In Silico and in Vitro Metabolism Studies Support Identification of Designer Drugs in Human Urine by Liquid Chromatography/quadrupole-Time-of-Flight Mass Spectrometry.

In silico and in vitro metabolism studies support identification of designer drugs in human urine by liquid chromatography/quadrupole-time-of-flight mass spectrometry.

Anal Bioanal Chem. 2013 Jun 25;
Tyrkkö E, Pelander A, Ketola RA, Ojanperä I

Human phase I metabolism of four designer drugs, 2-desoxypipradrol (2-DPMP), 3,4-dimethylmethcathinone (3,4-DMMC), ?-pyrrolidinovalerophenone (?-PVP), and methiopropamine (MPA), was studied using in silico and in vitro metabolite prediction. The metabolites were identified in drug abusers’ urine samples using liquid chromatography/quadrupole-time-of-flight mass spectrometry (LC/Q-TOF/MS). The aim of the study was to evaluate the ability of the in silico and in vitro methods to generate the main urinary metabolites found in vivo. Meteor 14.0.0 software (Lhasa Limited) was used for in silico metabolite prediction, and in vitro metabolites were produced in human liver microsomes (HLMs). 2-DPMP was metabolized by hydroxylation, dehydrogenation, and oxidation, resulting in six phase I metabolites. Six metabolites were identified for 3,4-DMMC formed via N-demethylation, reduction, hydroxylation, and oxidation reactions. ?-PVP was found to undergo reduction, hydroxylation, dehydrogenation, and oxidation reactions, as well as degradation of the pyrrolidine ring, and seven phase I metabolites were identified. For MPA, the nor-MPA metabolite was detected. Meteor software predicted the main human urinary phase I metabolites of 3,4-DMMC, ?-PVP, and MPA and two of the four main metabolites of 2-DPMP. It assisted in the identification of the previously unreported metabolic reactions for ?-PVP. Eight of the 12 most abundant in vivo phase I metabolites were detected in the in vitro HLM experiments. In vitro tests serve as material for exploitation of in silico data when an authentic urine sample is not available. In silico and in vitro designer drug metabolism studies with LC/Q-TOF/MS produced sufficient metabolic information to support identification of the parent compound in vivo. HubMed – drug


Mifepristone treatment affects the response to repeated amphetamine injections, but does not attenuate the expression of sensitization.

Psychopharmacology (Berl). 2013 Jun 25;
van der Veen R, Boshuizen MC, de Kloet ER

Glucocorticoid hormones facilitate sensitization to repeated administration of psychostimulants, an effect that is mediated by glucocorticoid receptors (GRs). It is still unclear, however, at which stage of psychomotor sensitization are stress and GR-mediated effects involved.In the present study, we have tested the hypothesis that GR-mediated effects during the phase of repeated amphetamine injections play a crucial role in the long-term expression of sensitization. For this purpose, we used DBA/2 mice, an inbred strain commonly used for the study of stress effects on psychostimulant sensitization.Animals were treated with the GR antagonist mifepristone (200 mg/kg) at 2.5 h before each daily injection of amphetamine (2.5 mg/kg) or saline in a 5-day protocol. The amphetamine or saline injections were given in the home or a novel context. This was followed by a 2.5-week withdrawal period, without any drug delivery. Following the withdrawal period, two low-dose amphetamine challenges (1.25 mg/kg) were given subsequently, without additional mifepristone.The animals receiving amphetamine in the novel context showed a higher expression of sensitization at challenge as compared to those in the home condition. Mifepristone treatment influenced locomotor response to repeated amphetamine injections, but this effect during the initial phase did not affect the expression of sensitization after a withdrawal period.Our results indicate that GR-related processes during the initial phase of sensitization are involved in, but not crucial for, the development of long-term sensitization. HubMed – drug


Extending the Dynamic Range of the Ion Trap by Differential Mobility Filtration.

J Am Soc Mass Spectrom. 2013 Jun 25;
Hall AB, Coy SL, Kafle A, Glick J, Nazarov E, Vouros P

A miniature, planar, differential ion mobility spectrometer (DMS) was interfaced to an LCQ classic ion trap to conduct selective ion filtration prior to mass analysis in order to extend the dynamic range of the trap. Space charge effects are known to limit the functional ion storage capacity of ion trap mass analyzers and this, in turn, can affect the quality of the mass spectral data generated. This problem is further exacerbated in the analysis of mixtures where the indiscriminate introduction of matrix ions results in premature trap saturation with non-targeted species, thereby reducing the number of parent ions that may be used to conduct MS/MS experiments for quantitation or other diagnostic studies. We show that conducting differential mobility-based separations prior to mass analysis allows the isolation of targeted analytes from electrosprayed mixtures preventing the indiscriminate introduction of matrix ions and premature trap saturation with analytically unrelated species. Coupling these two analytical techniques is shown to enhance the detection of a targeted drug metabolite from a biological matrix. In its capacity as a selective ion filter, the DMS can improve the analytical performance of analyzers such as quadrupole (3D or linear) and ion cyclotron resonance (FT-ICR) ion traps that depend on ion accumulation. HubMed – drug


An LC-MS based study of the metabolic profile of primaquine, an 8-aminoquinoline antiparasitic drug, with an in vitro primary human hepatocyte culture model.

Eur J Drug Metab Pharmacokinet. 2013 Jun 25;
Jin X, Pybus BS, Marcsisin SR, Logan T, Luong TL, Sousa J, Matlock N, Collazo V, Asher C, Carroll D, Olmeda R, Walker LA, Kozar MP, Melendez V

The 8-aminoquinoline drug primaquine (PQ) is currently the only drug in use against the persistent malaria caused by the hypnozoite-forming strains P. vivax and P. ovale. However, despite decades of research, its complete metabolic profile is still poorly understood. In the present study, the metabolism of PQ was evaluated by incubating the drug with pooled human hepatocytes cultured in vitro as well as with recombinant cytochrome P450 (CYP) isoenzymes, monoamine oxidases (MAO), and flavin-containing monooxygenases (FMO). Targeted LC-MS/MS analysis of hepatocyte incubations using chemical inhibitors indicated that PQ was predominantly metabolized by CYPs 3A4, 1A2 and 2D6, MAO-A, -B and FMO-3. Confirmation of these results was sought by incubation of PQ with the corresponding recombinant enzymes. Small amounts of carboxyprimaquine (CPQ), the major observed PQ metabolite in vivo, were detected in recombinant MAO-A incubations along with another peak at m/z 261, and no significant formation of CPQ with any other recombinant enzymes was observed. Incubations with all recombinant enzymes identified as potentially active towards PQ from the hepatocyte-based assay resulted in significant parent loss over the course of 1 h. These results suggest that several enzymes, including CYPs in combination with FMOs and MAOs, play a role in the overall metabolism of PQ and indicate a major role for MAO-A. Future studies to elucidate the potential role in cytotoxicity and/or efficacy of the PQ metabolite observed at m/z 261, as observed in MAO-A isoenzyme studies, are needed. HubMed – drug



Psychology Career Profile: University of Waikato – Rickardt van Dyk studied Psychology at the University of Waikato and is now working as a Service Coordinator at Community Living, working with people who hav…