Grape seed extract (GSE) is a source of naturally occurring compounds known as proanthocyanidins and flavan-3-ols, which are recognized to exert a protective effect on human health, so GSE is widely used mainly as a nutritional supplement. However, polyphenols may have, in some cases, estrogenic effects or may interfere with the endocrine system. For that reason, it was considered of interest to investigate the beneficial or detrimental effects induced by low monomer content grape seed extract (LMC-GSE) in a teleost experimental model, the juvenile goldfish (Carassius auratus); therefore, biomarkers of estrogenic exposure together with cholesterol titers were assessed in both plasma and tissue samples taken from fish fed with different doses of LMC-GSE for 4 weeks. Dietary LMC-GSE (71 or 35 mg/g diet) did not affect vitellogenin (VTG) synthesis; on the contrary, VTG production was exclusively induced in fish fed with an estradiol-17beta (E2)-incorporated diet. In addition, it was found that both plasma E2 levels and hepatic total cholesterol were not affected by LMC-GSE dietary regimens.