individual and inter-ethnic variations in cardiotoxicity can be predicted by combining in vitro toxicity and metabolic data, PBK modelling and Monte Carlo simulations. The novel methodology can be used to enhance cardiac safety evaluations and risk assessment of chemicals.

Pharmacokinetic modeling of R and S‐Methadone and their metabolites to study the effects of various covariates in post‐operative children

Methadone is a synthetic opioid used as an analgesic and for the treatment of opioid abuse disorder. The analgesic dose in the pediatric population is not well‐defined. The pharmacokinetics (PKs) of methadone is highly variable due to the variability in alpha‐1 acid glycoprotein (AAG) and genotypic differences in drug‐metabolizing enzymes. Additionally, the R and S enantiomers of methadone have unique PK and pharmacodynamic properties. This study aims to describe the PKs of R and S methadone and its metabolite 2‐ethylidene‐1,5‐dimethyl‐3,3‐diphenylpyrrolidine (EDDP) in pediatric surgical patients and to identify sources of inter‐ and intra‐individual variability. Children aged 8–17.9 years undergoing orthopedic surgeries received intravenous methadone 0.1 mg/kg intra‐operatively followed by oral methadone 0.1 mg/kg postoperatively every 12 h. Pharmacokinetics of R and S methadone and EDDP were determined using liquid chromatography tandem mass spectrometry assays and the data were modeled using nonlinear mixed‐effects modeling in NONMEM. R and S methadone PKs were well‐described by two‐compartment disposition models with first‐order absorption and elimination. EDDP metabolites were described by one compartment disposition models with first order elimination. Clearance of both R and S methadone were allometrically scaled by bodyweight. CYP2B6 phenotype was a determinant of the clearance of both the enantiomers in an additive gene model. The intronic CYP3A4 single‐nucleotide polymorphism (SNP) rs2246709 was associated with decreased clearance of R and S methadone. Concentrations of AAG and the SNP of AAG rs17650 independently increased the volume of distribution of both the enantiomers. The knowledge of these important covariates will aid in the optimal dosing of methadone in children.

Stereoselective methadone disposition after administration of racemic methadone to anesthetized Shetland ponies assessed by capillary electrophoresis.

The enantioselectivity of the pharmacokinetics of methadone was investigated in anesthetized Shetland ponies after a single intravenous (0.5 mg/kg methadone hydrochloride; n = 6) or constant rate infusion (0.25 mg/kg bolus followed by 0.25 mg/kg/h methadone hydrochloride; n = 3) administration of racemic methadone. Plasma concentrations of l‐methadone and d‐methadone and their major metabolites, l‐ and d‐2‐ethylidene‐1,5‐dimethyl‐3,3‐diphenylpyrrolidine (EDDP), respectively, were analyzed by CE with highly sulfated γ‐cyclodextrin as chiral selector and electrokinetic analyte injection from liquid/liquid extracts prepared at alkaline pH. In both trials, the d‐methadone concentrations were lower than those of l‐methadone and the d‐EDDP levels were lower than those of L‐EDDP. For the case of a single intravenous bolus injection, the plasma concentration versus time profile of methadone enantiomers was analyzed with a two‐compartment pharmacokinetic model. l‐methadone showed a slower elimination rate constant, a lower body clearance, and a smaller steady‐state volume of distribution than d‐methadone. d‐methadone and d‐EDDP were eliminated faster than their respective l‐enantiomers. This is the first study that outlines that the disposition of racemic methadone administered to anesthetized equines is enantioselective.

