Abstract
Gut microbiota plays essential roles in maintaining gut homeostasis. The composition of gut microbes and their metabolites are altered in response to diet and remedial agents such as antibiotics. However, little is known about the effect of antibiotics on the gut microbiota and their volatile metabolites. In this study, we evaluated the impact of a moderate level of ampicillin treatment on volatile fatty acids (VFAs) of gut microbial cultures using an optimized real-time secondary electrospray ionization coupled with high-resolution mass spectrometry (SESI-HRMS). To evaluate the ionization efficiency, different types of electrospray solvents and concentrations of formic acid as an additive (0.01, 0.05, and 0.1%, v/v) were tested using VFAs standard mixture (C2–C7). As a result, the maximum SESI-HRMS signals of all studied m/z values were observed from water with 0.01% formic acid than those from the aqueous methanolic solutions. Optimal temperatures of sample inlet and ion chamber were set at 130 °C and 85 °C, respectively. SESI spray pressure at 0.5 bar generated the maximum intensity than other tested values. The optimized SESI-HRMS was then used for the analysis of VFAs in gut microbial cultures. We detected that the significantly elevated C4 and C7 VFAs in the headspace of gut microbial cultures six hours after ampicillin treatment (1 mg/L). In conclusion, our results suggested that the optimized SESI-HRMS method can be suitable for the analysis of VFAs from gut microbes in a rapid, sensitive, and non-invasive manner.
Highlights
Over the past two decades, the mass spectrometry-based methodologies such as proton-transfer-reaction mass spectrometry (PTR-MS) and selected ion flow tube mass spectrometry (SIFT-MS), in addition to the gold-standard gas chromatography-mass spectrometry (GC-MS), have been used for the detection and analysis of gas-phase volatile organic compounds (VOCs) [1,2,3,4]
To optimize Secondary Electrospray Ionization (SESI) parameters, 50 μmol/L of fresh standards of a volatile free acid mixture consisting of acetic acid (C2 -volatile fatty acids (VFAs)), propionic acid (C3 -VFA), isobutyric acid/butyric acid (C4 -VFA), isovaleric acid/valeric acid (C5 -VFA), isocaproic acid/hexanoic acid (C6 -VFA), and heptanoic acid (C7 -VFA) were diluted in deionized water and used as the source of volatiles for SESI-high-resolution mass spectrometry (HRMS) detection
Our findings indicate that the optimization of SESI parameters may be pre-required to obtain the maximum and accurate intensity based on our observation that as the five-times higher signal intensity can be obtained after systematic optimization in comparison to the results from the least optimized condition
Summary
Over the past two decades, the mass spectrometry-based methodologies such as proton-transfer-reaction mass spectrometry (PTR-MS) and selected ion flow tube mass spectrometry (SIFT-MS), in addition to the gold-standard gas chromatography-mass spectrometry (GC-MS), have been used for the detection and analysis of gas-phase volatile organic compounds (VOCs) [1,2,3,4]. SESI coupling to high-resolution mass spectrometry (HRMS) has been shown to enhance the capability to capture as many VOC metabolites as possible in real-time mass spectrometric analysis [10]. SESI was invented as a variant of the electrospray ionization source (ESI) that can be utilized for gaseous analytes [11,12]. It is an ambient electrospray ionization type that can produce charging agents from evaporating nano-droplets/ions to charge neutral vapor molecules in the gas phase [13,14]. While efforts have been made to understand the exact ionization mechanism of SESI, the impact of some major SESI parameters on its signal intensities and ionization efficiency remains to be fully defined [14]
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