Determination of six short‐chain fatty acids in rat feces using headspace solid‐phase dynamic extraction coupled with GC‐MS

Abstract

Short‐chain fatty acids (SCFAs) are organic acids with carbon atoms less than six, released through fermentation products by intestinal microbiome, having multiple physiological activities. Considering weak acidity and high volatility, derivatization or liquid–liquid extraction is essential, which is time consuming. Headspace‐solid‐phase dynamic extraction (HS‐SPDE) coupled with gas chromatography‐mass spectrometry is automated and effortless to determine SCFAs in rat feces. The extraction procedure is performed by aspirating and discharging the headspace cyclically through a steel needle, coated with an inner polyethylene glycol sorbent. The key parameters of SPDE were optimized including coating type, incubation time and temperature, and number of extraction strokes. Besides, salting‐out was conducted. Then, a method by HS‐SPDE‐GC‐MS was established and validated. It only took 3‐min incubation time, 4.5 min extraction time, and 13 min chromatographic separation in a run. The recovery, linearity, limit of quantification, and stability were evaluated. Then, the proposed method was applied to analyze rat feces including 18 rats with liver injury and 23 normal controls. Mann–Whitney U test indicated that the concentrations of six SCFAs in normal rat feces were higher than those with liver injury. This method provides a choice for fast, solvent‐free, automated, and high‐throughput analysis of SCFAs.