Investigation of the Anticancer Potential of Vernonia amygdalina Methanol Extract: A New Hope for MDA-MB-231 Breast Cancer Therapy
Abstract
MDA-MB-231 is a breast cancer cell line derived from metastatic adenocarcinoma and classified as a triple-negative breast cancer (TNBC) subtype. This breast cancer subtype is highly aggressive, has a high recurrence rate, and offers limited treatment options due to the absence of estrogen, progesterone, and HER-2 receptor expression. Therefore, the development of advanced therapeutic strategies is urgently needed to inhibit the metastasis of TNBC cancer cells. This study aimed to investigate the potential of the methanol extract of Vernonia amygdalina leaves as a natural chemotherapeutic agent for metastatic breast cancer therapy, with a focus on inducing cell death through apoptosis mechanisms. This study was an experimental research that began with the collection and identification of raw materials, followed by the preparation of simplicia, extraction processes, and phytochemical screening. Subsequently, cytotoxicity testing was conducted using the MTT assay, cell cycle analysis was performed using the PI-staining assay, and apoptosis was assessed using the Annexin V/PI-staining assay, all of which were analyzed through flow cytometry. The methanol extract of V. amygdalina demonstrated a moderate cytotoxic effect with an IC50 value of 109.36 µg/mL. The extract induced S-phase cell cycle arrest in a dose-dependent manner, indicating its ability to reduce cell viability by inhibiting DNA replication. In the apoptosis assay, the highest percentage of cell death (3.12%) was observed at the concentration of 54.5 µg/mL, suggesting that this dose produced the strongest apoptotic response among the treatments tested. The methanol extract of V. amygdalina leaves shows potential as a natural chemotherapeutic agent for metastatic breast cancer. The extract can induce cancer cell death through apoptosis, indicating its promise for further development as a supportive anticancer therapy.
Keywords
Full Text:
PDFReferences
Adascalului, M., Tih?uan, B.-M., Berca, L. M., Martinez Sanmartin, A., Nica, S., Cimponeriu, D., & Du??, D. (2020). Experimental in vitro cytotoxicity evaluation of plant bioactive compounds and phytoagents: a review Cytotoxicity evaluation of plant bioactive compounds. Romania. All Rights Reserved Rom Biotechnol Lett, 25(4), 1832–1842. https://doi.org/10.25083/rbl/25.4/1832.1842
Alara, O. R., Abdurahman, N. H., Ukaegbu, C. I., & Kabbashi, N. A. (2019). Extraction and characterization of bioactive compounds in Vernonia amygdalina leaf ethanolic extract comparing Soxhlet and microwave-assisted extraction techniques. Journal of Taibah University for Science, 13(1), 414–422. https://doi.org/10.1080/16583655.2019.1582460
Alshehade, S. A., Almoustafa, H. A., Alshawsh, M. A., & Chik, Z. (2024). Flow cytometry-based quantitative analysis of cellular protein expression in apoptosis subpopulations: A protocol. Heliyon, 10(13), e33665. https://doi.org/10.1016/j.heliyon.2024.e33665
Amarante-Mendes, G. P., Adjemian, S., Branco, L. M., Zanetti, L. C., Weinlich, R., & Bortoluci, K. R. (2018). Pattern Recognition Receptors and the Host Cell Death Molecular Machinery. Frontiers in Immunology, 9. https://doi.org/10.3389/fimmu.2018.02379
Bianchini, G., Balko, J. M., Mayer, I. A., Sanders, M. E., & Gianni, L. (2016). Triple-negative breast cancer: challenges and opportunities of a heterogeneous disease. Nature Reviews Clinical Oncology, 13(11), 674–690. https://doi.org/10.1038/nrclinonc.2016.66
Bob I.A. Mgbeje, James Ofem Otu, & Flora O. Ugoanyanwu. (2020). Phytochemical Components of Butanol Fraction of Vernonia amygdalina Leaf Extract Using GC-MS Analysis. Merit Research Journal of Medicine and Medical Sciences , 8(4), 74–79.
