Mitochondria play a fundamental role in generating of energy in cells. Electron transfer in electron transport chain (ETC) involved in mitochondria was one of the biological functions of heme. Heme biosynthesizes increased upon the addition of 5-aminolevulinic acid (ALA). Complex I and II protein expression were downregulated, while complex IV and cytochrome c expression were upregulated by the addition of ALA. Administration of ALA significantly increased Complex IV (COX) activity and cellular ATP level. Surprisingly, the mitochondrial content and mitochondrial membrane potential were unchanged. Consistently, the relative mRNA-expression of transcription factors affecting mitochondrial biogenesis was unchanged after the addition of ALA.