Analysis of short chain fatty acids in faecal samples : development and validation of a new method

Sammanfattning: Short chain fatty acids (SCFA) are produced by the gut microflora following the intake of complex carbohydrates and have been suggested as a contributor for some of the health benefits associated with dietary fibre (DF) intake. For further research of the mechanisms and effects of SCFA, reliable and accurate methods for analysis of SCFA in different biological samples are needed. In this project the aim was to develop and validate a rapid and ease-of use method for analysis of SCFA content in faecal samples. The contents of acetic, propionic, iso-butyric, n-butyric, iso-valeric, n-valeric and capronic acid were analysed through a method where fecal water was obtained through centrifugation and SCFA were further extracted with propyl formate. Extracts were analysed on a gas chromatograph coupled with a flame-ionization detector (GC-FID). The method was validated including estimation of accuracy and precision which were measured through calculation of recovery, deviation from linearity and by the coefficient of variance (CV), respectively. An exploratory test was also performed to analyse the distribution of SCFA in faecal matter. Moreover, the method was used to analyse SCFA concentrations in faecal samples (n=40) from two Norwegian bowel cancer projects, NORCCAP and BCSN. The results of the validation (recovery, CV and deviation from linearity) varied somewhat between the three batches, and thereby also the limits of quantification (LOQ), which were limited, but in most cases included the physiologically relevant concentrations. Through the exploratory distribution test, a significant difference in SCFA content was found between the in- and outside of the faecal sample, which emphasises the importance of homogenization of samples or consistent sampling from faecal samples. SCFA contents of faecal samples from the NORCCAP and BCSN samples were consistent with those reported by other studies, which indicate that the method produces reliable results. For a higher accuracy and precision, it is suggested that the amount of faecal material analysed is increased, followed by further validation of both lower and higher concentrations. Additionally, the effects of storage with and without buffer should also be investigated for more comparable results.