Methamphetamine Enantiomers Research Articles

Post mortem chiral analysis of MDMA and MDA in human blood and hair

Drug-related fatalities in the EU are predominantly associated with opioids. MDMA (Ecstasy) consumption results in fewer lethal intoxications despite its widespread use. This study investigates MDMA-related fatalities, focusing on enantiomer ratios of MDMA and its metabolite MDA to explore the role of metabolism in fatal outcomes.MDMA induces euphoria, increased empathy, and physiological effects such as tachycardia, hypertension, and hyperthermia. Metabolism mainly involves CYP1A2 and CYP2D6, with polymorphism of the latter influencing metabolism rates. Our institute observed several MDMA-related fatalities, which prompted an investigation into the potential role of inefficient drug metabolism in these cases.A novel quantitative chiral analysis method was developed and validated for MDMA, MDA, amphetamine and methamphetamine enantiomers in human blood. Analysis of post mortem blood samples from eleven MDMA-related fatalities exhibited a wide range of concentrations and enantiomer ratios. Variability in R/S MDMA ratios, however, could be linked to the time period of metabolism. Hair analysis revealed high MDMA concentrations in all segments, irrespective of prior drug abuse anamnesis. Therefore, hair analysis may not be suitable for the assessment of past drug use in ecstasy-related fatalities.The results indicated that elevated levels of the MDMA enantiomer are correlated with longer survival times in cases of intoxication. However, there was no clear evidence for slowed MDMA metabolism as a cause of lethal intoxications. While challenges remain due to the diversity of cases, this study contributes valuable insights into ecstasy intoxications, aiding future interpretation of post mortem analysis.

The potential of the use of deep eutectic solvents and amino acid-based ionic liquids to enhance the chiral discrimination ability of different chiral selectors in capillary electrophoresis

The effect of the combined use of amino acid-based ionic liquids (AAILs) and deep eutectic solvents (DESs) with either cyclodextrin- (CD) or cyclofructan- (CF) based chiral selectors for the chiral separation of amphetamine derivatives was investigated in the present study. A non-significant improvement in enantiomeric separation of target analytes was observed when AAILs were combined with either CF or CD. On the other side, a markedly improved chiral separation of enantiomers was obtained using the dual carboxymethyl-β-cyclodextrin/DES system, highlighting the existence of a synergistic effect. After the addition of 0.5% v/v of choline chloride-ethylene glycol, the resolution of the enantiomers of amphetamine, methamphetamine and 3-fluorethamphetamine, increased from 1.4, 1.1, 1.0 to 1.8, 1.8, and 1.5 min, and the analysis times increased from 19.54, 20.48, 18.71 to 35.71, 35.78 and 32.90 min, respectively. This was not the case for the CF/DES dual system, in which the separation of amphetamines worsened, indicating an antagonistic effect. In conclusion, DESs are a very promising additive in capillary electrophoresis that can improve the separation of chiral molecules in combination with CDs but not CFs.

Monitoring Fast Synaptic Transmission of Neuromodulatory Transmitters To Define Drug Effects in the Brain.

Catecholaminergic systems are involved in a range of psychiatric disorders and are central mediators of the effects of stimulants on the brain and behavior. Advances in analytical detection methods paired with creative application of these approaches allow for recording noradrenergic and dopaminergic systems in the brain in isolation with subsecond resolution. Pauly et al. ( ACS Chem. Neurosci. 2023, 10.1021/acschemneuro.2c00689) define the distinct and differential mechanisms of methamphetamine enantiomers on dopamine and norepinephrine release, giving critical insights into the pharmacodynamic properties of a stimulant that is both abused and used clinically.

Enantioselective determination of plasma protein binding of common amphetamine-type stimulants

