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Metabonomic Evaluation of Fecal Water Preparation Methods: The Effects of Ultracentrifugation

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Background: Fecal water provides information on the state of the gut microbiota and host physiology and is a common biofluid to study, particularly for metabonomic purposes. Despite the interest in fecal water, a method of preparation has yet to be standardized. Current methods of preparing fecal water usually involve an extraction protocol combined with ultracentrifugation and filtration steps. If metabonomic analysis is the final means of examining fecal water samples, consideration should be given to the potential impacts of these preparation steps on the metabolites observed.

Objective: This work examines metabonomic effects of ultracentrifugation during fecal water preparation, and provides an easily adaptable method based on considerations to metabonomic impacts of the procedure.

Methods: Both fecal water (prepared from human stool samples) as well as a controlled surrogate for fecal water samples (prepared from in vitro fecal microbial communities) were used to perform systematic evaluations of various preparation steps, and used metabonomic profiles to assess the protocols’ impact on the sample.

Results: 52 metabolites were observed and found that ultracentrifugation speed decreased metabolite concentrations by approximately -2% per 10000 rpm increase, with one exception. P-cresol increased by approximately +50% per 10000 rpm. Upon investigation of the potential sources of p-cresol, we found evidence that these were tyrosine-rich cell proteins which broke down upon ultracentrifugation.

Conclusion: Based on these findings, we suggest that ultracentrifugation is an effective means of preparing fecal water samples, with negligible impact on metabonomic analyses, though measurement of p-cresol concentrations should be treated with discretion.
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Keywords: Fecal water; MLR; metabolomics; metabonomics; microbiota; preparation; ultracentrifugation

Document Type: Research Article

Publication date: 01 April 2018

More about this publication?
  • Current Metabolomics publishes timely reviews, original research articles, thematic issues, and technical notes covering all aspects of the recent advancements and applications of metabolomics technology to systems biology, disease diagnosis, personalized medicine, and drug discovery. Original research articles and reviews in the following areas of metabolomics are of special interest to the readers of this journal:

    Application to humans, plants, microbes, cell cultures, tissues, organs, and biofluids
    Automation and high-throughput technology
    Bioinformatics
    Chemical and structural characterization of new metabolites
    Chemometrics, statistical analysis, and other approaches to data analysis
    Databases, including collections of reference spectra
    Disease biomarkers
    Drug discovery, efficacy, toxicity, and personalized medicine
    Integration of analytical techniques and/or various OMICS datasets
    Metabolic profiling and directed metabolite analysis
    New analytical experiments and techniques (NMR, MS, FTIR, etc)
    Novel sensors and instrumentation for detecting metabolites
    Sample preparation, extraction techniques and chromatography
    Software

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