Ischemic heart disease involves the occlusion of blood vessels resulting in a cessation of oxygenated blood flow to the heart. This hypoxia, and the necessary reperfusion to salvage surviving myocytes, induces cellular damage. Notably mitochondrial dysfunction occurs, increasing the production of reactive oxygen and reactive nitrogen species (ROS/RNS). This increase in ROS/RNS overwhelms cellular antioxidant defence and can alter protein structure / function via various protein post-translational modifications (PTMs). One target of ROS/RNS is the most redox active amino acid cysteine (Cys). Cys redox PTMs can be broken down into two classes, those that are biologically reversible (e.g. S-glutathionylation) or those considered ‘irreversible’ (sulfinic and sulfonic acid; Cys-SO2H/SO3H). Irreversible Cys redox PTM occur with prolonged exposure to high levels of ROS/RNS and are associated with protein dysfunction and/or degradation. A mass spectrometry technique based on parallel reaction monitoring was employed to quantify changes in irreversible Cys modification in a Langendorff model of myocardial ischemia/reperfusion injury (I/R). Due to the low abundance of Cys, and low abundance of Cys PTMs, an enrichment strategy utilising strong cation exchange and hydrophilic interaction chromatography was used to better profile the changes in irreversible Cys PTM. I/R significantly increased the abundance of Cys-SO2H/SO3H-modified peptides from proteins involved in the tricarboxylic acid (TCA) cycle. By using a targeted metabolomic workflow we also observed concurrent perturbations in the abundance of metabolites involved in the TCA cycle occurred during I/R. The addition of an aminothiol antioxidant MPG (N-2-mercaptopropionylglycine) in reperfusion improved functional recovery of hearts, ameliorated irreversible modification of Cys, and improved the recovery from TCA cycle metabolic dysfunction induced by ischemia.