METHODS article
Front. Cell. Infect. Microbiol.
Sec. Bacteria and Host
Volume 15 - 2025 | doi: 10.3389/fcimb.2025.1584487
This article is part of the Research TopicNavigating the Microbial Landscape: Integrating Mass Spectrometry Imaging and Other Multimodal Imaging Approaches for Spatially Resolved Microbial StudiesView all 3 articles
On-Tissue Derivatization for Mass Spectrometry Imaging Reveals the Distribution of Short Chain Fatty Acids in Murine Digestive Tract
Provisionally accepted
- 1Kobe University, Kobe, Japan
- 2Division of Dermatology, Department of Internal Medicine, Graduate School of Medicine, Kobe University, Kobe, Hyōgo, Japan
- 3Shimadzu Corporation, Kyoto, Kyōto, Japan
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Short-chain fatty acids (SCFAs), which are produced by microorganisms in the digestive tract of animals, play an important role in maintaining homeostasis in the host, including immune function. Different types of SCFAs are produced by different intestinal bacterial communities. However, visualizing their spatial distribution within tissue sections has been difficult. This is primarily due to the volatility of SCFAs, which makes detection challenging, even with matrix-assisted laser desorption/ionization (MALDI) mass spectrometry imaging (MSI) using an atmospheric pressure ion source. To address this issue, we minimized the volatility of SCFAs in fresh tissue sections. Then, we used N,N,N-trimethyl-2-(piperazin-1-yl)ethan-1-amine iodide (TMPA) and 1- ((dimethylamino)(dimethylimino)methyl)-1H-[1,2,3]triazolo[4,5-b]pyridine-3-oxide hexafluorophosphate (HATU) to chemically derivatize the carboxylic acid into a quaternary amine. This in situ derivatization enabled visualization of SCFAs using MALDI-MSI. In the cecum of mice, strong signals for butyrate and propionate were detected in areas with high bacterial density, as identified by hematoxylin staining. This indicates that these SCFAs are produced by bacteria. Anaerobic bacteria were cultured from the cecum of another individual raised under the same environment. Strain identification was performed using MALDI mass spectrometry of bacterial protein finger prints which confirmed the presence of bacteria that produce SCFAs. This approach, which combines minimizing volatility and in situ derivatization, provides a powerful tool for elucidating the spatial relationship between intestinal bacteria and metabolites including SCFAs.
Keywords: short-chain fatty acids, SCFAs, MALDI, MSI, TMPA, HATU
Received: 27 Feb 2025; Accepted: 01 Sep 2025.
Copyright: © 2025 Kubo, Nakagawa, Okamoto, Terashima and Kobayashi. This is an open-access article distributed under the terms of the Creative Commons Attribution License (CC BY). The use, distribution or reproduction in other forums is permitted, provided the original author(s) or licensor are credited and that the original publication in this journal is cited, in accordance with accepted academic practice. No use, distribution or reproduction is permitted which does not comply with these terms.
* Correspondence: Akiko Kubo, Kobe University, Kobe, Japan
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