Redox status influences the course of the inflammatory, metabolic, and proliferative liver diseases. Oxidative stress is thought to play a crucial and sustained role in the pathological progression of early steatosis to severe hepatitis, fibrosis, cirrhosis, and hepatocellular carcinoma. Oxidative stress induced by reactive oxygen species which are generated in the mitochondria can lead to chronic organelle damage in hepatocytes. Currently, the diagnosis of liver disease requires liver biopsy, which is invasive and associated with complications. The present report describes the development of a novel molecular probe, EDA-PROXYL, with higher reactivity and mitochondrial selectivity than standard carboxyl-PROXYL and carbamoyl-PROXYL probes. The membrane permeability of our probe improved in aqueous environments which led to increased accumulation in the liver and interaction of EDA-PROXYL with the carnitine transporter via the amine (NH3+) group further increased accumulation. This increased mitochondrial sensitivity and enhanced accumulation highlight the potential of EDA-PROXYL as a molecular probe for determining metabolic reactions of the mitochondria. Thus, this novel probe could be a tool for the evaluation of redox status of the mitochondria to assess the degree of liver injury and, ultimately, the response to pharmacological therapy.