Enantiomeric methadone quantitation on real post-mortem dried matrix spots samples: Comparison of liquid chromatography and supercritical fluid chromatography coupled to mass spectrometry

This study describes two bioanalytical methods for the quantitation of the two methadone enantiomers in dried matrix spots using high performance liquid chromatography coupled to tandem mass spectrometry (HPLC-MS/MS) and high performance supercritical chromatography tandem mass spectrometry (HPSFC-MS/MS). Dried matrix spots were obtained by spotting 10 µL of each sample fluid on a Whatman paper. Methadone and its main metabolite, EDDP, were extracted with 100 µL methanol and subsequently injected into the LC-MS/MS and SFC-MS/MS systems. Enantiomeric separation was achieved with AGP-column for the LC conditions and with Chiralpak IH-3 in SFC. The two methods were fully validated and 93 post-mortem samples were analysed with both analytical methods. Results from validation parameters and results obtained for all post-mortem samples were compared with a significant spearman correlation of rs = 0.9978 for R-methadone and rs = 0.9981 for S-methadone. The LC method provided better results in terms of uncertainty, retention factor and resolution, whereas SFC provides better sensitivity, with lower LOD. Median R-/S-methadone ratio in peripheral blood was found equal to 1.60 (N = 32), varying from 0.79 to 4.23. The reported values were in good agreement with previously published results.Based on the results obtained here, SFC-MS/MS can be considered a reliable alternative to the widely used LC-MS/MS for the quantitation of methadone enantiomers in bioanalysis and should be evaluated for other bioanalytical methods. Both methods can be easily and quickly used in toxicological routine analysis for the methadone quantitation in human fluids matrices, even if considering that the polysaccharide coated column IH-3 used in SFC does not allow the enantiomeric EDDP separation.

Relevance of CYP2B6 and CYP2D6 genotypes to methadone pharmacokinetics and response in the OPAL study.

Our study aimed to evaluate the impacts of the cytochrome P450 (CYP) 2B6-G516T and CYP2D6 genetic polymorphisms on pharmacokinetic and clinical parameters in patients receiving methadone maintenance treatment. Opioid PhArmacoLogy (OPAL) was a clinical survey of the sociodemographic characteristics, history and consequences of pathology associated with methadone maintenance treatment response and current addictive comorbidities. A subgroup of 72 methadone patients was genotyped. When comparing the three CYP2B6 genotype groups, the methadone (R)- and (S)-methadone enantiomer concentrations/doses (concentrations relative to doses) were different (P = .029, P = .0019). The CYP2D6 phenotypes did not seem to be relevant with regard to methadone levels. On multivariate analysis, neither the CYP2B6 genotype nor the CYP2D6 phenotype explained the (R)-methadone concentration/dose values (P= .92; P = .86); the (S)-methadone concentration/dose values (P = .052; P = .95 [although there was a difference between the TT group and GT and GG groups {P = .019}]); or opiate cessation (P = .12; P = .90). The genotyping of CYP2B6 G516T could be an interesting tool to explore methadone intervariability.

Inverse cationic ITP for separation of methadone enantiomers with sulfated β-cyclodextrin as chiral selector.

Chiral ITP of the weak base methadone using inverse cationic configurations with H+ as leading component and multiple isomer sulfated β-CD (S-β-CD) as leading electrolyte (LE) additive, has been studied utilizing dynamic computer simulation, a calculation model based on steady-state values of the ITP zones, and capillary ITP. By varying the amount of acidic S-β-CD in the LE composed of 3-morpholino-2-hydroxypropanesulfonic acid and the chiral selector, and employing glycylglycine as terminating electrolyte (TE), inverse cationic ITP provides systems in which either both enantiomers, only the enantiomer with weaker complexation, or none of the two enantiomers form cationic ITP zones. For the configuration studied, the data reveal that only S-methadone migrates isotachophoretically when the S-β-CD concentration in the LE is between about 0.484 and 1.113mM. Under these conditions, R-methadone migrates zone electrophoretically in the TE. An S-β-CD concentration between about 0.070 and 0.484mM results in both S- and R-methadone forming ITP zones. With >1.113mM and < about 0.050mM of S-β-CD in the LE both enantiomers are migrating within the TE and LE, respectively. Chiral inverse cationic ITP with acidic S-β-CD in the LE is demonstrated to permit selective ITP trapping and concentration of the less interacting enantiomer of a weak base.

