The metabolism of feline follicles has been largely overlooked in reproductive science. The primary objective of this study was to determine differences in metabolite profiles and glucose carbon fluxes in feline follicles of different developmental stages and subjected to different developmental environments. Glycolytic flux, cholesterol (P <0.0001), pyruvic acid (P <0.0001), myo-Inositol (P =0.0003) and proline (P <0.0001) were higher in late versus early antral follicles. Serine (P =0.0085), threonine (P <0.0001), and valine (P =0.0025) were significantly higher in early antral follicles cultured in vitro than those derived directly from feline ovaries; however, glucose carbon flux was not significantly different. The current study suggests that early antral follicles undergo significant metabolic changes before progressing to the late antral stage. While it appears that early antral follicles have diminished capacity to progress to the late antral stage in vitro, it does not appear to be due to fundamental metabolic differences.