Nepthea sp. is a soft coral that grows abundantly in the seas of Southeast Sulawesi, Indonesia. However, there is no information available regarding its pharmacological or chemical characteristics. As a result, the goal of this research was to uncover the chemical profile of the Nepthea sp. ethyl acetate subfractions, as well as their antioxidant and anticancer potential. The sample was extracted with ethyl acetate and then fractionated using vacuum liquid chromatography with Si-gel as an adsorbent and a chosen solvent as an eluent. Phytochemical tests, Liquid Chromatography-Mass Spectroscopy/Mass Spectroscopy (LC-MS/MS), and total phenolic content were used to determine the chemical content. The 2,2-diphenyl-1-picrylhydrazyl (DPPH) and 2,2-azino-bis(3-ethylbenzothiazoline-6-sulfonic acid (ABTS) radicals were used to test the antioxidant potency, whereas MCF-7 cell lines were used in the 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide experiment to evaluate cytotoxicity. The fractionation of the ethyl acetate extract (160 g) produced six subfractions: Fractions A (35.2 g), B (4.3 g), C (5.9 g), D (10.7 g), E (26.5 g), and F (15.4 g). According to the DPPH and ABTS results, fraction E has the highest antioxidant potency (IC 50 = 67.39 ± 1.56 and 54.12 ± 0.95 mgl −1 , respectively), and fraction C has the highest anticancer activity (IC 50 = 72.82 ± 1.30 mgl −1 ). Fraction C components include 3-acetyl-3,4-dihydro-5,6-dimethoxy-2(1)H-benzopyrone, oxyphyllenone B, and unidentified chemicals, according to LC-MS/MS data (C 15 H 21 NO, C 21 H 33 NO 2 , C 15 H 23 NO 3 , C 15 H 21 NO 2 , C 15 H 21 NO 3 , and C 45 H 84 O 14). Rengyolester, piperolactam-C9:1(8E), valine, and unidentified chemicals (C 52 H 79 N 3 O, C 32 H 51 NO 7 ) make up fraction E. As a result, the ethyl acetate extract and its subfractions from Nepthea sp., especially fractions C and E, can be used as a source of raw materials for anticancer agents and antioxidants, respectively.