In contrast to plasma free fatty acids (FFA) remarkably little work has been carried out on myocardial FFA content directly. Reported values of normal FFA content vary so widely (29.0 to 25 000 nmol/g wet wt) that clearly the usual methods for the determination of FFA are difficult or not at all applicable to myocardial tissue. A method has been developed for the mass fragmentographic determination of myocardial FFA content and distribution and has been validated for sensitivity, reproducibility, specificity, recovery, cross-reaction with other lipids and reliability. The FFA content of normoxic canine myocardium was found to be 55.6 ± 20.2 nmol/g wet wt. This value is considerably below most of the previously reported values and much of the reported work on myocardial FFA content and changes must be re-examined.