Metabolite Profiling and Structural Elucidation

Biotransformation describes the conversion of a compound to a metabolite through the action of enzymes while structural elucidation is the process of determining the chemical structure of those metabolites of interest. Charles River offers a comprehensive metabolite profiling and structural elucidation service for the profiling, isolation, analysis, and identification of metabolites in biological matrices in support of discovery and safety studies.

Our team of scientists can design bespoke experiments to overcome unique and complex analytical challenges across the pharmaceutical, agrochemical, and veterinary industries while working hand-in-hand to ensure timely delivery of data packages and detailed scientific reports that meet the relevant regulatory requirements. Our flexible, custom metabolite profiling and structural elucidation solutions are designed to meet your needs, available as part of a wider program of work or as stand-alone studies.

The metabolite profiling and structural elucidation process is an iterative one that aids in the prediction of human in vivo metabolites throughout the drug discovery and development process. Data from nonclinical in vitro and in vivo models informs lead selection, optimization decisions and helps design safety and clinical studies.

Study Types

In early discovery, understanding the fate of metabolized compounds is important for compound design, as it identifies metabolic hot-spots within a family of chemicals and supports the refinement of structure to afford more favorable physicochemical and pharmacokinetic properties, not limited to only metabolic stability. For adequate safety assessment, it is important to ensure coverage of human metabolites in the appropriate toxicology species, and to understand any toxic effects as early as possible. Gathering metabolic profiling and structural elucidation data earlier in the process delivers valuable insights for any development program.

For agrochemicals and veterinary medicines, the situation can be complicated by a secondary metabolism effect. This happens when a compound is first metabolized by the intended primary species (e.g., a treatment for livestock ailment or plant protection product) and later metabolized again in humans, as many of these commodities find their way into the food chain. As a result, it is critical to understand the nature, identity, and subsequent concentration of these metabolites, i.e., metabolite profiling and structural elucidation, for ongoing safety assessment and continued use of the product.

Sample Analysis

The application of high-resolution LC-MS and LC-MS/MS for metabolite identification remain the primary tools in industry. These analytical techniques afford the sensitivity and precision required to observe very low concentration analytes of interest and the ability to discern them from unrelated and abundant naturally occurring compounds. By combining robust accurate mass, extreme resolution, and fragmentation with separation techniques, mass spectrometers provide superior metabolite profiling and structural information.

Using a suite of state-of-the-art instruments that deliver unrivalled sensitivity and resolution and an array of powerful new experiments, we can produce highly detailed metabolite structural data and quantify very low metabolite concentration analytes.

Using Liquid Chromatography High-Resolution Mass Spectrometry for Metabolite Profiling

Using Liquid Chromatography High-Resolution Mass Spectrometry for Metabolite Profiling

Identify your metabolites, as active metabolites can interact with other targets and produce undesirable effects.
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Data Mining and Interpretation

Deciphering structural information for complex biological samples requires not only sophisticated instrumentation, but software that can process the chromatographic and mass spectral data to differentiate parent and metabolized compounds from endogenous material. This is even more challenging with low concentration samples. Our team of scientists use both manual screening techniques and Thermo Scientific’s Compound Discoverer software to find and elucidate the structures of degradates, hydrolysates, and metabolites across a diverse array of sample types.

Structure Elucidation

Structure elucidation is the process of using all the available chromatographic and spectral data to identify a component of interest. Data such as retention time, isotopic pattern, accurate mass measurement, and product ion fragmentation are routinely used to reinforce the identity of the proposed structure. Where available, components can be confirmed against authentic reference standards to afford increased structural certainty.

Offering further support for your program, our synthetic chemistry service can perform synthesis of metabolites, reference standards, and internal standards to assist with method development and metabolite identification.

    • Radio profiling
    • Degradates
    • Hydrolysates
    • Metabolites
    • (U)HPLC including 2D chromatography
    • Sample concentration, matrix and spectral clean-up
    • Chiral resolution of enantiomeric analytes
    • High resolution accurate mass LC-MSn
    • Thermo Orbitrap Fusion and Fusion LUMOS Tribrid MS
    • Thermo Scientific Q-Exactive™ and Q-Exactive™ Plus MS
    • Waters Synapt G2-S
    • GC and GC-MS
    • The latest metabolism software packages
    • Radiochemical repurification service
    • Use of in vitro synthetic models for biosynthesis of ‘awkward’ metabolites
    • High field multinuclear NMR with cryoprobe for absolute structure determination

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Frequently Asked Questions (FAQs) for Metabolite profiling

  • What is metabolite identification and profiling?

    The conversion of a parent compound to a metabolite or multiple metabolites through the action of biological enzymes can cause undesirable biological effects. Understanding this conversion can aid the anticipation of safety concerns arising from disproportionate toxic metabolites, predicting drug-drug interactions, and understanding routes of elimination.

    Metabolite identification can be classified in three steps:

    1. Metabolite profiling, the process of finding compound-related metabolites in a complex biological sample
    2. Metabolite quantitation, the determination of abundance.
    3. Metabolite characterization, the structural elucidation, and assessment of (bio)transformation pathway
  • What is metabolite profiling vs metabolomics profiling?

    Metabolite profiling measures the conversion of an applied compound of interest (test item) to its metabolites and intermediates in biological samples as a result of the test system, e.g., enzymes, hydrolysis, genetic modification, and physiological pathways. Metabolomics takes a broader view and examines what a test item or compound of interest does to the wider biological system and the complete set of metabolites in cells, tissues and organs. It is the study of what is left behind as a result of the cellular process and state of the biological system.

  • What are the best techniques for metabolite profiling and structural elucidation?

    Mass spectrometry (MS), particularly high-resolution mass spectrometry (HRMS) and nuclear magnetic resonance (NMR) spectroscopy are the most powerful analytical methods for metabolite identification, profiling and structural elucidation. By combining these techniques, our scientists can optimize sensitivity, resolution, and absolute structure determination. These techniques can detect individual molecular species with short acquisition times, though they require careful sample preparation, skilled analysis & and data mining, and experienced interpretation of the resulting datasets. If working with radiolabels, then LC-radioactivity detection is an option as well.