Amphetamine-type stimulants (ATS) like amphetamine (‘speed’), methamphetamine (‘crystal meth’) and 3,4-methylenedioxy-N-methylamphetamine (MDMA, ‘ecstasy’) represent some of the most frequently abused drugs worldwide. Another less frequently abused ATS is 4-fluoroamphetamine (4-FA). The enantiomers of these four compounds exhibit different pharmacokinetic and pharmacodynamic properties. According to the free drug theory, the pharmacological properties of a substance are dependent on its plasma protein binding (PPB). However, data on PPB of stimulant enantiomers in humans are rare or non-existent.Human plasma samples were spiked with racemic mixtures of the stimulants and subjected to ultrafiltration to extract the unbound fraction. Enantioselective liquid chromatography − tandem mass spectrometry (LC-MS/MS) methods were applied using a chiral Phenomenex® Lux3 μm AMP column. Method validation showed satisfactory selectivity, linearity (0.5 250 ng/mL), accuracy and precision. Enantiomers were quantified before and after ultracentrifugation to determine PPB.For all analytes, low to medium plasma protein binding was found. For (R)-amphetamine a slightly but significantly higher PPB was found compared to the (S)-enantiomer (31.7 % vs 29.0 %). (R)-MDMA also showed only slightly but significantly significantly higher PPB than (S)-MDMA, although the mean difference was negligible (21.6 % vs 21.3 %). For the enantiomers of methamphetamine and 4-FA, no significant differences in PPB were found.In summary, there were no or only minor differences in PBB for the enantiomers of all investigated compounds. The different pharmacological properties of the stimulant enantiomers can therefore not be explained by differences in PPB.

NMR Study of the Discrimination of Enantiomers of Methamphetamine and Its Raw Materials Using Chiral Solvating Agents

Some controlled substances, such as stimulants and narcotics, have asymmetric carbons in their molecules. Because the enantiomers do not always show the same pharmacological effects, and there are substances with different controls due to differences in their stereochemistry, a simple and unambiguous method for assessment of the composition of enantiomers is necessary. In this study, to develop a simple and rapid stereoscopic identification method for methamphetamine and its raw materials (ephedrine and pseudoephedrine), the 1H-NMR method was studied using three commercially available chiral solvating agents (CSAs); 1,1'-bi(2-naphthol)(BINOL), 2,2,2-trifluoro-1-(9-anthryl)ethanol (TFAE) and α-methoxy-α-(trifluoromethyl)phenylacetic acid (MTPA). In addition, the accuracy of the optical purity, which was measured using samples mixed with enantiomers in various ratios, was investigated. The NMR peaks of the enantiomers were separated by adding (R)- or (S)-form of BINOL, TFAE or MTPA to the chloroform-d solution of methamphetamine, ephedrine or pseudoephedrine. A sufficient discrimination of enantiomers was obtained by adding about 10 equal amounts of each CSA to the solutions. With regard to the optical purity, it was possible to determine accurately the mixing of small amounts of enantiomers of about 5% even if the NMR peaks did not reach the baseline separation, when impurity peaks do not overlap. This method will be one of the useful techniques for the rapid and simple discrimination of enantiomers of illegal methamphetamine and its raw materials.

Chiral Analysis of Amphetamine and Methamphetamine in Urine by Liquid Chromatography-Tandem Mass Spectrometry Applying Mosher Derivatization

Methamphetamine (MA) and amphetamine (AM) are central nervous system doping agents, listed in the WADA Prohibited List, which in the past years showed the highest prevalence among all stimulants. Specifically, concerning these stimulants, it is essential to distinguish between d and l-isomers of AM and MA due to their distinction in illicit or legal nonprescription use. The main separation methods used for the urinary determination of MA and AM enantiomers are based on chromatographic procedures, such as gas chromatography after chiral derivatization or liquid chromatography (LC) applying chiral columns, followed by the mass spectrometer (MS) detection. The use of the optically pure reagent R-(-)-α-methoxy-α-(trifluoromethyl) phenylacetic acid chloride (R-MTPA-Cl – Mosher’s Reagent) allows the isomeric separation by both chromatographic procedures while preserving the isomeric composition of the substances. Therefore, in the present work, an LC-MS2 approach has been developed for AM and MA detection in urine after Mosher derivatization. The method was validated and can be used for the enantioselective confirmation of AM and MA for doping control purposes and forensic analysis in general, being useful for the distinction between the medicinal and illicit use of both stimulants.

Research Progress on Chiral Separation Methods and Toxic Effects of Methamphetamine Enantiomers.

