Aging is a natural and inevitable process that leads to the irreversible impairment of biological functions and increased vulnerability to death. Great efforts have been spent to study the mechanisms underlying the aging process, with the ultimate goal of developing effective interventions to delay the onset of aging and to extend lifespan. Mitochondria, more than just being the powerhouses of the cell, are involved in a wide range of cellular and metabolic processes. Multiple lines of evidence have suggested that mitochondria play a vital role in the process of aging. Therefore, my research aims to investigate the effect of both dietary supplements and mitochondrial DNA content on lifespan. Chapter one reviews the contemporary knowledge of mitochondrial DNA genetics and the implication of mitochondrial dysfunction in human diseases. It also summarizes the interaction between mitochondria and different dietary components or dietary patterns. I want to emphasize the importance of investigating the role of mitochondria in the development of complex diseases and to suggest future directions in identifying nutritional interventions to restore mitochondrial functions. Chapter two and Chapter three describe the discoveries of two compounds that extend the lifespan of Drosophila melanogaster. The first one is konjac glucomannan hydrolysate, which can promote longevity in both genders across different genetic backgrounds, and regardless of mating status. By investigating the potential underlying mechanisms, we found it extends the lifespan through promoting the growth of gut microbiome and preserving the intestinal proliferative homeostasis. The second compound is oligofructose, the hydrolyzed form of inulin. It can also prolong the lifespan but is restricted to specific genetic backgrounds and mating status. Revealed by the transcriptome analysis, it is very likely that it extends lifespan through the inhibition of mitochondrial respiration and suppression of stress signaling pathways. Chapter four describes an exploratory project on the relationship between mitochondrial DNA content and lifespan in both normal and stress conditions. By implementing both survival test and transcriptome analysis, we found a possible connection between mtDNA copy number and the lifespan in male flies through regulation of fertility.