It is considered that high air temperature and humidity during the summer are the main factors which adversely affect both the health and production-reproductive performance of high yielding dairy cows. The resulting heath stress leads to a series of changes in endocrine regulation of homeostasis. The changes in hormonal status reflect in some way to the indicators of metabolic status of the cows. The objective of this work was to investigate the influence of heat stress on metabolic status of cows. The experiment was carried out on 20 cows of Holstein-Friesian breed during the summer, in the period from 18th to 45th day of lactation. During the performance of the experiment, the value of heat index (THI) was determined hourly and then the value of average morning (from 10 pm the previous day to 9 am the current day), afternoon (from 10 am to 9 pm the current day) and all-day THI was calculated. Blood sampling was carried out on the 1st, 2nd, 8th, 11th, 14th, 18th, 25th, 29th and 37th day of the experiment, in the morning and the afternoon. On the basis of hourly THI values, whole experimental period was divided into three periods: period A during which the cows were exposed to a extreme high heat stress (THI?78) at least 7 hours in 24 hours; period B during which the cows were exposed to a moderate heat stress (72?THI?78) at least 7 hours in 24 hours; period C during which the cows were not exposed to a heat stress (THI?72) in 24 hours. The average daily THI in period A (73,25?0,89) was significantly higher (p<0,01, individually) in regard to period B (71,45?0,96) and period C (65,41?2,09). THI was significantly higher in the period B than in the period C (p<0,01). Significantly lower blood glucose value (p<0,05) during the afternoon period in the cows exposed to the extreme heat stress (3,02?0,31 mmol/L) in regard to the morning period (3,14?0,41 mmol/L) points to the fact that in such conditions, metabolism redirects to use of glucose as an energy source because in that way less thermal energy is produced than during decomposition of fatty acids. Concentration of most important metabolic profile parameters in blood (cholesterol, triglycerides, total proteins, albumin, urea, total bilirubin, calcium, inorganic phosphorus as well as AST and ALT activity) did not vary significantly under the influence of heat stress. The exception was ionic calcium concentration which, under the conditions of extreme heat stress, was on the lower limit of physiological values (1.17?0.16 mmol/L).