Chen, X., Li, H., Zhang, B., & Deng, Z. (2022). The synergistic and antagonistic antioxidant interactions of dietary phytochemical combinations. Critical Reviews in Food Science and Nutrition, 62(20), 5658–5677. https://doi.org/10.1080/10408398.2021.1888693
Degu, S., Meresa, A., Animaw, Z., Jegnie, M., Asfaw, A., & Tegegn, G. (2024). Vernonia amygdalina: a comprehensive review of the nutritional makeup, traditional medicinal use, and pharmacology of isolated phytochemicals and compounds. Frontiers in Natural Products, 3. https://doi.org/10.3389/fntpr.2024.1347855
Farmasita Nabilla Cahyani, Rachmawati Ardiana, Dewi Uswatun Khasanah, Adinda Sukma Dewi, & Oktavia Rahayu Adianingsih. (2021). Development and Optimation of Microsphere Capsule of Licorice Extract as Oral Extended Release Dosage Form for Breast Cancer . Pharmaceutical Journal of Indonesia, 7(1), 63–70.
Garrido-Castro, A. C., Lin, N. U., & Polyak, K. (2019). Insights into Molecular Classifications of Triple-Negative Breast Cancer: Improving Patient Selection for Treatment. Cancer Discovery, 9(2), 176–198. https://doi.org/10.1158/2159-8290.CD-18-1177
Hardiansyah, M., Musa, Y., & Jaya, A. (2021). The Effectiveness of Giving Plant PGPR Rhizosphere Bamboo on Cocoa Seeds Germination at The Nursery Level. Biology, Medicine, & Natural Product Chemistry, 10(1), 1-5. doi:https://doi.org/10.14421/biomedich.2021.101.1-5
Hasibuan, P. A. Z., Harahap, U., Sitorus, P., & Satria, D. (2020). The anticancer activities of Vernonia amygdalina Delile. Leaves on 4T1 breast cancer cells through phosphoinositide 3-kinase (PI3K) pathway. Heliyon, 6(7), e04449. https://doi.org/10.1016/j.heliyon.2020.e04449
Huang, Z., Yu, P., & Tang, J. (2020).
Characterization of Triple-Negative Breast Cancer MDA-MB-231 Cell Spheroid Model
. OncoTargets and Therapy, Volume 13, 5395–5405. https://doi.org/10.2147/OTT.S249756Imran, M., Rauf, A., Abu-Izneid, T., Nadeem, M., Shariati, M. A., Khan, I. A., Imran, A., Orhan, I. E., Rizwan, M., Atif, M., Gondal, T. A., & Mubarak, M. S. (2019). Luteolin, a flavonoid, as an anticancer agent: A review. Biomedicine & Pharmacotherapy, 112, 108612. https://doi.org/10.1016/j.biopha.2019.108612
Kadiri, O., & Olawoye, B. (2016). Vernonia amygdalina: An Underutilized Vegetable with Nutraceutical Potentials – A Review. Turkish Journal of Agriculture - Food Science and Technology, 4(9), 763. https://doi.org/10.24925/turjaf.v4i9.763-768.570
Kemenkes RI. (2017). FARMAKOPE HERBAL INDONESIA EDISI II 2017 KEMENTERIAN KESEHATAN REPUBLIK INDONESIA 615.1 Ind f.
Lin, Y.-T., Lin, J., Liu, Y.-E., Chen, Y.-C., Liu, S.-T., Hsu, K.-W., Chen, D.-R., & Wu, H.-T. (2022). USP7 Induces Chemoresistance in Triple-Negative Breast Cancer via Deubiquitination and Stabilization of ABCB1. Cells, 11(20), 3294. https://doi.org/10.3390/cells11203294
Maadi, H., Soheilifar, M. H., & Wang, Z. (2022). Analysis of Cell Cycle by Flow Cytometry (pp. 183–195). https://doi.org/10.1007/978-1-0716-2736-5_14
Mohammed, M. F., Raman, N., Alhoot, M. A., & Alwan, R. (2020). Antibacterial Activities of Allium Sativum (Garlic) Extracts Against Staphylococcus Aureus and Escherichia Coli. In European Journal of Molecular & Clinical Medicine (Vol. 7, Number 11).