Simultaneous chiral separation of tramadol and methadone in tablets, human urine, and plasma by capillary electrophoresis using maltodextrin as the chiral selector

The stereoselective analysis and separation of racemic drugs play an important role in pharmaceutical industry to eliminate the unwanted isomer and find the right therapeutic control for the patient. Present study suggests a maltodextrin-modified capillary electrophoresis method for a single-run chiral separation of two closely similar opiate pain relief drugs: tramadol (TRA) and methadone (MET). The best separation method possible for the both enantiomers was achieved on an uncoated fused-silica capillary at 25°C using 100 mM phosphate buffer (pH8.0) containing 20% (wv-1 ) maltodextrin with dextrose equivalent of 4-7 and an applied voltage of 16kV. Under optimal conditions, the baseline resolution of TRA and MET enantiomers was obtained in less than 12minutes. The relative standard deviations (n=3) of 20μgmL-1 TRA and MET were 2.28% and 3.77%, respectively. The detection limits were found to be 2μgmL-1 for TRA and 1.5μgmL-1 for MET. This method was successfully applied to the measurement of drugs concentration in their tablets, urine, and plasma samples.

Can Saliva and Plasma Methadone Concentrations Be Used for Enantioselective Pharmacokinetic and Pharmacodynamic Studies in Patients With Advanced Cancer?

PurposeMethadone is a potent analgesic used to treat refractory cancer pain. It is administered as a racemic mixture, with the l-enantiomer being primarily a μ-receptor agonist, whereas the d-enantiomer is an N-methyl-d-aspartate antagonist and inhibits serotonin and norepinephrine reuptake. Dose requirements vary greatly among patients to achieve optimal pain control and to avoid the risk of adverse effects. The relationship between plasma and saliva methadone enantiomer concentrations was investigated to determine if saliva could be a substitute for plasma in pharmacodynamic and pharmacokinetic studies for clinical monitoring and dose optimization of methadone in patients with advanced cancer. MethodsPatients with advanced cancer who were prescribed varying doses of oral methadone for pain management were recruited to obtain paired plasma and saliva samples. Pain scores were recorded at the time of sampling. The total and unbound plasma and saliva concentrations of the l- and d-enantiomers of methadone were quantified by using an HPLC-MS/MS method. The relationship between plasma (total and unbound) and saliva concentrations were compared. The saliva-to-plasma concentration ratio was compared versus the dose administered and the time after dosing for both enantiomers. The association of methadone concentrations with reported pain scores was compared by using a Mann-Whitney U test for significance. FindingsFifty patients receiving a mean dose of 11mg/d of methadone provided 151 paired plasma and saliva samples. The median age of the population was 61 years with an interquartile range of 53-71 years with total body weight ranging from 59-88 kg. Median (interquartile) total plasma concentrations for l- and d-methadone were 50.78 ng/mL (30.6–113.0 ng/mL) and 62.0 ng/mL (28.7–116.0 ng/mL), respectively. Median (interquartile range) saliva concentrations for l- and d-methadone were 81.5 ng/mL (28.0–203.2 ng/mL) and 44.2 (16.2–149.7 ng/mL). No relationship could be established between plasma and saliva concentrations for l- and d-methadone (r2 = 0.35 and 0.25). The saliva-to-plasma concentration analyzed with the methadone dose showed higher saliva concentrations at lower doses. Dose-normalized saliva concentrations followed a similar pattern over time compared with plasma concentrations. No correlation was found between l-methadone plasma, d-methadone plasma, l-methadone saliva, d-methadone saliva concentrations, and pain score. ImplicationsSaliva concentration was not a better predictor of pain control than plasma concentration for dose optimization and monitoring studies of methadone in patients with cancer. Although the saliva-to-plasma ratio of the concentration of methadone enantiomers was stable across the dosing range, due to the variability in individual saliva-to-plasma ratios, saliva sampling may not be a valid substitute in pharmacokinetic studies of methadone in cancer.

