Thesis

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

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Awarding institution
  • University of Strathclyde
Date of award
  • 2013
Thesis identifier
  • T13485
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Abstract
  • 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.
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DOI
Date Created
  • 2013
Former identifier
  • 991433

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