Ndayambaje, M., Habyarimana, T., Niyonsaba, T., Mwiseneza, A., Nshizirungu, J. P., Yadufashije, C., Naya, A., & Oudghiri, M. (2025). Vernonia amygdalina Delile and cancer: a comprehensive review of its chemopreventive mechanisms and efficacy. Discover Plants, 2(1), 30. https://doi.org/10.1007/s44372-025-00104-8
Nugraha, A. T., Prayitno, G., Situmorang, M. E., & Nasution, A. (2020). The role of infrastructure in economic growth and income inequality in Indonesia. Economics & Sociology, 13(1), 102–115. https://doi.org/10.14254/2071-789X.2020/13-1/7
Pandey, P., Lakhanpal, S., Mahmood, D., Kang, H. N., Kim, B., Kang, S., Choi, J., Choi, M., Pandey, S., Bhat, M., Sharma, S., Khan, F., Park, M. N., & Kim, B. (2025). An updated review summarizing the anticancer potential of flavonoids via targeting NF-kB pathway. Frontiers in Pharmacology, 15. https://doi.org/10.3389/fphar.2024.1513422
Peng, B.-Y., Singh, A. K., Chan, C.-H., Deng, Y.-H., Li, P.-Y., Su, C.-W., Wu, C.-Y., & Deng, W.-P. (2023). AGA induces sub-G1 cell cycle arrest and apoptosis in human colon cancer cells through p53-independent/p53-dependent pathway. BMC Cancer, 23(1), 1. https://doi.org/10.1186/s12885-022-10466-x
Tumilaar, S. G., Hardianto, A., Dohi, H., & Kurnia, D. (2024). A Comprehensive Review of Free Radicals, Oxidative Stress, and Antioxidants: Overview, Clinical Applications, Global Perspectives, Future Directions, and Mechanisms of Antioxidant Activity of Flavonoid Compounds. Journal of Chemistry, 2024, 1–21. https://doi.org/10.1155/2024/5594386
Ugbogu, E. A., Emmanuel, O., Dike, E. D., Agi, G. O., Ugbogu, O. C., Ibe, C., & Iweala, E. J. (2021). The Phytochemistry, Ethnobotanical, and Pharmacological Potentials of the Medicinal Plant-Vernonia amygdalina L. (bitter Leaf). Clinical Complementary Medicine and Pharmacology, 1(1), 100006.https://doi.org/10.1016/j.ccmp.2021.100006
Urbain, A., & Simões?Pires, C. A. (2020). Thin?Layer Chromatography for the Detection and Analysis of Bioactive Natural Products. In Encyclopedia of Analytical Chemistry (pp. 1–29). Wiley. https://doi.org/10.1002/9780470027318.a9907.pub2
Valentine Mbatchou. (2017). Vernonia amygdalina Leaf: Unveiling its antacid and carminative properties In Vitro. International Scientific Organization.
WHO. (2018). Quality control methods for herbal materials. World Health Organization.
Wu, L., Lin, Y., Gao, S., Wang, Y., Pan, H., Wang, Z., Pozzolini, M., Yang, F., Zhang, H., Yang, Y., Xiao, L., & Xu, Y. (2023). Luteolin inhibits triple-negative breast cancer by inducing apoptosis and autophagy through SGK1-FOXO3a-BNIP3 signaling. Frontiers in Pharmacology, 14, 1200843. https://doi.org/10.3389/fphar.2023.1200843
DOI: https://doi.org/10.14421/biomedich.2026.151.885-893
Refbacks
- There are currently no refbacks.
Copyright (c) 2026 Ivan Junius Mesak, Ardhi Broto Sumanto
Biology, Medicine, & Natural Product Chemistry |



