Derivatization strategy combined with parallel reaction monitoring for the characterization of short-chain fatty acids and their hydroxylated derivatives in mouse

Short-chain fatty acids (SCFAs) and hydroxylated short-chain fatty acids (OH–SCFAs) are crucial intermediates related to a variety of diseases, such as bowel disease, cardiovascular disease, renal disease and cancer. A global profiling method to screen SCFAs and OH–SCFAs was developed by tagging these analytes with d₀/d₆-N, N-dimethyl-6,7-dihydro-5H-pyrrolo [3,4-d] pyrimidine-2-amine (d₀/d₆-DHPP) and using ultra-high performance liquid chromatography coupled with high-resolution tandem mass spectrometry (UHPLC-MS/MS) in parallel reaction monitoring (PRM) mode. The derivatization procedure was simple and rapid. The targeted compounds could be derivatized within 3 min under mild condition and analyzed without the need of further purification. The derivatization significantly improved the chromatographic performance and mass spectrometry response. The d₆-DHPP tagged standards were used as internal standards, which remarkably reduced the matrix effects. The use of high resolution PRM mode made it possible to locate unknown SCFA and OH-SCFA species, and greatly reduced the false positive identification results. The developed method was successfully applied to the analysis of mouse fecal, serum, and liver tissue samples harvested from the breast cancer nude mice that had been exposed with 2,2′,4,4′-tetrabromodiphenyl ether (BDE-47). Results showed that 40 analytes (10 SCFAs and 30 OH–SCFAs) were characterized. Semi-quantitative analysis indicated that the exposure of BDE-47 to the mice altered the SCFA and OH-SCFA metabolism, especially in the high dose group. This study provides a high-throughput method to characterize SCFAs and OH–SCFAs in mouse samples.