Lung transplantation is the established treatment for patients with chronic, end-stage lung diseases such as chronic obstructive pulmonary disease (COPD), idiopathic pulmonary fibrosis (IPF) and cystic fibrosis (CF). However, its utility remains limited by the chronic shortage of donor lungs, limited lung preservation strategies and post-transplant complications leading to graft failure. Although efforts have been made to expand the limited pool of viable donor lungs via novel preservation strategies such as ex vivo lung perfusion (EVLP), our limited understanding of the mechanism and progression of donor lung injury continues to inhibit our ability to fully exploit these advances to improve lung transplant outcomes. Furthermore, the clinical standard for post-transplant assessment is limited to whole lung measurement such as pulmonary functional tests (PFTs) and structural imaging via radiography or HRCT, both of which lack the necessary sensitivity to detect lung rejection early. Given these limitations of currently available pre- and post-transplant lung assessment tools, a novel metabolic biomarker may provide higher sensitivity for determining the viability of donated lungs, as well as for assessing the onset of rejection before permanent structural changes in the lungs become apparent. We proposed that hyperpolarized (HP) [1-13C]pyruvate magnetic resonance imaging (MRI)—which provides real-time metabolic assessment of tissue based on the conversion of [1-13C] pyruvate to [1-13C]lactate via glycolysis, or to 13C bicarbonate via oxidative phosphorylation—may be an effective tool for assessing the health of donated lungs and may also serve as an early biomarker for detecting pulmonary graft dysfunction (PGD)-associated inflammation or acute lung rejection. In a rat model, we demonstrated the feasibility of using HP [1-13C]pyruvate nuclear magnetic resonance (NMR) spectroscopy to assess the viability of ex vivo perfused lungs. We further showed that our technique can be used to measure the improved viability of those lungs after treatment with ascorbic acid. Finally, translating our previously developed technique to in vivo HP [1-13C]pyruvate imaging of an inflamed rat lung, we not only demonstrated its utility for detecting lung transplantation rejection, but found that the HP lactate-to-pyruvate ratio is a better predictor of acute lung rejection in a rat model than computed tomography.