Abstract Purpose This study explored the association of surgical traumatic stress with volatile organic compounds (VOCs) in breath. Methods Exhaled breath and blood samples were collected from 105 patients under general anesthesia at three time points: prior to incision (Pre-op), 2 h after incision (Intra-op), and prior to extubation (End-op). Differential mass spectral features between these time points were screened. Traumatic stress-related biomarker detection and mixed-effects model analysis were performed to define correlations and significance between parameters. Subgroup analysis was conducted to test the ability of mass spectral features to distinguish different surgical routes (open vs. laparoscopic). Results The abundances of mass spectral features changed significantly between these time points. All stress-related biomarker increased dramatically in the Intra-op group and decreased significantly after operation. Mixed-effects model analysis revealed that several features were significantly correlated with stress biomarkers. Most importantly, subgroup analysis revealed that different surgical routes failed to be distinguished by mass spectral features. Conclusion Surgical traumatic stress may change the exhaled breath mass spectral features in perioperative patients, providing preliminary evidence for mass spectral features use in future monitoring of this stress response.