Methamphetamine （MAMP） is a kind of amphetamine-type stimulants （ATS） which contains one chiral carbon atom in its structure. Therefore a pair of enantiomers, S-（+）-MAMP and R-（-）-MAMP exist. R type and S type methamphetamines possess similar physicochemical properties, but has largely different pharmacological and toxic effects. S-（+）-MAMP is the main component of addictive drug "Ice" at present, seriously affecting human health and public safety. The separation analysis and mechanism of toxic effects discussions on MAMP are the current research focuses. This paper reviews the research progress of separation analysis methods and toxic effects of methamphetamine enantiomers to provide reference for forensic study and forensic practice.

Chromatographic separation of R-(-)/S-(+)-enantiomers of amphetamine and methamphetamine: differentiation between single methamphetamine consumption and co-consumption with amphetamine using enantioselective quantitative LC-MS/MS analysis.

The differentiation between single methamphetamine consumption and co-consumption with amphetamine is difficult, however possible by enantioselective analysis due to different preferred synthesis pathways of both substances. We quantified (R)-(-) and (S)-(+)-enantiomers of methamphetamine and amphetamine by a fast liquid chromatographic tandem-mass spectrometric method using a Lux® 3-μm AMP 150 × 3.0mm analytical column after simple protein precipitation with methanol. Method validation for quantitative detection showed limits of quantification < 5ng/mL, linearity in a range between 5 and 300ng/mL and bias and imprecision data < 15%. Overall, 134 plasma samples of police cases from the German regions of Franconia and Northrhine-Westphalia were analyzed for the enantiomers of methamphetamine and amphetamine. In 28 cases, the intake of racemic illicit amphetamine could be demonstrated; (R)-(-) / (S)-(+)-amphetamine concentration ratios in these cases were between 1.38 and 4.50 with most of the ratios being < 2.0. These ratios were compared to a subgroup of 25 consumers with a co-consumption of (S)-(+)-methamphetamine and racemic amphetamine detected by the qualitative proof of (R)-(-)-amphetamine but also by (R)-(-) / (S)-(+)-amphetamine concentration ratios (< 1 in 11 of 25 cases). Within our collective of 106 plasma samples after methamphetamine use, 25 samples showed co-consumption with amphetamine which shows that co-consumption of both stimulants is not a rare scenario. Furthermore, we could show that if non-stereoselective methods are used and the concentration ratio of total methamphetamine/total amphetamine is determined, a reliable estimation of co-consumption is not possible.

Chromatographic separation of R/S-enantiomers of amphetamine and methamphetamine: Pathways of methamphetamine synthesis and detection in blood samples by qualitative enantioselective LC–MS/MS analysis

Methamphetamine can be synthesized either enantiopure or in its racemic form. We separated (R)- and (S)-enantiomers of methamphetamine and amphetamine by a fast LC–MS/MS-method using a Lux® 3μm AMP 150×3.0mm analytical column after simple protein precipitation with methanol. Sufficient resolution could be achieved. Method validation for qualitative detection showed limits of quantification <5ng/mL while only little (maximum 14.5%) ion suppression could be shown. Stability in the processed sample could be achieved using isotopically labelled internal standards. Plasma samples of police cases from the german regions of Franconia and Northrhine revealed that in the majority of 106 tested samples (>99%) only (S)-methamphetamine was detected which leads to the conclusion that, in Germany, predominantly enantiopure (S)-methamphetamine is consumed which is synthesized via (1R,2S)-ephedrine or (1S,2S)-pseudoephedrine. However, racemic methamphetamine seems also to be on the market.

Enantiomeric profiling of amphetamine and methamphetamine in wastewater: A 7-year study in regional and urban Queensland, Australia

Enantiomeric profiling was used in this study to investigate the consumption of amphetamine and methamphetamine in regional and urban Southeast Queensland, Australia over a period of seven years. S(+) methamphetamine was predominantly consumed in both urban and regional areas, showing a two and three fold increase in urban and regional catchments respectively between 2011 and 2017. The ratio of amphetamine to methamphetamine (AMP/METH) in wastewater reflected the expected excretion profile of methamphetamine consumption indicating the presence of amphetamine in this study was primarily the result of methamphetamine metabolism. However, the occasional occurrence of R(−) amphetamine in samples containing higher AMP/METH ratios, suggested the consumption of racemic amphetamine. The R(−) methamphetamine enantiomer was also identified in several samples, possibly indicative that the phenyl-2-propanone (P2P) synthesis process rather than the more typical reduction of ephedrines was also being used to manufacture methamphetamine. Furthermore, we identified two samples with a significantly different enantiomer ratio for the METH and AMP as well as a much lower AMP/METH concentration ratio suggesting contribution from direct disposal of methamphetamine into the sewer. This study demonstrated that enantiomeric profiling in wastewater-based epidemiology can provide valuable information for evaluating the origin of amphetamine in wastewater as either a metabolite of methamphetamine consumption or amphetamine itself.

