Acute coronary syndrome (ACS) can cause arrhythmia, heart failure, and even sudden death. Our aim in this study was to identify potential metabolic biomarkers in patients with ACS. The serum metabonomics approach based on ultra-performance liquid chromatography (UPLC)/Orbitrap mass spectrometer (MS) was used to analyze the serum samples from 45 patients with ACS and 29 healthy controls. Multivariate statistical analysis was used to screen for ACS biomarkers. In total, 69 biomarkers were identified to be enriched in 19 metabolic pathways; 43 biomarkers were significantly up-regulated, while 26 biomarkers were significantly down-regulated in the ACS group. The main classes were lyso-sphingolipid (SM), cinnamic acids, cholines, and primary amides. Receiver operating characteristic (ROC) curve analysis showed that lysoPC(20:4(8Z,11Z,14Z,17Z)/0:0) (ROC area under the curve, AUC = 0.936), SM(d18:0/16:0) (ROC AUC = 0.932), and SM(d18:1/14:0) (ROC AUC = 0.923) had a high ACS diagnostic ability. The AUC value of the diagnostic model constructed using these combined biomarkers was 0.96. Therefore, these biomarkers may improve the diagnostic efficacy of ACS. The findings of this study also implied that glycerophospholipid metabolism; the biosynthesis of unsaturated fatty acids; linoleic acid metabolism; and valine, leucine, and isoleucine biosynthesis played important roles in ACS. Network analysis by ingenuity pathway analysis (IPA) showed these biomarkers were correlated to the cardiac hypertrophy signaling pathway, ERK/MAPK signaling pathway, NF-kappa B signaling pathway, nitric oxide (NO) signaling pathway in cardiovascular system, and TLR-signaling pathway. These findings will help to improve the ability of accurate diagnosis and intervention of ACS.