The objective of this study was to confirm the effect of maternal genistein exposure on body weight of male offspring and the metabolic alterations associated with maternal genistein-induced obesity. Pregnant female Sprague–Dawley (SD) rats were supplemented with 300 mg/kg diet of genistein (GEN) or no genistein (CON) throughout pregnancy and lactation. The growth of male offspring was investigated until 12 week age and the mechanism of obesity was studied using metabonomics by ultra performance liquid chromatography and quadrupole time-of-flight (UPLC Q-TOF) MS with electrospray ionization in positive ESI mode (ESI+). Compared with the CON group, body weight, fat pad and food intake of male offspring in GEN group were increased significantly at the age of weeks 10 to 12 ( p <0.05). Ten urine principal metabolites contributing to the clusters were identified, including increased 8-Isoprostaglandin F2a, and decreased L-Proline, Betaine, L-Acetylcarnitine, Norsalsolinol, Indoleacrylic acid, L-Tryptophan, Lysophosphatidylcholines (LysoPC) (20 : 4), Lysophosphatidylethanolamines (LysoPE) (18 : 1) and LysoPC (O-18 : 0). Our results confirmed weight-increasing effects of maternal genistein exposure, accompanied by favorable changes in metabolites in the male offspring’ urine. Therefore, this research enables us to better understand obesity and predict risk of obesity-related disease by studying metabolites present in the urine.