Integration of Serum and Liver Metabolomics with Antioxidant Biomarkers Elucidates Dietary Energy Modulation of the Fatty Acid Profile in Donkey Meat

Donkey meat is valued for its high protein, unsaturated fats, and low cholesterol. Fatty acid (FA) composition critically influences meat quality and is modulated by dietary energy levels. Twenty-four meat donkeys (male) were randomly divided into three groups: a low-energy group (LEG), a medium-energy group (MEG), and a high-energy group (HEG). The trial lasted for 135 days, with dietary digestible energy levels adjusted during the pre-fattening, mid-fattening, and late-fattening phases according to the experimental design. The results showed that MEG and HEG interventions significantly upregulated tissue polyunsaturated fatty acid (PUFA) and n-3 PUFA content while reducing n-6/n-3 ratios, concomitant with enhanced activity and gene expression of most lipid-metabolizing enzymes. Notably, MEG further elevated antioxidant enzyme activities and anti-inflammatory mediators while suppressing pro-inflammatory factors. MEG and HEG significantly upregulated serum cholestane-3,7,12,25-tetrol-3-glucuronide and cortisol, along with hepatic choline, lysoPC(20:2(11Z,14Z)), glycocholic acid, and cholestane-3,7,12,25-tetrol-3-glucuronide. These modified metabolites were predominantly enriched in key metabolic pathways: pentose and glucuronate interconversions, primary bile acid biosynthesis, steroid hormone biosynthesis, glycerophospholipid metabolism, purine metabolism, and glutathione metabolism. Additionally, compared to HEG, MEG improved the antioxidant activities and immune signaling molecule levels with elevated pyroglutamic acid, glutathione, choline, inosine, adenine, and uric acid. Thus, moderately elevated dietary energy levels may enhance FA profiles in muscular and adipose tissues through coordinated regulation of lipid-metabolizing enzymes and associated gene expression, with serum and hepatic metabolites actively participating in these regulatory pathways. However, excessive energy intake could induce oxidative stress in donkeys.