The human hepatoblastoma cell line, HepG2, has been used for investigating a wide variety of physiological and pathophysiological processes. However, less information is available about the phospholipid metabolism in HepG2 cells. In the present report, to clarify the relationship between cell growth and phospholipid metabolism in HepG2 cells, we examined the phospholipid class compositions of the cells and their intracellular organelles by using enzymatic fluorometric methods. In HepG2 cells, the ratios of all phospholipid classes, but not the ratio of cholesterol, markedly changed with cell growth. Of note, depending on cell growth, the phosphatidic acid (PA) ratio increased and phosphatidylcholine (PC) ratio decreased in the nuclear membranes, the sphingomyelin (SM) ratio increased in the microsomal membranes, and the phosphatidylethanolamine (PE) ratio increased and the phosphatidylserine (PS) ratio decreased in the mitochondrial membranes. Moreover, the mRNA expression levels of enzymes related to PC, PE, PS, PA, SM and cardiolipin syntheses changed during cell growth. We suggest that the phospholipid class compositions of organellar membranes are tightly regulated by cell growth. These findings provide a basis for future investigations of cancer cell growth and lipid metabolism.