While optimizing dietary energy and fat is important for laying hen performance, the effects of lysophospholipids under varying nutritional conditions remain unclear. This study aimed to evaluate the effects of fat source, metabolizable energy (ME) level, and lysophospholipid complex (LPL) supplementation on productive performance, egg quality, metabolic profile, yolk fatty acid composition, intestinal morphology, and nutrient digestibility in commercial laying hens during peak production (28–40 weeks). A total of 320 hens were allocated to a 2 × 2 × 2 factorial arrangement to receive diets varying in fat source (soybean oil or beef tallow), ME level [recommended (2800 kcal/kg) or reduced by 100 kcal/kg], and LPL supplementation (0 or 1 g/kg), with each treatment including eight replicates of five hens. Soybean oil reduced feed conversion ratio and serum alanine aminotransferase levels, while increasing villus height-to-crypt depth (VH:CD) ratio, as well as serum triglyceride and cholesterol concentrations, compared with beef tallow (P < 0.05). Additionally, soybean oil increased the proportion of polyunsaturated fatty acids (PUFA) while reducing saturated and monounsaturated fatty acids in egg yolks (P < 0.05). A 100 kcal/kg reduction in ME increased feed conversion ratio and serum uric acid, but decreased yolk color, fat digestibility, and apparent metabolizable energy corrected for nitrogen (AMEn) (P < 0.05). LPL supplementation enhanced nutrient digestibility, jejunal VH:CD ratio, and PUFA content in egg yolks (P < 0.05). Regarding factorial effects, LPL supplementation in beef tallow diets significantly increased egg production, egg mass, ileal fat and energy digestibility, and AMEn. An interaction between ME level and LPL supplementation also enhanced eggshell breaking strength, particularly under low-ME conditions (P < 0.05). These findings suggest that LPL supplementation, particularly in low-energy or high-saturated fat diets, can enhance productivity and digestive health in peak-phase laying hens. Further research should examine underlying mechanisms and long-term economic feasibility in commercial conditions.
