Supplementary Material for: Metabolomic markers for dietary carbohydrate, fat, and protein consumption: a two-sample Mendelian randomization study

Introduction: Observational studies have raised questions regarding the comprehensive metabolomic markers associated with dietary macronutrient intake. This study aimed to identify metabolites that are causally linked to dietary macronutrient consumption using a robust analytical framework. Methods: This genetic association study employed a two-sample Mendelian randomization approach. Genetic proxies for metabolites and their summary statistics were sourced from 64 genome-wide association studies, and a meta-analysis provided summary statistics for 3,362 metabolites. Summary statistics for dietary carbohydrate, fat, and protein intakes were obtained from the UK Biobank. Mendelian randomization associations were assessed using inverse variance-weighted, weighted median, and Egger’s regression models to enhance robustness. Significant metabolites were subjected to enrichment analysis to identify relevant metabolic pathways. Associations were expressed as odds ratios with 95% confidence intervals, and enrichment analysis was used to identify key KEGG pathways based on p-values. Results: Mendelian randomization analysis identified 540 metabolite-macronutrient associations: 186 for carbohydrate intake, 217 for fat intake, and 137 for protein intake. Metabolites inversely associated with carbohydrate and protein intake, but positively linked to fat intake, were enriched in KEGG pathways, particularly those governing the biosynthesis and metabolism of aromatic amino acids (tryptophan and phenylalanine) and branched-chain amino acids (tyrosine, valine, leucine, and isoleucine). Conclusion: This study comprehensively mapped the metabolite associations with dietary macronutrient intake using extensive genetic data. These findings highlight the critical roles of aromatic and branched-chain amino acid biosynthesis and metabolism as key metabolic pathways in macronutrient regulation.