Abstract Background A high carbohydrate-low protein diet can induce hepatic global DNA hypomethylation in trout. The mechanisms remain unclear. Objective We aimed to investigate whether increase in dietary carbohydrates (dHC) or decrease in dietary proteins (dLP) can cause hepatic global DNA hypomethylation, and to explore the underlying mechanisms in trout. Methods Two feeding trials were conducted on juvenile males, both of which involved a 4-day fasting and 4-day refeeding protocol. In Trial 1, trout were fed either a high protein-no carbohydrate (HP-NC, protein 60% dry matter (DM), carbohydrates 0% DM) or a moderate protein-high carbohydrate (MP-HC, protein 40% DM, carbohydrates 30% DM) diet. In Trial 2, fish were fed either a moderate protein-no carbohydrate (MP-NC, protein 40% DM, carbohydrates 0% DM), a MP-HC (protein 40% DM, carbohydrates 30% DM), or a low protein-no carbohydrate (LP-NC, protein 20% DM, carbohydrates 0% DM) diet to separate the effects of dHC and dLP on the hepatic methylome. Global CmCGG methylation, DNA demethylation derivative levels, and mRNA expression of DNA (de)methylation-related genes were measured. Differences were tested by one-way ANOVA when data were normally distributed or by Kruskal-Wallis non-parametric test if not. Results In both trails, global CmCGG methylation levels remained unaffected, but the hepatic 5-mdC content decreased after refeeding (1–3%). The MP-HC group had 3.4-fold higher hepatic 5-hmdC and a similar 5-mdC level compared to the HP-NC group in Trial 1. Both MP-HC and LP-NC diets lowered the hepatic 5-mdC content (1–2%), but only the LP-NC group had a significantly lower 5-hmdC level (P < 0.01) compared with MP-NC group in Trail 2. Conclusions dHC and dLP independently induced hepatic global DNA demethylation in trout. The alterations in other methylation derivatives levels indicated the demethylation process was achieved through an active demethylation pathway and probably occurred at non-CmCGG sites.