Rapid Quantification of Gut Microbial Short-Chain Fatty Acids by pDART-MS

Abstract

Short-chain fatty acids (SCFAs) are small molecules ubiquitous in nature. In mammalian guts, SCFAs are mostly produced by anaerobic intestinal microbiota through the fermentation of dietary fiber. Levels of microbe-derived SCFAs are closely relevant to human health status and indicative to gut microbiota dysbiosis. However, the quantification of SCFA using conventional chromatographic approaches is often time consuming, thus limiting high-throughput screening tests. Herein, we established a novel method to quantify SCFAs by coupling amidation derivatization of SCFAs with paper-loaded direct analysis in real time mass spectrometry (pDART-MS). Remarkably, SCFAs of a biological sample were quantitatively determined within a minute using the pDART-MS platform, which showed a limit of detection at the μM level. This platform was applied to quantify SCFAs in various biological samples, including feces from stressed rats, sera of patients with kidney disease, and fermentation products of metabolically engineered cyanobacteria. Significant differences in SCFA levels between different groups of biological practices were promptly revealed and evaluated. As there is a burgeoning demand for the analysis of SCFAs due to an increasing academic interest of gut microbiota and its metabolism, this newly developed platform will be of great potential in biological and clinical sciences as well as in industrial quality control.

Isotope-Coded Chemical Derivatization Method for Highly Accurately and Sensitively Quantifying Short-Chain Fatty Acids

“…The neonatal period is a crucial window phase in developing microbial colonies, which have profound influences on piglet health. During this period, the gut microbiota and the produced SCFAs undergo wide dynamic change. − Figure shows that the most abundant SCFAs in the cecum digesta was acetic acid, followed by propanoic acid and butyric acid, which demonstrated a trend consistent with previous studies . Since DMAQ derivatization significantly improved SCFA separation, 2-methylbutyric acid and 3-methylbutyric acid could be separately quantitated in cecum digesta and plasma (Figures B and A).…”

“…24−27 Figure 4 shows that the most abundant SCFAs in the cecum digesta was acetic acid, followed by propanoic acid and butyric acid, which demonstrated a trend consistent with previous studies. 28 Since DMAQ derivatization significantly improved SCFA separation, 2-methylbutyric acid and 3-methylbutyric acid could be separately quantitated in cecum digesta and plasma (Figures 4B and 5A). Comparing the changes in SCFAs during the neonatal period, it was found that there was an increase in the concentrations of most SCFAs and total SCFAs ranging from day 0 to 7 after birth and peaked on day 14.…”

Oral short-chain fatty acids administration regulates innate anxiety in adult microbiome-depleted mice

“…The analytical sample was prepared according to the method previously described (Weng et al, 2020). 25 mg of brain and spinal cord tissue was extracted with 500uL 70% MeOH and vortexed with beads (1.0 mm dia.…”

“…The sample was reconstituted with 25 μL MeOH. For mass analysis, each brain and spinal cord sample (2 μL) was injected into a vanquish ultra-high-performance liquid chromatography (UHPLC) system coupled with Thermo Orbitrap Elite TM (Weng et al, 2020).…”

A prebiotic diet modulates microglial states and motor deficits in α-synuclein overexpressing mice

“…The flow rate was 0.35 mL/min with a linear gradient elution over 15 min. Reagent 1: Pyridine (Sigma-Aldrich) was adjusted with methanol to 3% (V/V) ( Weng et al, 2020 ).…”