Disappearance of R/S-methamphetamine and R/S-amphetamine from human scalp hair after discontinuation of methamphetamine abuse

Methamphetamine (MA) and amphetamine (AM) are widely abused drugs. These compounds contain a chiral center, and their enantiomers exhibit different pharmacologic, pharmacokinetic, and metabolic properties due to differences in binding affinities to their receptor sites. Until now, there was a lack of information on the decline in the concentration of drugs in hair after abstinence. A simple procedure for the chiral separation and determination of methamphetamine (MA) and its metabolite amphetamine (AM) enantiomers by LC–MS/MS in hair samples has been developed and fully validated. The LODs and LLOQs were 0.02ng/mg and 0.05ng/mg for all analytes, respectively. This method was successfully applied to both real hair specimens from chronic MA users and after the discontinuation of MA. The concentration of total MA, and total AM in fifty-eight authentic hair specimens ranged from 7.8ng/mg to 521.0ng/mg, and from 0.3ng/mg to 84.0ng/mg, respectively. Both enantiomers of MA and/or AM were detected in seven of fifty-eight authentic hair specimens. Hair specimens were from thirteen women with a known history of MA abuse, who went to a rehabilitation center and ceased consuming MA (for 4–5 months). The S-isomers of MA and AM were detected in all the 5–6cm segments. Both enantiomers of MA were detected in nine of the 5–6cm segments and the enantiomers of AM were found in only five of the nine samples. Assuming a hair growth rate of 1cm/month, the mean hair elimination half-lives of S-MA, R-MA, S-AM, and R-AM were 0.64(95% CI, 0.46–0.96), 0.58(95% CI, 0.41–0.93), 0.62(0.49–0.88), and 0.50 months (95% CI, 0.42–0.56), respectively. With the developed method, R/S-MA and R/S-AM could be detected in the hair of former drug users for approximately 4 months after abstinence. S-MA is the most commonly found analyte in hair segments and is principally used by abusers. Our results suggest that to evaluate the discontinuation of MA abuse after a 6-month period of abstinence, a 3-cm proximal hair segment should be free of MA at the SOHT proposed cut-off level.

Enantioseparation of methamphetamine by supercritical fluid chromatography with cellulose-based packed column

The enantiomers of methamphetamine were differentiated by supercritical fluid chromatography (SFC) with an enantioselective cellulose-based packed column. The optimization of the chromatographic conditions was achieved by changing column temperature, co-solvent proportion, additive concentration, flow rate and back pressure. In particular, the additive concentration crucially changed the resolution between the enantiomers. After determining the optimized conditions, the enantiomers of methamphetamine were successfully separated. The analytical precision, accuracy and limit of detection were checked by using the authentic standard and seized real samples. We believe that chiral SFC is a promising method for enantioseparation of forensic samples.

Determination of methamphetamine enantiomer composition in human hair by non-chiral liquid chromatography-tandem mass spectrometry method.

