BACKGROUND: Long-term aerobic exercise during endurance racing places high demands on equine homeostasis. This study aimed to use proteomic analysis to elucidate complex biological responses during endurance exercise. It was hypothesized that different serum proteome changes would be noted, reflecting physiological processes as a response to race. The serum has been taken before and after an 80 km race from 13 endurance horses. Proteomic analysis of samples has been performed by TMT-based quantitative method. Apolipoprotein and haptoglobin values have been validated by enzyme-linked immunosorbent assay and biochemical assay respectively. The difference in protein abundance between pre and post-race values has been determined. RESULTS: In serum samples, 10 master proteins with significant p value differences between pre- and post-race abundances were detected. Increased protein abundance after the race was noted for the apolipoprotein groups: ApoA IV and E, Microfibril-associated glycoprotein 4 (MFAP4), transferrin, and antithrombin-III. Decreases in apolipoprotein C-II, C-III and R, alpha-1-microglobulin/bikunin precursor protein (AMBP) and haptoglobin abundance were reported after the race compared to before the race. Gene Ontology analysis revealed changes in triglyceride and acylglycerol homeostasis, lipid localization regulation, triglyceride catabolic processes, cholesterol binding, antioxidant activity and several cellular components. CONCLUSIONS: The endurance race caused several homeostatic imbalances characterized by various alterations in serum protein levels. The most pronounced changes emphasize the adaptation of energy metabolism to a more pronounced consumption of lipids.