This study was designed to monitor the biochemical profiles of serum and follicular fluid (FF) of postpartum dairy cows during the summer (n = 30) and winter (n = 30). Blood and FF (follicles ≥ 9 mm) were obtained from Girolando cows at 30, 45, 60, 75 and 90 days postpartum. The samples were collected and analysed to determine glucose, total cholesterol (TC), triglyceride (TG), urea, sodium (Na), potassium (K) and calcium (Ca) levels. Throughout the study, the following clinical variables were measured: rectal temperature (RT), respiratory rate (RR) and body condition score (BCS). In addition, the temperature humidity index (THI) was calculated for each season. During the summer season, THI was higher, BCS decreased, there was an increase in RT, and glucose, urea, Na and K serum levels were decreased (P < 0.05). The levels of TC, TG, urea, K and Ca in follicular fluid increased (P < 0.05). Positive correlations (P < 0.05) were observed between the serum and FF levels for glucose (r = 0.29), TC (r = 0.24) and Ca (r = 0.30). Therefore, the biochemical profile of serum and FF of dairy cows under summer heat-stress conditions demonstrates marked changes that may impair fertility during lactation.