Metabolomics is a valuable tool for biomarker screening of colorectal cancer (CRC). In this study, we profiled the urinary metabolomes of patients enrolled in a prospective patient cohort (ColoCare). We aimed to determine changes in the metabolome of the longer clinical follow-up and ascertain candidate markers with possibly prognostic significance. In total, 199 urine samples from CRC patients prior to surgery (n = 97) or 1–8 days post-surgery (n = 12), and then after 6 (n = 52) and 12 months (n = 38) were analyzed using both GC–MS and 1 H-NMR. Both datasets were analyzed separately with built in uni- and multivariate analyses of Metaboanalyst 2.0. Furthermore, adjusted linear mixed effects regression models were constructed. Many concentrations of the metabolites derived from the gut microbiome were affected by CRC surgery, presumably indicating a tumor-induced shift in bacterial species. Associations of the microbial metabolites with disease stage indicate an important role of the gut microbiome in CRC. We were able to differentiate the metabolite profiles of pre-surgery CRC patients from those at any post-surgery timepoint using a multivariate model containing 20 marker metabolites (AUCROC = 0.89; 95 % CI 0.84–0.95). This is one of the first metabolomic studies to follow CRC patients in a prospective setting with repeated urine sampling over time. We were able to confirm markers initially identified in case–control studies and metabolites which may represent prognostic biomarker candidates of CRC.