Chiral separation is crucial for investigating methamphetamine positive cases. While (S)-(+)-enantiomer of methamphetamine (S-MAMP) is a schedule II controlled substance, (R)-(-)-enantiomer (R-MAMP) is an active ingredient of a few over-the-counter drugs in the United States. Among biological specimen types, hair provides greater detection window than blood, urine or oral fluid, and are therefore regarded with particular interest. Herein we describe a novel non-chiral liquid chromatography-tandem mass spectrometry (LC-MS/MS) method to directly determine methamphetamine enantiomeric composition (percentage) in hair specimens. Hair samples were washed once with acetone, powdered, incubated overnight at 53°C in 0.1M hydrochloric acid (HCl), and subjected to a solid phase extraction (SPE). The extracts were derivatized using Marfey's reagent at 53°C for 60min. The final mixture was analyzed by LC-MS/MS. Chromatographic separation was achieved using a C18 Kinetex analytical column and 60% (v/v) aqueous methanol as mobile phase (isocratic). Triple quadrupole mass spectrometer was equipped with an electro-spray ionization (ESI) source operating in negative mode and the chromatograms were acquired using a multiple-reaction monitoring (MRM) approach. The results were expressed as ratio of R- to S-MAMP and then derived to composition percentages without requiring quantitating each enantiomer. The method was precise and accurate across 0-100% S-composition at a range of 80-18,000pg/mg. The performance of the new method was compared with an (S)-(-)-N-trifluoroacetylprolyl chloride (S-TPC) derivatization and gas chromatography-mass spectrometry (GC-MS) method on authentic methamphetamine-positive hair samples. Not only the new Marfey's reagent approach presented satisfactory correlation with the S-TPC approach, but it also exhibited significantly improved quality (e.g., S/N) of the chromatograms. In summary, our protocol employs cost effective and minimally hazardous Marfey's reagent to derivatize trace amounts of methamphetamine extracted from hair samples and a non-chiral LC-MS/MS approach to separate and identify the two enantiomers. The method allows determination of the methamphetamine enantiomeric composition without requiring quantitation of each enantiomer and is therefore well suited for further investigate previously determined methamphetamine positive cases. This method represents a viable tool for evaluation of long-term drug exposure.

Improved Chiral Separation of Methamphetamine Enantiomers Using CSP-LC-MS-MS.

To determine the true enantiomeric composition of methamphetamine urine drug testing results, chiral separation of dextro (D) and levo (L) enantiomers is necessary. While enantiomeric separation of methamphetamine has traditionally been accomplished using gas chromatography-mass spectrometry (GC-MS), chiral separation of D- and L-methamphetamine by chiral stationary phase (CSP) liquid chromatography-mass spectrometry/mass spectrometry (LC-MS-MS) has proved more reliable. Chirally selective detection of methamphetamine by GC-MS is often performed using L-N-trifluoroacetyl-prolyl chloride (TPC). L-TPC, a chiral compound, is known to have impurities that can affect the chiral composition percentages of the methamphetamine sample, potentially leading to inaccurate patient results. The comparative analysis of the samples run by GC and LC methods showed preferential bias of the GC method for producing error rates, consistent with previous research, of 8-19%. The CSP-LC-MS-MS method produces percent deviation errors of <2%. Additionally, the GC method failed to produce results that were 100% D- or L-isomer even for enantiomerically pure standards. A higher rate of D- and L-methamphetamine isomer racemization is seen in samples when analyzed by GC-MS using L-TPC-derivatizing agent. This racemization is not seen when these samples are tested with CSP-LC-MS-MS. Thus, a more accurate method of enantiomeric analysis is provided by CSP-LC-MS-MS.

Enantiomer Profiling of Methamphetamine in White Crystal and Tablet Forms (Ma Old) Using LC-MS-MS.

At the global level, seizures of crystalline methamphetamine (MA) and MA tablets have risen to a new high, indicating that the substance is an imminent threat. MA enantiomer profiling was a useful tool to investigate the prevalence of MA abuse, the intrinsic characteristics of the seized samples and the trends of precursors. In this work, the distribution of enantiomers in clandestine MA crystals and tablets seized mainly in the Yangtze River Delta region, China, from 2008 to 2014 were identified. The MA samples were diluted with internal standard methanol solution, and analyzed by LC-MS-MS. The detection limits of the enantiomers were 0.04 μg/L. The limit of quantification was 0.1 μg/L. As little as 0.2% of the R-enantiomer ratio could be determined. Standard calibration curves of S- and R-MA showed good linearity in the range of 0.1-80 μg/L (r(2) > 0.995). All of the seizures were optically pure S-enantiomer in the years 2008 and 2009. Seized samples containing a slight amount of R-MA began to appear in 2010 and increased in the year 2014. No racemic mixture or R-isomer of MA was seized. From this study, we could find out that smuggling routes and/or precursors might be silently changing in the Yangtze River Delta region, China.

