All--retinoic acid (aRA) is the essential derivative of vitamin A and is of interest due to its various biological key functions . As shown in the recent literature, aRA also plays a role in the failing heart during myocardial infarction, the leading cause of death globally . To date insufficient mechanistic information has been available on related hypoxia-induced cell damage and reperfusion injuries . However, it has been demonstrated that a reduction in cellular aRA uptake abrogates hypoxia-mediated cell and tissue damage, which may offer a new route for intervention . Consequently, in this study, the effect of the novel cardio-protective compound 5-methoxyleoligin (5ML) on cellular aRA uptake was tested in human umbilical-vein endothelial cells (HUVECs). For this purpose, a high-performance liquid chromatography tandem mass spectrometry (HPLC-MS/MS) method was developed to assess intra-cellular levels of the active substance and corresponding levels of vitamin A and its derivatives, including potential / isomers . This work also focused on light-induced isomerization and the stability of biological sample material to ensure sample integrity and avoid biased conclusions . This study provides evidence of the inhibitory effect of 5ML on cellular aRA uptake, a promising step toward a novel therapy for myocardial infarction.