Abnormal mitochondria are an increasingly recognized cause of neuromuscular disease. We have used phosphorus magnetic resonance spectroscopy to monitor noninvasively the metabolism of high‐energy phosphates and the intracellular pH of human skeletal muscle in vivo in 12 patients with mitochondrial myopathy. At rest, an abnormality could be demonstrated in 11 of 12 patients. Ten patients had evidence of a reduced muscle energy state with at least one of the following abnormalities: low phosphorylation potential, low phosphocreatine concentration, high adenosine diphosphate concentration, or high inorganic phosphate concentration. Two patients had abnormal resting muscle intracellular pH. In some patients phosphocreatine concentration decreased to low values during exercise despite limited work output. This was not accompanied by particularly severe intracellular acidosis. Evidence of impaired rephosphorylation of adenosine diphosphate to adenosine triphosphate during recovery from exercise was found in approately half the patients. Phosphorus magnetic resonance spectroscopy is useful in the noninvasive diagnosis of mitochondrial myopathies and in defining the pathophysiological basis of these disorders.