Findings in the hair of drug abusers using pressurized liquid extraction and solid-phase extraction coupled in-line with capillary electrophoresis

A suitable method has been developed and validated for simultaneously determining cocaine and its major metabolite, benzoylecgonine, 6-acetylmorphine, codeine, morphine and the methadone enantiomers in human hair samples by the in-line coupling between SPE and CyD-assisted CE with a previous sample pretreatment procedure based on pressurized liquid extraction. Optimal separation was achieved on a fused silica-capillary of 50μm i.d. and 80cm total length using 11mM α-CyD in an aqueous solution of 80mM sodium phosphate at pH 2.5 as the separation medium and an applied voltage of 30kV. The SPE-CE device consisted of a short length of capillary packed with Oasis HLB sorbent, which was inserted near to the inlet end of the CE capillary. Several parameters affecting the in-line preconcentration were evaluated. The LOQs reached for hair samples were in the range of 0.3–2.5ng/mg with satisfactory analytical precision in both intraday and day-to-day experiments (RSDs <13%). Relative recoveries greater than 80% were obtained. The method has successfully been applied to the determination of these drugs of abuse in segmented hair from drug abusers who were undergoing methadone maintenance treatment. The results were consistent with the patients' statements, indicating that the method established herein can be used for verifying a history of drug abuse.

Determination of the unbound fraction of R- and S-methadone in human brain.

According to the free drug hypothesis, only the unbound fraction (f u ) of a given drug is biologically available in terms of its pharmacologic activity. Methadone shows large interpersonal variation in toxicity. The goal of the work presented here was to examine whether isolating the unbound fraction of the active R-methadone enantiomer from brain matter could be of value as a forensic tool. A method applying equilibrium dialysis to postmortem brain samples was validated and showed good reproducibility for the previously published f u values for eight common drugs (alprazolam, citalopram, codeine, methadone, morphine, diazepam, oxycodone, tramadol), as well as methadone enantiomers. This method was then applied to approximately 50 authentic case samples with R-methadone and S-methadone concentrations ranging from 0.03 to 13 and 0.6 to 6.8mg/kg, respectively; median f u values (R-and S-methadone) were 3.9% (range 3.0-5.3%) and 3.7% (range 2.9-4.9%). No overall correlation between the active R-methadone concentration and f u were found. Small but statistically significant differences in median f u for the R-methadone enantiomer were identified between case-categories (i.e., poisoning with multiple drugs, methadone poisoning, and deaths unrelated to methadone), but these are thought to be too low to be of any forensic value.

Genome-Wide Pharmacogenomic Study on Methadone Maintenance Treatment Identifies SNP rs17180299 and Multiple Haplotypes on CYP2B6, SPON1, and GSG1L Associated with Plasma Concentrations of Methadone R- and S-enantiomers in Heroin-Dependent Patients.

Methadone maintenance treatment (MMT) is commonly used for controlling opioid dependence, preventing withdrawal symptoms, and improving the quality of life of heroin-dependent patients. A steady-state plasma concentration of methadone enantiomers, a measure of methadone metabolism, is an index of treatment response and efficacy of MMT. Although the methadone metabolism pathway has been partially revealed, no genome-wide pharmacogenomic study has been performed to identify genetic determinants and characterize genetic mechanisms for the plasma concentrations of methadone R- and S-enantiomers. This study was the first genome-wide pharmacogenomic study to identify genes associated with the plasma concentrations of methadone R- and S-enantiomers and their respective metabolites in a methadone maintenance cohort. After data quality control was ensured, a dataset of 344 heroin-dependent patients in the Han Chinese population of Taiwan who underwent MMT was analyzed. Genome-wide single-locus and haplotype-based association tests were performed to analyze four quantitative traits: the plasma concentrations of methadone R- and S-enantiomers and their respective metabolites. A significant single nucleotide polymorphism (SNP), rs17180299 (raw p = 2.24 × 10−8), was identified, accounting for 9.541% of the variation in the plasma concentration of the methadone R-enantiomer. In addition, 17 haplotypes were identified on SPON1, GSG1L, and CYP450 genes associated with the plasma concentration of methadone S-enantiomer. These haplotypes accounted for approximately one-fourth of the variation of the overall S-methadone plasma concentration. The association between the S-methadone plasma concentration and CYP2B6, SPON1, and GSG1L were replicated in another independent study. A gene expression experiment revealed that CYP2B6, SPON1, and GSG1L can be activated concomitantly through a constitutive androstane receptor (CAR) activation pathway. In conclusion, this study revealed new genes associated with the plasma concentration of methadone, providing insight into the genetic foundation of methadone metabolism. The results can be applied to predict treatment responses and methadone-related deaths for individualized MMTs.

