Glycerol is a naturally occurring 3-carbon alcohol in the human body. It is the structural backbone of triacylglycerol molecules, and can also be converted to a glycolytic substrate for subsequent metabolism. Serum glycerol concentrations approximate 0.05 mmol/L at rest, and can increase to 0.30 mmol/L during increased lipolysis associated with prolonged exercise or caloric restriction. When glycerol is ingested or infused at doses greater than 1.0 g/kg bodyweight, serum concentrations can increase to approximately 20 mmol/L, resulting in more than a 10 mOsmol/kg increase in serum osmolality. Glycerol infusion and ingestion have been used in research settings for almost 60 years, with widespread clinical use between 1961 and 1980 in the treatment of cerebral oedema resulting from acute ischaemic stroke, intraocular hypertension (glaucoma), intracranial hypertension, postural syncope and improved rehydration during acute gastrointestinal disease. Since 1987, glycerol ingestion with added fluid has been used to increase total body water (glycerol hyperhydration) by up to 700 ml, thereby providing benefits of improved thermoregulation and endurance during exercise or exposure to hot environments. Despite the small number of studies on glycerol hyperhydration and exercise, it appears to be an effective method of improving tolerance to exercise and other heat-related stressors.