Mitochondrial β-Oxidation of Fatty Acids: Regulation, Disorders, and Therapeutic Perspectives

Background: Mitochondrial β-oxidation of fatty acids is a central catabolic pathway responsible for energy production during fasting, exercise, and metabolic stress. It involves the sequential degradation of long-chain fatty acids into acetyl-CoA units, fueling the tricarboxylic acid cycle and ATP synthesis via oxidative phosphorylation. Dysregulation of this pathway is implicated in a spectrum of metabolic and mitochondrial disorders, highlighting the need for a deeper mechanistic understanding and improved therapeutic interventions. Methods and Results: This review provides a comprehensive analysis of the regulatory architecture governing mitochondrial β-oxidation, including transcriptional control by PPARα, AMPK-mediated signaling, substrate availability, and carnitine-dependent transport mechanisms. Key enzymatic steps—from acyl-CoA dehydrogenases to thiolases—are dissected in the context of substrate specificity, cofactor interactions, and mitochondrial compartmentalization. Genetic defects in β-oxidation enzymes result in inherited metabolic disorders such as medium-chain acyl-CoA dehydrogenase deficiency (MCADD), very-long-chain acyl-CoA dehydrogenase deficiency (VLCADD), and carnitine transport defects, often presenting with hypoglycemia, myopathy, and hepatic dysfunction. Advances in mass spectrometry-based metabolomics and next-generation sequencing have enhanced diagnostic accuracy and expanded the known mutational spectrum. Therapeutically, current strategies include dietary management, carnitine supplementation, and avoidance of catabolic stress, while emerging approaches such as PPAR agonists, gene therapy, and enzyme replacement are under investigation. Furthermore, secondary β-oxidation impairments contribute to complex diseases like heart failure, non-alcoholic fatty liver disease, and neurodegeneration, suggesting broader clinical relevance. Conclusion: Mitochondrial β-oxidation of fatty acids is a tightly regulated and clinically significant pathway whose disruption contributes to both rare and common diseases. Continued integration of biochemical, genetic, and therapeutic research is essential for advancing precision medicine approaches aimed at restoring mitochondrial metabolic function.