Mephedrone pharmacokinetics after intravenous and oral administration in rats: relation to pharmacodynamics

Fe d'errates disponible a: http://​dx.​doi.​org/​10.​1007/​s00213-013-3283-6Rationale Mephedrone (4-methylmethcathinone) is a still poorly known drug of abuse, alternative to ecstasy or cocaine. Objective The major aims were to investigate the pharmacokineticsa and locomotor activity of mephedrone in rats and provide a pharmacokinetic/pharmacodynamic model. Methods Mephedrone was administered to male Sprague-Dawley rats intravenously (10 mg/kg) and orally (30 and 60 mg/kg). Plasma concentrations and metabolites were characterized using LC/MS and LC-MS/MS fragmentation patterns. Locomotor activity was monitored for 180-240 min. Results Mephedrone plasma concentrations after i.v. administration fit a two-compartment model (α=10.23 h−1, β=1.86 h−1). After oral administration, peak mephedrone concentrations were achieved between 0.5 and 1 h and declined to undetectable levels at 9 h. The absolute bioavailability of mephedrone was about 10 % and the percentage of mephedrone protein binding was 21.59±3.67%. We have identified five phase I metabolites in rat blood after oral administration. The relationship between brain levels and free plasma concentration was 1.85±0.08. Mephedrone induced a dose-dependent increase in locomotor activity, which lasted up to 2 h. The pharmacokinetic-pharmacodynamic model successfully describes the relationship between mephedrone plasma concentrations and its psychostimulant effect. Conclusions We suggest a very important first-pass effect for mephedrone after oral administration and an easy access to the central nervous system. The model described might be useful in the estimation and prediction of the onset, magnitude,and time course of mephedrone pharmacodynamics as well as to design new animal models of mephedrone addiction and toxicity

Identification of Novel Metabolic Pathways of Sitagliptin (STG) by LC/MS and LC/MS2 after Incubations with Rat Hepatocytes

Sitagliptin (STG), a drug for treating Type Ii Diabetes Mellitus (T2DM), has been associated with severe joint pain in some patients. In this paper the metabolic profile of the drug has been investigated in order to determine metabolism and formation of reactive compounds which may contribute to this adverse effect. Metabolism of STG was investigated in vitro by incubation with freshly isolated Sprague-Dawley rat hepatocytes, to characterize Phase I and II metabolites, and the reaction mixture analysed on a zwitter ionic hydrophilic interaction (ZIC®-HILIC) column using LC-MS and LC-MS2 utilising electrospray ionization (ESI) in the positive ion mode. STG was metabolised to yield eleven metabolites, but in total only 3.1% of the parent drug was metabolised over 2 hrs incubation. These metabolites were structurally characterized on the basis of accurate mass analyses and the major metabolic routes for STG determined to be via aromatic oxidation (0.86%) and desaturation of N-C and C-C of the piperazine (0.44%). Novel metabolites of STG detected using these methods included STG N-glucuronide (M6) and a di-ketone metabolite (M4), hydroxylation of both the amine group and aromatic ring followed by formation of glucuronide metabolites (M5, M5’), oxidative desaturation of NH2 and di-hydroxylation of metabolites followed by loss of HF. Also, observed was an N-sulfate metabolite (0.07%) and acetylation followed by glucuronide conjugation was also found in trace amounts (<0.01%). MS2 fragment ions provide additional structural confirmation providing a possible structure for most metabolites such as by fragment ion loss of the glucuronide group (176 Da) from metabolite M5 and loss of the phenolic sulfate (80 Da) of N-Sulfate metabolite (M7). Reduction reaction of piperazine ring probably generates highly electrophilic metabolite of STG, which may be susceptible to produce adverse effects. Furthermore, N-oxidation reaction forming reactive intermediates metabolic to give a hydroxylamine metabolite that may undergo further reactions to yield electrophilic intermediate metabolites

Origami Chips: Development and validation of a paper-based Lab-on-a-Chip device for the rapid and cost-effective detection of 4-methylmethcathinone (mephedrone) and its metabolite, 4-methylephedrine in urine