Quantitative determination of the enantiomers of methadone in human plasma and saliva by chiral column chromatography coupled with mass spectrometric detection

Methadone is a potent lipophilic synthetic opioid that is effective in the treatment of cancer pain and perceived benefit in difficult pain control scenarios (especially in cases of neuropathic pain). The use of methadone in clinical practice is challenging however, due to the narrow therapeutic window and large inter- and intra-individual variability in therapeutic response. Quantitation of the enantiomers d- and l-methadone (d- and l-MTD) in plasma and saliva provides a basis for studying its pharmacokinetics in patients with cancer and for monitoring efficacy, toxicity and side-effects. This assay involves quantitation of the enantiomers of methadone using their respective deuterated internal standards, in plasma and saliva matrices with no impact of ion suppression in either matrix. The analytical recoveries of d- and l-MTD from the saliva collection devices (Salivette®) are optimised in this novel method with an accurate and simple extraction method employing dichloromethane. Optimal enantioselective separations were achieved using an α1-acid glycoprotein chiral stationary phase and triple quadrupole tandem mass spectrometer. Linearity was demonstrated over 0.05–1000µg/L for both enantiomers in plasma and in saliva with correlation coefficients greater than 0.998. The lower limit of quantitation (LLOQ) was determined to be 0.1µg/L in plasma and saliva for d- and l-MTD. Accuracy of the method ranges from 100% to 106% even at the LLOQ and total precision, expressed as the coefficient of variation, was between 0.2% and 4.4% for both analytes in both matrices. A simple one step extraction procedure resulted in recoveries greater than 95% for both analytes, at concentrations as low as 0.5µg/L, from the Salivette®. The validated method was applied successfully in 14 paired plasma and saliva samples obtained from adult patients with cancer pain receiving methadone.

Assessment of drug-drug interactions between daclatasvir and methadone or buprenorphine-naloxone.

Hepatitis C virus (HCV) infection is common among people who inject drugs, including those managed with maintenance opioids. Pharmacokinetic interactions between opioids and emerging oral HCV antivirals merit evaluation. Daclatasvir is a potent pangenotypic inhibitor of the HCV NS5A replication complex recently approved for HCV treatment in Europe and Japan in combination with other antivirals. The effect of steady-state daclatasvir (60 mg daily) on stable plasma exposure to oral opioids was assessed in non-HCV-infected subjects receiving methadone (40 to 120 mg; n = 14) or buprenorphine plus naloxone (8 to 24 mg plus 2 to 6 mg; n = 11). No relevant interaction was inferred if the 90% confidence interval (CI) of the geometric mean ratio (GMR) of opioid area under the plasma concentration-time curve over the dosing interval (AUCτ) or maximum concentration in plasma (C max) with versus without daclatasvir was within literature-derived ranges of 0.7 to 1.43 (R- and S-methadone) or 0.5 to 2.0 (buprenorphine and norbuprenorphine). Dose-normalized AUCτ for R-methadone (GMR, 1.08; 90% CI, 0.94 to 1.24), S-methadone (1.13; 0.99 to 1.30), and buprenorphine (GMR, 1.37; 90% CI, 1.24 to 1.52) were within the no-effect range. The norbuprenorphine AUCτ was slightly elevated in the primary analysis (GMR, 1.62; 90% CI, 1.30 to 2.02) but within the no-effect range in a supplementary analysis of all evaluable subjects. Dose-normalized C max for both methadone enantiomers, buprenorphine and norbuprenorphine, were within the no-effect range. Standardized assessments of opioid pharmacodynamics were unchanged throughout daclatasvir administration with methadone or buprenorphine. Daclatasvir pharmacokinetics were similar to historical data. Coadministration of daclatasvir and opioids was generally well tolerated. In conclusion, these data suggest that daclatasvir can be administered with buprenorphine or methadone without dose adjustments.

