Ovarian reserve rather than chronological age dominates follicular metabolic remodeling: evidence from a 2 × 2 factorial metabolomics study

Advanced age and diminished ovarian reserve (DOR) are both associated with impaired reproductive potential, but their independent and combined associations with the follicular metabolic microenvironment remain unclear. We performed untargeted metabolomic profiling of follicular fluid from 126 women classified by age and ovarian reserve status in a 2 × 2 factorial design. Within each age stratum, DOR versus normal ovarian reserve (NOR) comparisons were adjusted for continuous age. Complementary analyses evaluated reserve-adjusted age associations, age × ovarian reserve interactions, pathway enrichment, and associations with oocyte and embryo outcomes. Multiple testing was controlled using the Benjamini–Hochberg false discovery rate (BH-FDR). Distinct metabolic alterations associated with DOR were identified in both age strata. In contrast, evidence for an independent association with chronological age was limited and depended on model specification, and no robust age × ovarian reserve interaction remained significant after multiple-testing correction. Exploratory interaction profiles nevertheless suggested that DOR-associated metabolic alterations may differ between age groups. Directional pathway enrichment implicated steroid hormone biosynthesis in older women with DOR and bile acid pathways in younger women with DOR. Several metabolic features were also associated with oocyte and embryo outcomes. These findings suggest that follicular-fluid metabolic alterations are more strongly associated with ovarian reserve status than with chronological age. Age-related associations were model-dependent, whereas age × ovarian reserve interaction patterns remained exploratory and require validation in independent cohorts. Not applicable.