4-methylmethcathinone (mephedrone) has emerged in drug seizures as a new psychoactive substance (NPS) causing a public health risk of global concern. Currently, there are no commercial microfluidic devices for the selective detection of mephedrone and so this study presents a simple, low cost and portable paper-based Lab-on-a-Chip (LOC) device with colorimetric detection to fill this gap. Limits of detection for mephedrone in spiked urine and dissolved powder (aqueous) samples are clinically relevant at 4.34 ng mL-1 and 2.51 ng mL-1 respectively. No cross-reactivity for commonly encountered cutting agents, interferents and adulterants were detected. Mephedrone and its main metabolite were detectable in aqueous samples within 3 minutes. Stability and reproducibility measurements showed no significant difference in signal intensity over eight weeks and no significant difference within or between devices. The proposed device has the potential to provide cost-effective, rapid, on-site testing within forensic or clinical settings and therefore has wide global applicability

Detection and quantification of new psychoactive substances (NPSs) within the evolved "legal high" product, NRG-2, using high performance liquid chromatography-amperometric detection (HPLC-AD)

The global increase in the production and abuse of cathinone-derived New Psychoactive Substances (NPSs) has developed the requirement for rapid, selective and sensitive protocols for their separation and detection. Electrochemical sensing of these compounds has been demonstrated to be an effective method for the in-field detection of these substances, either in their pure form or in the presence of common adulterants, however, the technique is limited in its ability to discriminate between structurally related cathinone-derivatives (for example: (±)-4′-methylmethcathinone (4-MMC, 2a) and (±)-4′-methyl-N-ethylmethcathinone (4-MEC, 2b) when they are both present in a mixture. In this paper we demonstrate, for the first time, the combination of HPLC-UV with amperometric detection (HPLC-AD) for the qualitative and quantitative analysis of 4-MMC and 4-MEC using either a commercially available impinging jet (LC-FC-A) or custom-made iCell channel (LC-FC-B) flow-cell system incorporating embedded graphite screen-printed macroelectrodes. The protocol offers a cost-effective, reproducible and reliable sensor platform for the simultaneous HPLC-UV and amperometric detection of the target analytes. The two systems have similar limits of detection, in terms of amperometric detection [LC-FC-A: 14.66 μg mL−1 (2a) and 9.35 μg mL−1 (2b); LC-FC-B: 57.92 μg mL−1 (2a) and 26.91 μg mL−1 (2b)], to the previously reported oxidative electrochemical protocol [39.8 μg mL−1 (2a) and 84.2 μg mL−1 (2b)], for two synthetic cathinones, prevalent on the recreational drugs market. Though not as sensitive as standard HPLC-UV detection, both flow cells show a good agreement, between the quantitative electroanalytical data, thereby making them suitable for the detection and quantification of 4-MMC and 4-MEC, either in their pure form or within complex mixtures. Additionally, the simultaneous HPLC-UV and amperometric detection protocol detailed herein shows a marked improvement and advantage over previously reported electroanalytical methods, which were either unable to selectively discriminate between structurally related synthetic cathinones (e.g. 4-MMC and 4-MEC) or utilised harmful and restrictive materials in their design

An overview of recent developments in the analytical detection of new psychoactive substances (NPSs)

New psychoactive substances (NPSs), sometimes referred to as “legal highs” in more colloquial environments/ the media, are a class of compounds that have been recently made available for abuse (not necessarily recently discovered) which provide similar effects to the traditional well studied illegal drugs but are not always controlled under existing local, regional or international drug legislation. Following an unprecedented increase in the number of NPSs in the last 5 years (with 101 substances discovered for the first time in 2014 alone) its, occasionally fatal, consequences have been extensively reported in the media. Such NPSs are typically marketed as ‘not for human consumption’ and are instead labelled and sold as plant food, bath salts as well as a whole host of other equally nondescript aliases in order to bypass legislative controls. NPSs are a new multi-disciplinary research field with the main emphasis in terms of forensic identification due to their adverse health effects, which can range from minimal to life threatening and even fatalities. In this mini-review we overview this recent emerging research area of NPSs and the analytical approaches reported to provide detection strategies as well as detailing recent reports towards providing point-of-care/in-the-field NPS (“legal high”) sensors

Developing methods for profiling of cathinone derivatives and elucidation of their metabolites formed in rat and human hepatocytes

