The human fecal metabolome is increasingly studied to explore the impact of diet and lifestyle on health and the gut microbiome. However, systematic differences and confounding factors related to age, sex, and diet remain largely unknown. In this study, absolute concentrations of fecal metabolites from 205 healthy Danes (105 males and 100 females, 49 ± 31 years old) were quantified using 1H NMR spectroscopy and the newly developed SigMa software. The largest systemic variation was found to be highly related to age. Fecal concentrations of short-chain fatty acids (SCFA) were higher in the 18 years old group, while amino acids (AA) were higher in the elderly. Sex-related metabolic differences were weak but significant and mainly related to changes in SCFA. The concentrations of butyric, valeric, propionic, and isovaleric acids were found to be higher in males compared to females. Sex differences were associated with a stronger, possibly masking, effect from differential intake of macronutrients. Dietary fat intake decreased levels of SCFA and AA of both sexes, while carbohydrate intake showed weak correlations with valeric and isovaleric acids in females. This study highlights some possible demographic confounders linked to diet, disease, lifestyle, and microbiota that have to be taken into account when analyzing fecal metabolome data.
Keywords: 1H NMR; age; amino acids; carbohydrates; fat; fecal metabolome; sex; short-chain fatty acids.