Standardization of Fecal Metabolomics Using Microbiome Preservation Kits: Implications for Multiomics Integration.

Area of Science:

• Human Health and Nutrition
• Metabolomics
• Multiomics

Background:

• Integrative omics research is growing in human health and nutrition.
• Optimal storage and preprocessing of labile fecal samples for metabolomics remain challenging.

Purpose of the Study:

• To systematically evaluate three normalization methods (wet weight, dry weight, protein quantification) for quantitative fecal metabolomic profiling.
• To assess the reproducibility of fecal metabolite profiling using the OMNIgene·GUT kit.

Main Methods:

• Evaluation of wet weight, dry weight, and protein quantification normalization methods.
• Analysis of 41 metabolites in fresh fecal samples from healthy individuals and obese patients collected via the OMNIgene·GUT kit.
• Assessment of metabolite reproducibility using coefficient of variation (CV < 30).

Main Results:

• Wet and dry weight normalization showed high reproducibility in fresh samples (24 metabolites, CV < 30).
• OMNIgene·GUT kit samples showed improved reproducibility with wet weight (20 metabolites, CV < 30) and protein quantification (19 metabolites, CV < 30) normalization.
• Dry weight normalization failed to meet the CV < 30 criterion for obese patient samples.
• Direct analysis without drying enhanced chromatographic clarity.

Conclusions:

• Wet weight normalization offers a robust and standardized framework for fecal metabolite profiling, consistently minimizing variation.
• The OMNIgene·GUT kit is compatible with broad-spectrum metabolomic analyses and suitable for multiomics workflows.
• Reproducible normalization protocols are crucial for accurate, comparable, and scalable fecal metabolomics in clinical and nutritional research.