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This study investigated the acute endocrine responses to a 164-km road cycling event in a hot environment. Thirty-four male experienced cyclists (49.1 ± 8.3 years, 86.8 ± 12.5 kg, 178.1 ± 5.1 cm) participating in a 164-km road cycling event were recruited. Blood samples were collected within 0.3–2.0 h before the start (PRE: ~0500–0700 h) and immediately following the ride (POST). Samples were analysed for testosterone, growth hormone (GH), cortisol and interleukin-6 (IL-6). The temperature and humidity during the event were 35.3 ± 4.9°C and 47.2 ± 14.0%, respectively. Based on the finishing time, results for the fastest (FAST, 305 ± 10 min) and the slowest (SLOW, 467 ± 31 min) quartiles were compared. At POST, testosterone concentration was significantly (P < 0.05) lower (PRE, 20.8 ± 8.6; POST, 18.2 ± 6.7 nmol · L⁻¹), while GH (PRE, 0.3 ± 0.1; POST, 2.3 ± 0.3 µg · L⁻¹), cortisol (PRE, 661 ± 165; POST, 1073 …