Bile acids (BA) are signaling molecules with a wide range of biological effects, also identified amongst the most responsive plasma metabolites in the post-prandial state. We here describe this response to different dietary challenges and report on key determinants linked to its inter-individual variability. Healthy men and women (N=72, 62 ± 8 years) were enrolled into a 12-week weight loss intervention. All subjects underwent an oral glucose tolerance test (OGTT) and a mixed meal tolerance test (MMTT) before and after the intervention. BA were quantified in plasma by LC-MS/MS combined with whole genome exome sequencing and fecal microbiota profiling. Considering the average response of all 72 subjects, no effect of the successful weight loss intervention was found on plasma BA profiles. Fasting and post-prandial BA profiles revealed high inter-individual variability and 3 main patterns in post-prandial BA response were identified using multivariate analysis. Although the women enrolled were postmenopausal, gender effects in BA response were evident. Exome data revealed the contribution of preselected genes to the observed inter-individual variability. In particular, a variant in the SLCO1A2 gene, encoding the small intestinal BA transporter OATP1A2 was associated with delayed post-prandial BA increases. Fecal microbiota analysis did not reveal evidence for a significant influence of bacterial diversity and/or composition on plasma BA profiles. The analysis of plasma BA profiles in response to two different dietary challenges revealed a high inter-individual variability, which was mainly determined by genetics and gender of host with minimal effects of the microbiota.