Abstract: A pre-column derivatization high performance liquid chromatography (PCD-HPLC) method was developed for simultaneous analysis of short-chain fatty acids (SCFAs) and lactic acid in complex biological samples including intestinal contents and serum. The contents of SCFAs and lactate in mouse feces were determined by PCD-HPLC and compared with those determined by gas chromatography (GC) and high performance liquid chromatography (HPLC). Then, methodological evaluation was performed on PCD-HPLC. PCD-HPLC was applied to analyze intestinal contents and blood. Results showed that GC could rapidly detect acetate, propionate, butyrate and valerate, but not lactic acid. HPLC yielded many impurity peaks and inaccurate results for lactic acid. The PCD-HPLC method was applicable to a wide range of samples and allowed for simultaneous analysis of lactate and five SCFAs in intestinal contents and serum. Only a few impurity peaks appeared in the chromatogram, and a good separation of all analytes was achieved. PCD-HPLC was accurate, giving a coefficient of variation (CV) less than or equal to 8.87% for each analyte. In conclusion, this method is superior to GC and HPLC in determining SCFAs and lactic acid in complicated biological samples such as intestinal contents and serum, and can meet the demand for accurate analysis of SCFAs and lactic acid in complicated biological samples in response to the rapid development of microbial metabolomics.

Key words: short-chain fatty acids; lactic acid; gas chromatography; high performance liquid chromatography; pre-column derivatization; feces; cecal contents; serum