The Lithuanian Zemaitukai horse breed is one of the oldest in Europe. Currently, there is a lack of information about the effect of endurance competition on blood indices and acid-base balance in the Zemaitukai horses. The aim of this study was to evaluate the effect of endurance exercise on the acid-base balance and electrolyte indicators of the Zemaitukai horses in an official endurance competition. In total, 48 horses of the Zemaitukai breed competed in endurance competitions over the same distance (60 km). Samples were taken before and immediately after the exercise. The following indicators were analysed: the hydrogen potential (pH), partial carbon dioxide pressure (pCO(2)), partial oxygen pressure (pO(2)), base excess in blood (BE), base excess in the extracellular fluid [BE (ecf)], cHCO(3) (bicarbonate), cSO(2) (oxygen saturation), tCO(2) (blood total carbon dioxide), haematocrit (HCT), haemoglobin concentration (cHgb), sodium (Na), potassium (K), calcium (Ca), chlorides (Cl), and lactate (Lac). Increase in pH showed adaptation of the horse metabolism to exercise. Blood changes were caused by the development of dehydration and metabolic alkalosis. According to Lac results, all horses were fit and tolerated well physical activity. The exercise did not significantly affect the demand for minerals. Based on the results obtained, it can be stated that horses of the Zemaitukai breed are suitable for endurance competing.