Sexual dimorphism in the serum metabolome following acute exhaustive exercise.

BACKGROUND: Sex differences in exercise metabolism have been recognized for decades, but the molecular metabolic landscape in which men and women reach standardized physiological exhaustion criteria remains unexplored. OBJECTIVE: We systematically characterized serum metabolic sexual dimorphism following acute exhaustive exercise using standardized termination criteria. METHODS: In a cross-sectional study (ChiCTR2400089036), forty healthy adults (20 males, 20 females; aged 22.4 ± 3.4 years, BMI 22.4 ± 2.1 kg/m2; V̇O2peak 40.92 ± 5.69 ml/kg/min) underwent cardiopulmonary exercise testing via objective termination criteria, with female participants tested during the mid-luteal phase. Serum samples were collected at baseline, immediately post-exercise, and at 15 and 30 min post-exercise for comprehensive metabolomics and targeted triacylglycerol (TAG) analysis via mass spectrometry. RESULTS: All participants achieved standardized exhaustion endpoints. Despite equivalent fat-free mass-normalized V̇O2peak, males expended 20% more energy per unit fat-free mass (P = 0.015). While both sexes showed similar numbers of altered metabolites (290-308), their molecular compositions differed markedly. Lipids comprised the largest fraction of sex-specific responses, with hypoxanthine, sarcosine, and lysophospholipids as key discriminators. Females showed sustained lipid downregulation while males demonstrated recoverable patterns. Notably, 37 TAGs showed sexually antagonistic regulation, and 41.7% of fitness-correlated metabolites exhibited opposite associations between sexes. CONCLUSIONS: This study reveals distinct metabolic response patterns between males and females when standardized exhaustion endpoints are reached. Key exercise-induced sex-discriminating metabolites were identified and opposing metabolic-fitness associations were observed between sexes. These findings emphasize the necessity of sex-stratified analysis in exercise metabolism research and metabolic biomarker interpretation. While men and women respond differently to exercise, molecular differences at complete exhaustion have never been studied. We studied 20 men and 20 women who exercised to exhaustion using standardized criteria, then analyzed hundreds of blood molecules before, during, and after exercise. Although both sexes showed similar numbers of changed molecules, the specific types were remarkably different, with fat-related molecules showing the largest differences. We identified molecular markers that distinguished male from female responses, found certain fat storage molecules responded in opposite directions between sexes, and discovered that molecules like kynurenine and androsterone sulfate showed opposite fitness relationships in men versus women. These findings reveal that even at identical exhaustion levels, men and women use fundamentally different molecular strategies. By understanding these biological differences, we could develop personalized exercise and nutrition plans that work better for everyone's biology, potentially improving health outcomes and athletic performance. HIGHLIGHTS: 1. Males and females exhibit distinct serum metabolomic profiles when they reach standardized physiological exhaustion endpoints, with similar numbers of altered metabolites but different molecular compositions. 2. Lipid species composed the largest fraction of sex-differential responses, including LPLs, bile acids, and fatty acids, showing female-dominant sustained downregulation and male-dominant recoverable downregulation patterns. 3. Targeted analysis revealed the sexually antagonistic regulation of 37 triacylglycerol species. These species are characterized primarily by C50‒C52 carbon chain lengths and enrichment with monounsaturated fatty acids, particularly those containing palmitoleic acid. 4. Key metabolites showing sex-related differential characteristics, including hypoxanthine, sarcosine, and LPLs, were identified, providing molecular markers for sex differences in exercise responses. 5. Among the 115 metabolites correlated with V̇O2peak, 41.7% exhibited opposing correlations between sexes, with L-kynurenine and androsterone sulfate as key examples, highlighting the necessity of sex-stratified analysis and the limitations of pooled approaches.