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

Short-chain fatty acids (SCFAs) are major gut microbiota-derived metabolites, which can reshape the intestine andregulate gut immunity. The application of conventional GC methods has been hampered for quantifying low-concentrated SCFAs,such as in serum, saliva, and digesta of germ-free animals. Herein, we established a LC-MS method to quantify SCFAs after 5-(dimethylamino)-1-carbohydrazide-isoquinoline (DMAQ) derivatization. The DMAQ derivatization significantly enhanced thedetection sensitivity and improved separation of SCFAs. 2-methylbutyric acid and 3-methylbutyric acid were separately quantitated.Moreover, the matrix effect was diminished using DMAQ-13C/15N-tagged SCFAs as internal standards. The established quantitationmethod was successfully applied in the analysis of plasma and cecum digesta collected from neonatal piglets, revealing that significantincreases in biological SCFA contents in cecum digesta were closely related to the variation of gut microbial diversity. Theestablished quantitation method is capable of sensitively and comprehensively quantifying SCFAs that may provide insights intounderlying gut-microbiota functions

Automated summary

Short-chain fatty acids (SCFAs) are important metabolites produced by gut microbiota, playing a crucial role in intestinal and immune regulation. A LC-MS method was developed to quantify low-concentrated SCFAs, and successfully applied in analyzing plasma and cecum digesta of neonatal piglets, revealing their relationship with gut microbial diversity.