People with obesity tend to have altered functional connectivity of reward-related areas in the brain, contributing to overeating and weight gain. The gut-brain axis may function as a mediating factor, with gut-derived short-chain fatty acids (SCFAs) as possible intermediates in the relationship between microbiota and functional connectivity. We investigated the influence of SCFA turnover on resting state functional connectivity in healthy individuals with extremely high and extremely low levels of intestinal SCFA turnover. In this study, we included individuals with low or high intestinal SCFA turnover (estimated by fecal concentration of the butyryl-coenzyme A (CoA)-transferase (ButCoA) gene). Resting state functional magnetic resonance imaging (rs-fMRI) was used to assess functional connectivity of eight regions of interest (ROIs) either directly involved in the mesolimbic dopaminergic network (amygdala, hippocampus, caudate nucleus, putamen and nucleus accumbens) or primary projection regions of this network (middle frontal gyrus, superior frontal gyrus, insula). Functional connectivity was assessed using connectivity strength and eigenvector centrality. No differences in connectivity strength or eigenvector centrality were observed between the high and the low ButCoA group in any of our ROIs, suggesting SCFA turnover is not associated with resting state functional connectivity of central reward-related areas. Although previous studies provide evidence for an association between gut microbiota and resting state functional connectivity of reward-related areas, our findings do not support the hypothesis that this relationship is mediated by SCFAs. This suggests the existence of alternative mechanisms via which the intestinal microbiota may affect appetite, beyond local SCFA production.