Urine and blood samples from patients with different clinical disorders were examined for 2-hydroxybutyric acid (2-HB) by gas chromatography and mass spectrometry. All patients with combined lactic acidosis and ketoacidosis excreted 2-HB in the urine, in amounts up to 2.3 mmol/mmol creatinine. In blood from these patients, 2-HB was present only in trace amounts. Hard physical exercise resulted in lactic acidosis and accumulation of 2-HB. The lactate to pyruvate ratio in blood and urine was markedly increased in the patients excreting 2-HB but was normal in two patients with lactic acidosis who did not excrete 2-HB. It is concluded that an increased NADH2/NAD ratio is the most important factor for the production of 2-HB.