A 60-day feeding trial was conducted to evaluate the effects of different TMAO (trimetlylamine oxide) supple-ment levels (0%, 0.12%, 0.24%, and 0.48%, named G1, G2, G3, and G4, respectively) on growth performance, feed utilization, intestinal enzyme activities and appetite regulation of Eriocheir sinensis. The results showed that diet supplemented with 0.24% trimetlylamine oxide (TMAO) significantly improved the weight gain rate (WGR), specific growth rate (SGR), feed intake (FI) and feed utilization (p < 0.05), while having no effect on the survival rate (SR) and molting frequency (MF) of crabs (p > 0.05). Moreover, moderate dietary TMAO levels significantly improved the intestinal enzyme activities (& alpha;-amylase, trypsin, lipase and & gamma;-glutamyl transferase) of crabs. The RT-qPCR results showed that the expressions of the orexigenic neural signal-related factors neuropeptide Y (npy) and ghrelin were significantly up-regulated. The expressions of AKT/TOR/4E-BP signaling pathway related genes (protein kinase B (akt), target of rapamycin (tor) and 4E-binding protein (4e-bp)), anorexia-regulating peripheral hormones (leptin and cholecystokinin (cck)) and leptin receptor (lepr) were significantly down-regulated in the hepatopancreas, thoracic ganglia and cranial ganglia tissues. The anorexigenic neural signal-related factors cocaine and amphetamine regulated transcript (cart) and pro-opiomelanocortin (pomc) were significantly down-regulated in the hepatopancreas, thoracic ganglia and cranial ganglia tissues. Based on the quadratic regression analysis of WGR against dietary TMAO level, the optimum TMAO requirement of E. sinensis was suggested to be 0.27%. In conclusion, present results suggested that dietary supplementation with 0.27% TMAO can improve the growth performance, feed intake, intestinal enzyme activities and appetite regulation of E. sinensis.