Integrating blood biochemistry and plasma metabolomics to define age-related metabolic biomarkers in Alope village chickens

Abstract

Background and Ai: The Alope village chicken is an indigenous Indonesian line developed to combine local adaptability with improved growth potential. However, age-specific physiological and metabolic reference data remain limited, constraining evidence-based feeding and health management. This study aimed to integrate blood biochemical profiling and plasma metabolomics to characterize age-related metabolic changes, establish baseline reference values, and identify putative age-discriminant biomarkers in Alope village chickens. Materials and Methods: ), stratified into three age groups: G1 (8 weeks), G2 (12 weeks), and G3 (16 weeks) (n = 10 per group). Blood samples were collected for biochemical analysis of glucose, total protein, urea, and triglycerides using enzymatic colorimetric assays. Plasma metabolomic profiling was conducted using untargeted gas chromatography-mass spectrometry. Multivariate analyses, including principal component analysis and partial least squares-discriminant analysis, were used to explore age-associated metabolic patterns. Discriminant metabolites were identified based on variable importance in projection values and descriptive receiver operating characteristic analysis. Result: All biochemical parameters increased significantly with age (p < 0.05), remaining within physiological reference ranges, indicating normal metabolic development. Multivariate metabolomic analysis revealed clear age-dependent separation, with distinct metabolic signatures characterizing each growth stage. Exploratory pathway mapping indicated age-associated involvement of amino acid metabolism (glycine, serine, and threonine), lipid metabolism (glycerolipid and glycerophospholipid turnover), purine metabolism, the citrate cycle, and nodes linked to the pentose phosphate pathway. Seven metabolites demonstrated strong discriminatory performance (variable importance in projection ≈ 1.17-1.35; area under the curve ≥ 0.985) and were identified as putative age-related biomarkers, reflecting coordinated shifts in one-carbon/redox balance, membrane remodeling, and nucleotide and energy metabolism. Conclusio: Age was the primary determinant of metabolic organization in Alope village chickens from 8 to 16 weeks. The integration of blood biochemistry and plasma metabolomics revealed coordinated physiological transitions from early growth toward enhanced energy handling and metabolic stabilization. The identified candidate metabolites provide an exploratory framework for defining physiological age and support the potential application of age-informed metabolic indicators for precision nutrition and health monitoring in indigenous village chicken production systems.

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