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Background: Interleukin 6 (IL-6) has an inflammatory effect, and its concentration in serum increases during exercise. However, no studies have assessed acute changes in IL-6 concentration after consecutive days of extreme and long-term exercise., Objective: This study aimed to assess acute changes in serum IL-6 concentration during four days of long-distance walking., Methods: This prospective observational study assessed 25 athletes (aged 44.8 ± 9.1 years), who covered a total of 251 km in four days. Blood samples were collected daily to assess serum IL-6 concentrations. Repeated-measures analysis of variance (with Bonferroni's post hoc test) and the Kruskal-Wallis H -test (with Dunn's post hoc test) were used to investigate the differences between the measures., Results: The serum IL-6 concentrations were higher on the four days of walking (1st day: 26.8 ± 14.8; 2nd day: 14 ± 7.4; 3rd day: 9.4 ± 10.8; 4th day: 4.5 ± 0.2 pg/mL) when compared to pre-walk values (pre-walk: 2.2 ± 2.1 pg/mL; p < 0.001). On the first day, there was a tenfold increase compared to the pre-walk value., Conclusion: The inflammatory response increased the serum concentration of IL-6 after four days of exercise. With the passing of days, there were reductions but not to baseline values., Competing Interests: The authors declare that there are no conflicts of interest., (© 2020 Soares et al.)