Quantification of morphine, morphine 6-glucuronide, buprenorphine, and the enantiomers of methadone by enantioselective mass spectrometric chromatography in whole blood.

Deaths among drug addicts are frequently caused by intoxication with methadone and/or morphine. These drugs are often used in combination with other drugs, such as buprenorphine. In addition, methadone is generally used as a mixture of R- and S-enantiomers. To date, a method for separation and quantitation of these specific drugs has not been developed. The aim of this study was to develop a sensitive enantioselective method for quantitation of morphine, its active metabolite morphine 6-glucuronide, buprenorphine, and R- and S-methadone, in a single analytical run. Whole blood samples were diluted with 0.5 mol/L ammonium carbonate buffer and extracted on a Bond Elut C18 solid-phase extraction column with an automatic solid-phase extraction system. Chromatographic separation was performed on a chiral alpha-1-acid glycoprotein column with an acetonitrile/ammonium acetate buffer (10 mmol/L, pH 7.0, 22:78 v/v) mobile phase. The whole blood concentrations of the drugs were quantified by mass spectrometry using their stable isotope-labeled compounds as internal standards. The method was validated with respect to specificity, linearity, precision, limits of detection, and quantification and matrix effects. The precision (coefficient of variation) was below 15%, and the accuracy was between 90 and 115%. This method will be useful for routine analyses in forensic laboratories where blood samples are frequently analyzed for drugs of abuse. In some cases, sudden death from methadone overdose is caused by the enantiomeric form of the methadone, which makes the enantiomer separation capability of this method important.

Computer simulation of electrophoretic aspects of enantiomer migration and separation in capillary electrochromatography with a neutral selector.

A computer simulation study describing the electrophoretic separation and migration of methadone enantiomers in presence of free and immobilized (2-hydroxypropyl)-β-CD is presented. The 1:1 interaction of methadone with the neutral CD was simulated by using experimentally determined mobilities and complexation constants for the complexes in a low-pH BGE comprising phosphoric acid and KOH. The use of complex mobilities represents free solution conditions with the chiral selector being a buffer additive, whereas complex mobilities set to zero provide data that mimic migration and separation with the chiral selector being immobilized, that is CEC conditions in absence of unspecific interaction between analytes and the chiral stationary phase. Simulation data reveal that separations are quicker, electrophoretic displacement rates are reduced, and sensitivity is enhanced in CEC with on-column detection in comparison to free solution conditions. Simulation is used to study electrophoretic analyte behavior at the interface between sample and the CEC column with the chiral selector (analyte stacking) and at the rear end when analytes leave the environment with complexation (analyte destacking). The latter aspect is relevant for off-column analyte detection in CEC and is described here for the first time via the dynamics of migrating analyte zones. Simulation provides insight into means to counteract analyte dilution at the column end via use of a BGE with higher conductivity. Furthermore, the impact of EOF on analyte migration, separation, and detection for configurations with the selector zone being displaced or remaining immobilized under buffer flow is simulated. In all cases, the data reveal that detection should occur within or immediately after the selector zone.

