This study utilized in vitro colonic fermentation to examine the impact of astaxanthin on the microbial catabolism of tryptophan. Astaxanthin significantly altered the gut microbiota and raised the tryptophan catabolism metabolite levels in an in vitro human colonic fermentation system. To eliminate the influence of substrate availability, we conducted in vitro colonic fermentation of the gut microbiota of astaxanthin-domesticated mice. We observed that the capacity of astaxanthin-domesticated gut microbiota to catalyze the conversion of tryptophan to indole and derivatives was considerably augmented. Astaxanthin significantly increased the relative abundance of Akkermansia, Ruminococcus, Bacteroides and Lactobacillus and elevated the levels of indole-3-lactic acid and indole-3-propionic acid. These results demonstrated that astaxanthin regulates tryptophan metabolism by modifying gut microbiota and increasing the levels of indole metabolites, such as indole-3-lactic acid and indole-3-propionic acid. This study provides insights into the physiological activity of astaxanthin and sheds light on the potential for enhancing tryptophan metabolism through dietary manipulation of the gut microbiota.