The most common form of toxemia observed in large animals is endotoxemia. Endotoxemia is caused by the presence of lipopolysaccharide cell wall components of gram-negative bacteria in the blood and characterized clinically by abnormalities of many body systems. This study aimed to determine changes in serum inflammatory cytokine levels in calves with LPS-induced endotoxemia. Ten healthy Holstein breed calves were used in the study. To create experimental endotoxemia, LPS at 0.1 µg/kg dose was given intravenously to the calves from the vena jugularis as an infusion within 30 min. Blood samples to evaluate cytokines, biochemical, blood gases, and hematological parameters were collected before LPS infusion (0. hour) and at 1, 2, 3, 4, 6, 12, 24, and 36 h following LPS infusion. In the first 4 h following the LPS infusion, calves’ body temperature, heart and respiratory rate, and capillary refill time increased, while a decrease was observed in the following hours. Significant changes were detected in blood pH, WBC, and glucose levels after LPS administration. Serum TNF-α, IL-6, IL1-β, IL-10, and IFN-γ levels were low before the LPS infusion in the calves, while increased within the first 4 h following the LPS infusion, however, statistical significance was observed in TNF-α, IL-6, and IFN-γ. Cytokine levels decreased back to baseline levels after the fourth hour. As a result, it was found that 0.1 µg/kg dose of LPS administrated as infusion caused changes in clinical, hematological, blood gases, biochemical parameters, and significant increases in the levels of circulating cytokines. The results of this study showed that the changes observed in the organism during natural endotoxemia can also be revealed experimentally by administering LPS intravenously.