In many tissues the availability of L-cysteine is a rate-limiting factor in glutathione production, though this has yet to be fully tested in heart. This study aimed to test the hypothesis that supplying hearts with 0.5 mM L-cysteine would preserve glutathione levels leading to an increased resistance to ischaemia reperfusion. Left ventricular function was measured in isolated perfused rat hearts before, during and after exposure to 45 min global normothermic ischaemia. Control hearts received Krebs throughout, whilst in treated hearts 0.5 mM L-cysteine was added to the perfusate 10 min before ischaemia, and was then present throughout ischaemia and for the first 10 min of reperfusion. Reperfusion injury was assessed from the appearance of lactate dehydrogenase (LDH) in the effluent. In two separate groups of control and treated hearts, ATP and glutathione (GSH) contents were measured at the beginning and end of ischaemia. Hearts treated with 0.5 mM L-cysteine showed a significantly higher recovery of rate pressure product (16,256+/- 1288 mmHg bpm vs. 10,324+/- 2102 mmHg bpm, p < 0.05) and a significantly lower release of LDH (0.54+/- 0.16 IU/g wet weight vs. 1.44+/- 0.31 IU/g wet weight, p < 0.05) compared to controls. Also, the L-cysteine treated group showed significantly better preservation of ATP and GSH during ischaemia in comparison to control. These results suggest that the mechanisms underlying the cardioprotective effects of 0.5 mM L-cysteine may include: increased anaerobic energy production either directly or through reduced degradation of adenine nucleotides; direct scavenging of free radicals; and/or improved antioxidant capacity through glutathione preservation.