Chiral on-line solid phase extraction coupled to liquid chromatography–tandem mass spectrometry assay for quantification of (R) and (S) enantiomers of methadone and its main metabolite in plasma

The authors aimed at developing a liquid chromatography tandem mass spectrometry (LC-MS/MS) method with online extraction to determine (R)- and (S)- methadone enantiomers and its main metabolite 2-ethylidine-1,5-dimethyl-3,3 diphenylpyrrolidine (EDDP) in plasma.The analysis combined straightforward sample preparation, consisting of protein precipitation with acetonitrile, and an online enrichment by a flush/back-flush cycle before the second dimension chromatography.Using D3-deuterated internal standards allows overcoming significant relative matrix effect. Our method was linear up to 2000ng/mL. This simple sample preparation provides sensitive (the limit of quantitation is 25ng/mL for (R,S)-methadone and EDDP and 12.5ng/mL for (R)- and (S)- methadone), accurate and precise (the intra-day and inter-day imprecision and inaccuracy are lower than 15%) quantification of the plasma concentration of these drugs.We have developed a reliable LC-MS/MS method for both routine therapeutic drug monitoring and pharmacokinetics studies and for toxicology analysis in the setting of methadone treatment or intoxication

Distribution of enantiomers of methadone and its main metabolite EDDP in human tissues and blood of postmortem cases.

Knowledge concerning the distribution of methadone in postmortem human tissue and the effect of postmortem redistribution on methadone is today limited making the choice of a suitable substitute for femoral blood difficult when this is not available. Cardiac blood, femoral blood, muscle, and brain tissue concentrations of the enantiomers of methadone and its metabolite 2-ethyl-1,5-dimethyl-3,3-diphenylpyrrolinium were recorded for 155 postmortem cases. Brain and muscle tissue concentrations exceeded the femoral blood concentrations with a median fold of 2.3 and 1.6, respectively, but both had a better correlation than cardiac blood to femoral blood concentrations. The Kruskal-Wallis test showed a significant dependency on time and body mass index for some of the matrix ratios over femoral blood. We conclude brain or muscle tissue may constitute a better alternative for measurement of methadone than cardiac blood for situations in which femoral blood is not available, despite concentrations in both matrices being systematically higher.

Ethnic and genetic factors in methadone pharmacokinetics: A population pharmacokinetic study

BackgroundTreatment of opiate use disorders with methadone is complicated by wide interindividual variability in pharmacokinetics. To identify potentially contributing covariates in methadone pharmacokinetics, we used population pharmacokinetic modeling to estimate clearance (CL/F) and volume of distribution (V/F) for each methadone enantiomer in an ethnically diverse methadone maintained population. MethodsPlasma levels of the opiate-active R-methadone and opiate-inactive S-methadone were measured in 206 methadone maintained subjects approximately two and twenty-three hours after a daily oral dose of rac-methadone. A linear one-compartment population pharmacokinetic model with first-order conditional estimation with interaction (FOCE-I) was used to evaluate methadone CL/F and V/F. The influence of covariates on parameter estimates was evaluated using stepwise covariate modeling. Covariates included ethnicity, gender, weight, BMI, age, methadone dose, and 21 single nucleotide polymorphisms in genes implicated in methadone pharmacokinetics. ResultsIn the final model, for each enantiomer, Hmong ethnicity reduced CL/F by approximately 30% and the rs2032582 (ABCB1 2677G>T/A) GG genotype was associated with a 20% reduction in CL/F. The presence of the rs3745274 minor allele (CYP2B6 515G>T) reduced CL/F by up to 20% for S-methadone only. A smaller effect of age was noted on CL/F for R-methadone. ConclusionThis is the first report showing the influence of the rs2032582 and rs3745274 variants on methadone pharmacokinetics rather than simply dose requirements or plasma levels. Population pharmacokinetics is a valuable method for identifying the influences on methadone pharmacokinetic variability.