The recent global increase in the abuse of 4'-methylmethcathinone (mephedrone, 3a) and related compounds has led to a requirement for full chemical characterisation of these products. In this thesis the synthesis and characterisation of the hydrobromide salts of two mephedrone derivatives: 4'-methyl-N-ethylcathinone (4-MEC, 3c) and 4'-methyl-N-benzylcathinone (4-MBC, 3d) is reported. These compounds were previously identified in samples of the "legal high" NRG-2. Additionally, the first fully validated chromatographic methods for the detection and quantitative analysis of these substances both in their pure form, and in the presence of a number of common adulterants used in illicit drug manufacture are reported. (±)-mephedrone [4-MMC, 3a] is a synthetic "legal high", with a classical cathinone structure similar to methcathinone. The in vitro metabolism of 4-MMC (3a) was investigated in freshly Sprague-Dawley rat hepatocytes to characterize the associated Phase I and II metabolites , 2x106 cells mL-1 were incubated with 4-MMC (3a) The reaction mixture analysed on a (ZIC®-HILIC) column using LC-MS and LC-MS2 on Orbitrap instruments; 4-MMC yielded seventeen metabolites. These metabolites were structurally characterized on the basis of accurate mass analyses and LCMS2 fragmentation patterns and the major metabolic routes for 4-MMC(3a) determined to be via (i) oxidation of the 4'-methyl group and (ii) reduction of the β-keto moiety, with a range of Phase II metabolites also being formed including several glucuronides and acetates. The biotransformation of a modified 4'-trifluoromethyl- derivative (4-TFMMC, 3b) was also studied and showed significant differences in its metabolism compared to 4-MMC (3a). Key pharmacokinetic parameters for both drugs have been calculated [biological half-lives (t1/2) for 4-MMC (3a) = 61.9 minutes and for 4-TFMMC = 203.8 minutes] and these data may aid in the understanding of in vivo metabolism and the likely pharmacokinetic effects of chemical/structural modifications within this class of controlled substances. The same procedure was carried out with 4'-methyl-N-ethylcathinone [4-MEC (3c)] and similar pathways were observed. Cryopreserved human hepatocytes (0.5x106 cells mL-1) were incubated with 4-MMC (3a) and 4-TFMMC (3b) to investigate the metabolic pathways in human cells. Using IV the same analytical techniques, it was found that the metabolic pathways of in vitro cryopreserved human hepatocytes for 4-MMC (3a) and 4-TFMMC (3b) were similar to those of rat hepatocytes. However, human hepatocytes demonstrated slower metabolism and some metabolites were absent compared to the rat hepatocytes.The recent global increase in the abuse of 4'-methylmethcathinone (mephedrone, 3a) and related compounds has led to a requirement for full chemical characterisation of these products. In this thesis the synthesis and characterisation of the hydrobromide salts of two mephedrone derivatives: 4'-methyl-N-ethylcathinone (4-MEC, 3c) and 4'-methyl-N-benzylcathinone (4-MBC, 3d) is reported. These compounds were previously identified in samples of the "legal high" NRG-2. Additionally, the first fully validated chromatographic methods for the detection and quantitative analysis of these substances both in their pure form, and in the presence of a number of common adulterants used in illicit drug manufacture are reported. (±)-mephedrone [4-MMC, 3a] is a synthetic "legal high", with a classical cathinone structure similar to methcathinone. The in vitro metabolism of 4-MMC (3a) was investigated in freshly Sprague-Dawley rat hepatocytes to characterize the associated Phase I and II metabolites , 2x106 cells mL-1 were incubated with 4-MMC (3a) The reaction mixture analysed on a (ZIC®-HILIC) column using LC-MS and LC-MS2 on Orbitrap instruments; 4-MMC yielded seventeen metabolites. These metabolites were structurally characterized on the basis of accurate mass analyses and LCMS2 fragmentation patterns and the major metabolic routes for 4-MMC(3a) determined to be via (i) oxidation of the 4'-methyl group and (ii) reduction of the β-keto moiety, with a range of Phase II metabolites also being formed including several glucuronides and acetates. The biotransformation of a modified 4'-trifluoromethyl- derivative (4-TFMMC, 3b) was also studied and showed significant differences in its metabolism compared to 4-MMC (3a). Key pharmacokinetic parameters for both drugs have been calculated [biological half-lives (t1/2) for 4-MMC (3a) = 61.9 minutes and for 4-TFMMC = 203.8 minutes] and these data may aid in the understanding of in vivo metabolism and the likely pharmacokinetic effects of chemical/structural modifications within this class of controlled substances. The same procedure was carried out with 4'-methyl-N-ethylcathinone [4-MEC (3c)] and similar pathways were observed. Cryopreserved human hepatocytes (0.5x106 cells mL-1) were incubated with 4-MMC (3a) and 4-TFMMC (3b) to investigate the metabolic pathways in human cells. Using IV the same analytical techniques, it was found that the metabolic pathways of in vitro cryopreserved human hepatocytes for 4-MMC (3a) and 4-TFMMC (3b) were similar to those of rat hepatocytes. However, human hepatocytes demonstrated slower metabolism and some metabolites were absent compared to the rat hepatocytes

