Background: The aim was to compare the effects of high-flux hemodialysis (HF-HD) versus high-volume post-dilution hemodiafiltration (HV-HDF) on mortality risk. Methods: Retrospective observational study of prevalent patients on hemodialysis who were followed for two years and treated in 28 kidney centers in Colombia. In this study, we included all adult patients who had been on dialysis for at least 90 days treated with an arteriovenous fistula. They were classified as HF-HD if they underwent this treatment exclusively (100% of time). For HV-HDF, if they received this treatment in more than 90% of the observation period. The primary outcome variable was mortality, and the type of hemodialysis therapy was considered as the exposure variable. Propensity score matching (PSM) and Cox regression models were used to evaluate the effect of dialysis modality on the mortality risk. Results: A total of 2933 patients were analyzed, 2361 patients with HF-HD and 572 with HV-HDF. After PSM, 1010 prevalent patients remained; mortality rate was 14.2% (95% Confidence Interval—CI: 11.3–17.6%) and 5.9% (95%CI: 4.0–8.4%) in HF-HD and HV-HDF group, respectively. HV-HDF therapy was associated with a 55% reduction in mortality compared with the HF-HD group (Hazards ratio-HR: 0.45 [95%CI 0.32–0.64] p
< 0.001). Cardiovascular mortality rate was not statistically different between groups (HF-HD: 7.1% (36), HV-HDF: 3.4% (17), HR: 0.51 (95%CI: 0.21–1.28), p
: 0.152). However, in patients younger than 60 years, a beneficial effect was observed in favor to HV-HDF therapy with a 79% reduction in cardiovascular mortality risk (HR: 0.21, (95%CI: 0.05–0.79), p
: 0.021). Conclusion: After adjustment for different confounders, this study suggests that HV-HDF could reduce all-cause mortality compared to HF-HD therapy in prevalent patients on hemodialysis.