Evaluación del programa de mantenimiento con metadona del Hospital Mental de Risaralda

BackgroundPsychosocial care and methadone maintenance treatment (MMT) are the preferred strategies for the management of heroin addicts, but the results are still unsatisfactory, justifying the search and intervention of the factors influencing the response to treatment. MethodologyIn order to determine the contribution of demographic, clinical and genetic variables on serum concentrations and response to methadone, we investigated patients on MMT, who were receiving methadone in supervised and unchanged doses at least during the previous two weeks. The age, gender, body mass index (BMI), duration of heroin abuse, addiction to other drugs, criminal background, current daily methadone doses, time spent in the TMM, comorbidity and concomitant medication were recorded. Blood samples were taken for the determination of serum levels of racemic methadone and its R and S-enantiomers, and for typing of candidate alleles of POR, CYP2B6, ABCB1, GRIN1, OPRM1, SLC6A3, DßH and ARRB2 genes, all associated with the metabolism, tissue distribution and mechanism of action of methadone. Methadone quantification was by HPLC-DAD, and the detection of genetic markers by Real Time PCR and VNTR methods. ResultsA total of 80 subject volunteers were enrolled, with a mean age of 23.5 (5) years (86% male), all of them were addicts of multiple drugs, 60% with a criminal background, 5.1 (2.9) years taking heroin, and 5.3 (4) months on MMT, and taking a supervised dose of 41 (12) mg/day methadone. The (R), (S) and (R, S) methadone enantiomer trough plasma levels were, 84 (40), 84 (42), and 168 (77) ng/mL, respectively. All genotypes were in Hardy-Weinberg equilibrium. The two urine tests were negative for heroin in 61.3% (49/80) of the volunteers, the decline in cocaine/crack use was 83%, 30% of marijuana, and other psychoactives (inhalants, benzodiazepines, amphetamines) decreased to zero, while the consumption of snuff remained at 93.5% (75/80). Blood concentrations of racemic methadone and its enantiomers were significantly associated with the dose/day of the medication, but none of the other demographic, clinical or genetic variables impacted on serum levels of methadone. As for the results of the MMT, non-users and occasional users of heroin, as well as those who stopped taking other psychoactive drugs, and the ones who did not, were similar as regards the demographic, genetic and clinical variables. This included the blood metahdone concentrations, except for individuals who did not reduce their consumption of other psychoactives other than heroin, who had significantly (P=.03) higher blood levels of S-methadone, compared with those who did stop taking them. ConclusionsThere was a significant reduction in the consumption of heroin and other psychoactives, and social rehabilitation of patients. However, the extensive overlap between effective and ineffective doses of methadone suggests the presence of personal and social variables that transcend the simple pharmacological management. These probably need to be addressed more successfully from the psychosocial features, particularly as regards to identifying and overcoming relapse-trigger experiences, as well as certain features of the patient, such as their psychological distress level or their psychiatric disorders.

Dynamic high‐resolution computer simulation of isotachophoretic enantiomer separation and zone stability

The development of electrophoretic computer models and their use for simulation of electrophoretic processes has increased significantly during the last few years. Recently, GENTRANS and SIMUL5 were extended with algorithms that describe chemical equilibria between solutes and a buffer additive in a fast 1:1 interaction process, an approach that enables simulation of the electrophoretic separation of enantiomers. For acidic cationic systems with sodium and H3 0(+) as leading and terminating components, respectively, acetic acid as counter component, charged weak bases as samples, and a neutral CD as chiral selector, the new codes were used to investigate the dynamics of isotachophoretic adjustment of enantiomers, enantiomer separation, boundaries between enantiomers and between an enantiomer and a buffer constituent of like charge, and zone stability. The impact of leader pH, selector concentration, free mobility of the weak base, mobilities of the formed complexes and complexation constants could thereby be elucidated. For selected examples with methadone enantiomers as analytes and (2-hydroxypropyl)-β-CD as selector, simulated zone patterns were found to compare well with those monitored experimentally in capillary setups with two conductivity detectors or an absorbance and a conductivity detector. Simulation represents an elegant way to provide insight into the formation of isotachophoretic boundaries and zone stability in presence of complexation equilibria in a hitherto inaccessible way.