Adapting a commercial sample extraction protocol for biosafety level 3/4 compatible plasma metabolomics analysis.
INTRODUCTION: Severe infections and sepsis significantly impact military operational readiness and costs through loss of duty days, high treatment rates, and medical evacuations. Early diagnosis is critical for preventing sepsis progression and mortality, but it requires validated biomarkers to guide clinical decision-making. Study protocols for host biomarker discovery in infections usually require inactivation of high-risk pathogens prior to sample analysis, which limits the utility of metabolomics assays designed for untreated samples. METHODS: Matched blood plasma aliquots obtained from an international, observational sepsis cohort were analyzed to quantify metabolites following the commercial AbsoluteIDQ p180 protocol, with and without the addition of an organic solvent extraction method for metabolites, proteins, and lipids (MPLEx), previously validated for inactivating BSL-3/4 pathogens. We evaluated analyte detection rates and concentrations for each method, as well as differences in extraction efficiency. RESULTS: Levels of agreement between the unmodified AbsoluteIDQ p180 and combined MPLEx-p180 methods varied by metabolite class. Most targeted amino acids, glycerophospholipids, sphingolipids, and monosaccharides were reliably measured and correlated well between methods. However, the higher sample dilution in the MPLEx-p180 method significantly reduced detection rates for biogenic amines and acylcarnitines, and overall extraction efficiencies also differed. CONCLUSIONS: This study extends the applicability of commercial metabolomics assays designed for untreated samples by improving their suitability for high-risk infectious disease studies. Differences in metabolite extraction efficiencies and detection rates, as well as data harmonization strategies, should be considered if results from both protocols are to be combined.