Chiral separation and determination of R/S-methamphetamine and its metabolite R/S-amphetamine in urine using LC–MS/MS

Methamphetamine (MA) and amphetamine (AM) are widely abused drugs. Differentiation of MA and/or AM abuse from therapeutic ingestion of MA and/or AM or one of their precursor drugs is therefore of relevance in clinical and forensic toxicology. The aim of the study was to develop a simple, rapid, and accurate method for the chiral separation and determination of R/S-MA and R/S-AM in urine using liquid chromatography electrospray ionization tandem mass spectrometry operating in the positive ion multiple-reaction monitoring (MRM) mode. 20μL of urine was diluted 500 times and 20μL was injected. The chromatographic system consisted of a Chirobiotic™ V2 column (2.1mm×250mm, 5μm), and the mobile phase was methanol containing 0.1% (v/v) glacial acetic acid and 0.02% (v/v) ammonium hydroxide. The method was fully validated through assessments of its linearity (0.05–50.00mg/L, r2>0.994 for all analytes), and LOQ (0.05mg/L for all analytes). No matrix effect was observed. The method was successfully applied to 86 urine samples from suspected MA abusers. Only the S-isomers of MA and AM were detected in 72 samples. The concentrations of R-MA ranged from below the LOQ to 13.76mg/L in 14 urine samples with both enantiomers of MA and/or AM. Pure S-MA is the most common found analyte in urine and principally used by abusers.

Evaluation of the 20% D-Methamphetamine Requirement for Determining Illicit Use of Methamphetamine in Urine

In urine drug testing, enantiomer analysis is used to determine whether a positive methamphetamine result could be due to use of an over-the-counter (OTC) nasal inhaler containing L-methamphetamine. D-methamphetamine at more than 20% of the total is considered indicative of a source other than an OTC product. This interpretation is based on a 1991 Department of Health and Human Services (HHS) Technical Advisory. We performed studies to verify the methamphetamine enantiomer content of current OTC nasal inhalers and to evaluate current laboratory testing capabilities. This study demonstrated that OTC inhalers contain less than 1% D-methamphetamine. A proficiency testing (PT) set for HHS-certified laboratories performing methamphetamine enantiomer testing found D-methamphetamine percentages that were consistently 1 to 3% higher than theoretical due to optical impurity of the derivatizing reagent N-trifluoroacetyl-L-prolyl chloride (L-TPC). The PT results also demonstrate that laboratories can accurately determine 20% D-methamphetamine in samples with total methamphetamine concentrations down to 250 ng/mL. Based on these studies, the guideline of >20% D-methamphetamine is appropriate for interpreting results obtained using current laboratory methods.

Detection of the d (dextro) and l (levo) methamphetamine enantiomers in a population of those with pain

Detection of the d (dextro) and l (levo) methamphetamine enantiomers in a population of those with pain

Intradiscal pressurized physiologic saline injection drastically reduced pain from cervical and lumbar disc herniation

Intradiscal pressurized physiologic saline injection drastically reduced pain from cervical and lumbar disc herniation

Stereoselectivity in the human metabolism of methamphetamine

To characterize the formation and urinary elimination of metabolites of S-(+) and R-(-) methamphetamine (MA) in humans. In this 12-subject, six-session, double-blind, placebo-controlled, balanced, crossover design study, the formation of the MA metabolites para hydroxymethamphetamine (pOH-MA) and amphetamine (AMP) were determined in urine after intravenous doses of S-(+)-MA 0.25 and 0.5 mg kg(-1), R-(-)-MA 0.25 and 0.5 mg kg(-1), racemic MA 0.5 mg kg(-1), or placebo. Parent drug and metabolite levels in urine and plasma were measured by gas chromatography-mass spectrometry. Pharmacokinetic parameters were calculated by noncompartmental models using WinNonlin. An approximately threefold enantioselectivity difference in elimination was observed for AMP, with 7% of the dose converted to S-(+)-AMP vs. 2% to R-(-)-AMP (P < 0.001). Furthermore, less R-(-)-pOH-MA was excreted in the urine compared with S-(+)-pOH-MA (8% vs. 11%, P= 0.02). Correspondingly, S-(+)-MA excretion was less than R-(-)-MA (42% vs. 52%; P= 0.005). The metabolism of MA is enantioselective, with formation of AMP having the highest isomer selectivity. A greater percentage of MA is converted to pOH-MA (8-11%) than AMP (2-7%). The formation of pOH-MA was less affected by the MA enantiomer administered, suggesting that urine pOH-MA may be a more stable biomarker of MA metabolism.