Acute kidney injury (AKI) is described as a relatively common complication of exercise. In clinical practice the diagnosis of AKI is based on serum creatinine, the level of which is dependent not only on glomerular filtration rate but also on muscle mass and injury. Therefore, the diagnosis of AKI is overestimated after physical exercise. The aim of this study was to determine changes in uremic toxins: creatinine, urea, uric acid, asymmetric dimethylarginine (ADMA), symmetric dimethylarginine (SDMA), trimethylamine N-oxide (TMAO) and urinary makers of AKI: albumin, neutrophil gelatinase-associated lipocalin (uNGAL), kidney injury molecule-1 and cystatin-C (uCyst-C) after long runs. Sixteen runners, mean age 36.7 ± 8.2 years, (2 women, 14 men) participating in 10- and 100-km races were studied. Blood and urine were taken before and after the races to assess markers of AKI. A statistically significant increase in creatinine, urea, uric acid, SDMA and all studied urinary AKI markers was observed. TMAO and ADMA levels did not change. The changes in studied markers seem to be a physiological reaction, because they were observed almost in every runner. The diagnosis of kidney failure after exercise is challenging. The most valuable novel markers which can help in post-exercise AKI diagnosis are uCyst-C and uNGAL.