High‐resolution gas chromatography/mass spectrometry metabolomics of non‐human primate serum

Abstract

Our study provides insights into the benefits and limitations of the use of a higher mass resolution and mass accuracy instrument for untargeted GC/MS-based metabolomics with multi-dimensional chromatography in future studies addressing clinical conditions or exposome studies.

Comparison of a GC-Orbitrap-MS with Parallel GC-FID Capabilities for Metabolomics of Human Serum

“…These metabolites included S-adenosyl-L-methionine, adenosine monophosphate, S-adenosyl-L-homocysteine, glucose, alanine, lysine, formic acid, arginine, serine, tryptophan, phenylalanine, urea, 5-phosphorylribose 1-pyrophosphate, biotin, histidine, proline, citric acid, benzoic acid, valine, and threonine. The significantly higher number of metabolites detected in our current efforts, compared to our earlier analysis of a baboon serum sample (7) is attributed to a longer run time of 60 minutes as opposed to the shorter protocol of 23 minutes in the earlier study. Moreover, there are species specific metabolite differences among primate tissues (32) and biofluids.…”

“…We previously reported on the analysis of non-human primate serum from a baboon using HR-GC-MS alone (7). Here, we expanded our metabolomics analysis to human serum, and compared two orthogonal detection techniques attached to a GC, a MS and a FID detector.…”

“…Blanks were intermittently used to see that no carryovers occurred during randomized run orders and to manually filter out contaminating chemicals from the combined list of features obtained from the blanks. Samples were then sequentially derivatized with methoxyamine hydrochloride (MeOX) and 1% TMCS in N -methyl- N -trimethylsilyl-trifluoroacetamide (MSTFA) as described elsewhere (7, 18, 19). Steps involved addition of 10 μL of MeOX (20 mg/mL) in pyridine, followed by incubation under shaking at 55 °C for 60 min followed by trimethylsilylation at 60 °C for 60 min after adding 90 μL MSTFA as described (2, 7).…”

“…Particularly, gas chromatography mass spectrometry (GC-MS) is very amenable to polar primary metabolites (such as sugars, amino acids, amines, sugar phosphates, or sugar alcohols) (2) and fatty acids, in addition to excellent chromatographic resolution, thus lending itself to routine quantitative metabolomic applications (2). Newer high resolution (HR) instruments such as Orbitrap mass spectrometers are capable of providing sub-ppm mass accuracy at high mass resolutions (i.e., > 60,000), and hence allow calculation of predicted molecular formulas based on the mass defect of a detected metabolite ion (3–5), and generate mass spectral data at high resolving power with mass accuracies <1 ppm· However, these HRGC-MS platforms have found limited applications till date, baring handful applications in microbial metabolomics (6) and a recent use in non-human primate biofluid (i.e., baboon) serum metabolomics (7).…”

Drying enhances signal intensity for global GC-MS metabolomics

“…Due to the technical properties and its wide coverage of metabolites from central metabolic pathways to steroids and xenobiotic molecules, GC-MS metabolomics is an essential tool for the investigation of biological processes. Technical development is ongoing, with recent advances enabling the generation of ever more complex metabolic profiles containing 1000's of deconvoluted peaks per sample using the newest mass-spectrometry detectors 8,9 . Hardware advances aside, sample preparation approaches differ widely across sample types, as large divergence across different laboratories.…”