Multiple primary lung cancer (MPLC) is becoming increasingly common in clinical practice. Imaging examination is sometimes difficult to differentiate from intrapulmonary metastasis (IM) or single primary lung cancer (SPLC) before surgery. There is a lack of effective blood biomarkers as an auxiliary diagnostic method.A total of 179 patients who were hospitalized and operated in our department from January to June 2019 were collected, and they were divided into SPLC with 136 patients, MPLC with 24 patients, and IM with 19 patients. In total, 96 healthy people without lung cancer were enrolled. Medical history, imaging, and pathology data were assembled from all participants. Plasma metabolomics analysis was performed by quadrupole time-of-flight tandem mass spectrometry, and data were analyzed using SPSS19.0/Simca 14.1/MetaboAnalyst5.0 software. Significant metabolites were selected by variable importance in projection, P value, and fold change. The area under the receiver operating characteristic curve was used to evaluate their diagnostic ability.There were significant differences in plasma metabolite profiles between IM and MPLC. Seven metabolites were screened out. Two metabolites had higher levels in IM, and five metabolites had higher levels in MPLC. All had favorable discriminating capacity. Phosphatidyl ethanolamine (38:5) showed the highest sensitivity (0.95) and specificity (0.92). It was followed by l-histidine with sensitivity 0.92 and specificity 0.84. l-tyrosine can be used to identify SPLC and MPLC. The panel composed of related metabolites exhibited higher diagnostic ability. Eight principal metabolites caused remarkable differences between healthy people and MPLC, and five of them had area under the curves greater than 0.85, showing good discriminating power.Through the study of plasma metabolomics, it was found that there were obvious differences in the metabolite profiles of MPLC, IM, SPLC, and the healthy population. Some discovered metabolites possessed excellent diagnostic competence with high sensitivity and specificity. They had the potential to act as biomarkers for the screening and differential diagnosis of MPLCs.Copyright © 2023 the Author(s). Published by Wolters Kluwer Health, Inc.