Ovarian cancer is a heterogeneous disease of low prevalence but poor survival. Early diagnosis is critical for patients’ survival but often challenging because ovarian cancer symptoms are unclear and appear only in advanced stages of the disease. Finding more robust disease markers is therefore a clinical priority. Metabolomics is an emerging diagnostic tool that enables the detection of biomarkers reflecting alterations of tumor metabolism, a hallmark of cancer. We performed metabolomics profiling of serum and tumor tissue of high grade serous ovarian cancer (HGSOC) patients, and report novel diagnostic and prognostic biomarkers which indicate deregulation of hydroxybutyric acid (HBA) metabolism. The accumulation of HBA metabolites was associated with lower expression of succinic semialdehyde dehydrogenase (encoded by ALDH5A1), and with the presence of an epithelial-to-mesenchymal-transition (EMT) gene signature, implying a role for these metabolic alterations in cancer cell migration and invasion. In conclusion, our work reports the first comprehensive metabolomic investigation of HGSOC and proposes a new set of metabolites that can be used as diagnostic and prognostic biomarkers of the disease.