Metabolite Identification

Metabolite Identification

At XenoGesis we have two highly skilled people in metabolite identification with over 40 years’ combined experience. Metabolites are identified using their skill and experience to process the data manually on a scan by scan basis without the need to use metabolite identification software packages which, depending on how the software is configured, can miss the more unusual metabolites and generate a large number of possible metabolites that, on closer interrogation, are actually not related.

As well as providing a number of options for metabolite profiling in support of projects in Discovery, we have also provided support for clinical metabolite profiling studies in man by creating a single AUC0-24h pooled blood or plasma sample for identification of metabolites in support of the MIST guidelines.

We can offer a range of tailored services in order to meet your needs. This can be identification of the major metabolic route, cross species hepatocyte comparisons for identification of appropriate species for safety studies or animal In vivo studies at the later stages of the drug discovery process for potential drug candidates. We offer different reports depending on the level of detail required from a simple table of proposed structures based on the MS/MS data to provision of extracted ion chromatograms, MS and MS/MS spectra with interpretation of the data for each metabolite.

With the recent acquisition of our Thermo Scientific™ Q Exactive™ Focus Hybrid Quadrupole-Orbitrap™ mass spectrometer all metabolite identification work will be carried out on this instrument providing accurate mass data.

A number of suggested metabolite profiling packages are outlined below, which can easily be tailored to meet individual requirements.

Understanding the fate of highly metabolised research compounds is especially important in the design phase of drug discovery, as identifying the metabolic hot-spots within a chemical series is key to allow iterative chemical modifications to design compounds with improved metabolic stability. However, at the later stages of the discovery phase where potential drug candidates have been identified, it is important to ensure that there is coverage of the human metabolites in rodent and non-rodent species used for the safety studies.

For metabolite identification to be influential in the design of metabolically stable drug candidates a fast turnaround time is essential. Once the request has been made, the samples must be created, analysed and reported within a matter of days in order to be able to influence the next design cycle. To facilitate this, samples are analysed using a generic LC method with only the major metabolites being analysed by MS/MS and then reported in a simple table with the proposed structures.

For more detailed studies, such as cross species hepatocytes, it is necessary to determine the full metabolic profile to ensure there is coverage for the human metabolites in the proposed species for the safety studies. This can identify early on, any issues with human specific or disproportionate metabolites that need to be addressed.

The example below illustrates that dextromethorphan can undergo N- and O-demethylation. As both metabolites have an identical elemental formula, using the MS/MS fragmentation pattern is the only way to distinguish between both metabolites.

 

 

 

Full scan ESI spectra for Dextromethorphan and its mono- and di-de-methylated metabolites with a mass accuracy of 1ppm or less run on the Thermo Scientific Q-Exactive Focus obtained from a 60 minute rat hepatocyte incubation.
Product ion mass spectra for Dextromethorphan and its mono- and di-de-methylated metabolites acquired using variable Data Independent Acquisition mode allowing MS and MS/MS spectra to be acquired for parent and metabolites in one run with excellent mass accuracy for the fragments.