Denise Morrison

2-Aryl Indole NK1 receptor antagonists: optimisation of indole substitution

The synthesis and biological evaluation of a series of 2-aryl indoles with high affinity for the human neurokinin-1 (hNK1) receptor are reported, concentrating on optimisation of the indole substitution.

Approaches to higher-throughput pharmacokinetics (HTPK) in drug discovery.

With pressure on pharmaceutical companies to reduce time-to-market and improve the success rate of new drug candidates, higher-throughput pharmacokinetic (HTPK) support has become an integral part of many drug discovery programmes. This report details the amalgamation of robotics, new sample preparation technologies and highly sensitive and selective mass spectrometric detection systems to deliver the promise of HTPK. A historical perspective on automated bioanalysis with the current approaches and future prospects for the discipline are described.

Determination of the collisionally activated dissociation of a substituted indole by orthogonal acceleration quadrupole time-of-flight mass spectrometry ☆

The use of orthogonal acceleration quadrupole time-of-flight (Q-TOF) mass spectrometry to determine the collisionally activated dissociation (CAD) of a test compound 1-(3-[5-{1,2,4-triazol-4-yl}-1H-indol-3-yl]propyl)-4-(2-{3-fluorophenyl}ethyl)piperazine is described. At unitmass resolution the identity of many ions is ambiguous because of the complexity of the resulting product ion spectrum. Using the high resolution capabilities of the Q-TOF instrument, exact masses for each fragment were determined. These data were used to infer molecular formulas for each fragment through software interpretation and, by further applying chemical intuition, the majority of ions were fully assigned. Additionally, by utilizing in-source fragmentation at high cone voltage, analyses of second-generation products allowed derivation of a consistent sequential fragmentation pathway. This study clearly demonstrates the power of Q-TOF mass spectrometry to elucidate complex product ion spectra.

Determination of the in vitro metabolism of (+)- and (−)-epibatidine

The oxidative in vitro metabolism of epibatidine was investigated using liver microsomes from rat, dog, rhesus monkey and human. Analysis was performed using liquid chromatography-mass spectrometry (LC-MS) using both achiral and chiral stationary phases. Comparison of the metabolism of the (+)- and (-)-enantiomers revealed species differences in the extent of metabolism, with rhesus monkey>dog>rat=human. Furthermore, differences in the routes of metabolism for epibatidine enantiomers were also observed, with mass spectra consistent with hydroxylation of the azabicycle for (-)-epibatidine and with the formation of diastereomeric N-oxides for (+)-epibatidine being obtained. For chiral LC-MS, a volatile ion-pair reagent of heptafluorobutyric acid was used in place of pentanesulphonic acid with no deterioration in chiral selectivity. Analysis of the same samples by chiral LC-MS revealed no evidence for metabolic chiral interconversion and chiral analysis from a metabolic time course of racemic material revealed enantiomers to be metabolised to approximately the same extent. Such findings may be important particularly should epibatidine be investigated in non-rodent species.

Quantitative determination of L-775,606, a selective 5-hydroxytryptamine 1D agonist, in rat plasma using automated sample preparation and detection by liquid chromatography–tandem mass spectrometry

L-775,606 is under investigation as a selective 5-hydroxytryptamine 1D agonist for the treatment of migraine. A reliable and sensitive method for the analysis of L-775,606 in plasma was required in order to support preclinical evaluation of this compound. A semi-automated sample preparation method using the Beckman Biomek 2000 workstation to perform all liquid handling tasks has been established. The sample analysis was performed using HPLC-MS-MS with a cycle time of 3.5 min per sample. Intra- and inter-day assay accuracy and precision are excellent with a calibration range of 1-2000 ng/ml and a reproducible limit of quantification of 1 ng/ml.

Use of chiral liquid chromatography-tandem mass spectrometry to investigate the metabolism of racemic cholecystokinin-B antagonists

In an attempt to establish the enantiomeric specificity of metabolism for a series of racemic cholecystokinin-B receptor antagonists, chiral LC-MS-MS conditions were established using a Pirkle DNBL chiral stationary phase operating in the reversed-phase mode. Rat liver microsomal incubations of the compounds were analysed using these conditions and it was demonstrated that resolution of oxygenated and demethylated metabolites could be achieved. A single model compound was investigated in detail by obtaining product-ion spectra on all mono-oxygenated species in an attempt to correlate these and identify enantiomeric pairs of metabolites. In this example a lack of differentiation in the product ion spectra did not allow correlation but the results suggest that such an approach may still be viable for the chiral metabolic analysis of racemic material.

2-Aryl indole NK1 antagonists: optimisation of the amide substituent

The in vivo properties of a series of 2-arylindole NK(1) antagonists have been improved, by modification of the amide substituent. The 1-(2-methoxyphenyl)piperazine amide was identified as a major area of metabolism in the lead compound 1. Replacement of this amine moiety by a 4-benzyl-4-hydroxypiperidine resulted in a compound 18 with reduced clearance and improved central duration of action.