Determination of PPD in Hair Dyes Collected from Local Markets in El-Bieda City - Libya

This study aims to determine the para-phenylenediamine (PPD) in hair dyes by High-Performance Liquid Chromatography (HPLC). In the presented study, ten hair dye samples were collected from local markets in El-Bieda - Libya. A rapid, simple and reliable method is developed and validated for the determination of PPD in hair dyes samples using 50% methanol solution as solvent. The method was validated over a wide linear range of 5 – 25 µg/mL with correlation coefficients being consistently greater than 0.997. The LOD and LOQ were 1.21 µg/ ml and 3.67 µg/ ml, respectively. Relative standard deviation (% R.S.D.) ranged between 0.07 and 1.15 %. The minimum PPD level was observed in Beauty Touch (Blonde) sample (0.0855 % w/w), while Jourin sense Cosmetics (Black Blue) sample showed the highest PPD content (2.2526 % w/w). The HPLC measurments indicated that the results of PPD concentrations in Black colour samples were in the range of 0.3705 % - 2.2526 % w/w. Whereas its concentrations in anthor colour samples were in the range of 0.0855 % - 0.5936 % w/w. The level of PPD in Black colour samples were higher than other colour samples. The PPD content in all the samples analyzed in this study are well below the allowable limits set by the US Food and Drugs Administration.</jats:p

Detection and Quantitative Estimation of Toxic Acrylamide Levels in Selected Potatoes Chips and French Fries from the Libyan Market Using HPLC-UV Method

Acrylamide is a potential health hazardous compound occurring in baked and fried food as a result of excessive dry heating during the preparation and/or processing of foods. Exposure to a high level of acrylamide may cause cancer, neurotoxicity, and mutagenicity. In this study, an isocratic reversed-phase high-performance liquid chromatographic (HPLC) method using a C18 column was used for the determination of acrylamide in selected food. The mobile phase consisted of 0.1% formic acid in water: acetonitrile (98:02), and the flow rate was 1.0 mL min-1, elution was monitored at 200 nm. Validation in selected conditions showed that the chosen method is sensitive, selective, precise, and reproducible with a linear detector response for the determination of acrylamide. The limit of detection (LOD), and the limit of quantification (LOQ), were achieved at 0.41μg mL-1 and 1.25 μg mL-1respectively. The proposed method was also applied after validation to the most popular six brands of chips and French fries available in the Libyan market. Acrylamide was extracted by a simplified extraction method avoiding cleanup by solid-phase extraction (SPE), then analyzed by HPLC-UV. The highest level of acrylamide was found in one brand of chips with a concentration of 16.33 μg mL-1, whereas only one of the French fries products analyzed exhibited an acrylamide concentration of 10.26 μg mL-1.</jats:p

Tackling the forensic analytical challenges associated with the global increase in seizures of cathinone-derived "legal highs"

The increase in the global abuse of synthetic cathinones has given rise to significant legal and analytical challenges in their identification and quantification - thus rapid methods of testing (especially in the field) are urgently required. This paper presents synthesis; characterisation; validated presumptive and quantitative methods for these substances (both in pure and adulterated samples) and a rapid, novel NMR screening technique for street samples containing components which cannot normally be detected using standard chromatographic methods