Abstract: This study was conducted to investigate the effects of dietary glyceryl polyethylene glycol ricinoleate (PEGR) supplementation on growth performance and hepatic lipid metabolism in juvenile American eel (Anguilla rostrata), and to determine its optimal inclusion level. A total of 375 juvenile American eels with an initial body weight of (15.78±0.03) g were randomly assigned to five groups with three replicates per group and 25 fish per replicate. Fish were fed a basal diet (PEGR0) or diets supplemented with 0.50 (PEGR0.5), 1.00 (PEGR1), 2.00 (PEGR2), and 4.00 g/kg (PEGR4) PEGR for 10 weeks. The results showed that weight gain rate, specific growth rate, and feed intake in the PEGR0.5 and PEGR1 groups were significantly higher than those in the PEGR0 group (P<0.05), with the best growth performance observed in the PEGR0.5 group. Whole-body crude protein content was significantly increased (P<0.05), whereas hepatic crude lipid content was significantly decreased (P<0.05) in the PEGR0.5, PEGR1, and PEGR2 groups. Oil Red O staining further showed that dietary PEGR supplementation significantly reduced hepatic lipid droplet accumulation. However, no significant differences in the above parameters were observed in the PEGR4 group compared with the PEGR0 group (P>0.05). Serum total cholesterol levels in all PEGR-supplemented groups were significantly decreased (P<0.05), while triglyceride levels in the PEGR0.5 and PEGR1 groups were significantly reduced (P<0.05). In contrast, high-density lipoprotein cholesterol levels were significantly increased in the PEGR-supplemented groups (P<0.05). The activities of hormone-sensitive lipase and carnitine palmitoyltransferase-1 were significantly increased in the PEGR0.5 and PEGR1 groups (P<0.05), whereas fatty acid synthase activity was significantly decreased in the PEGR0.5 group (P<0.05). Untargeted metabolomics analysis revealed that lipid metabolism-related pathways, including glycerophospholipid metabolism, glycerolipid metabolism, linoleic acid metabolism, and α-linolenic acid metabolism, were enriched in the PEGR0.5 group, which might have contributed to improved lipid homeostasis. Based on the broken-line regression analysis of weight gain rate, the optimal dietary PEGR supplementation level for juvenile American eel was estimated to be 0.51 g/kg. In conclusion, appropriate dietary PEGR supplementation promoted growth performance and regulated lipid metabolism in juvenile American eel, whereas excessive supplementation failed to exert beneficial effects. The recommended dietary PEGR supplementation level was 0.50—0.51 g/kg.