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Premna odorata is a plant species with various medicinal properties. However, in Central Vietnam, the seeds of this plant have not been previously studied for their chemical constituents. The aim of this study was to isolate and identify compounds from the seeds of P. odorata collected from Phu Yen province, Vietnam. The seeds were extracted using dichloromethane. The chemical constituents were isolated and purified. Their structures were elucidated through spectroscopic methods, including one-dimensional (1D) and two-dimensional (2D) Nuclear Magnetic Resonance (NMR) spectroscopy, mass spectrometry, and comparison with existing literature data. A new triterpenoid (1) and four known flavonoids: diosmetin (2), luteolin (3), acacetin (4), and apigenin (5), were successfully isolated from the dichloromethane extract of P. odorata seeds. The structures of these compounds were determined based on spectroscopic data. Compound 1 was also tested for cytotoxicity on the KB cell line, showing significant activity with an IC50 value of 40.98 μM. This study presents the first chemical investigation of the seeds of P. odorata. The identification of one new triterpenoid and four known flavonoids contributes to the understanding of the chemical composition of this plant and provides a basis for further pharmacological studies.