Phytochemical screening, antibacterial and cytotoxic activities of Aristolochia philippinensis Warb. leaf extracts

Abstract

The Aristolochia philippinensis Warb. (Aristolochiaceae) is commonly used in traditional medicine among indigenous people in the Philippines. The leaves of A. philippinensis are used to treat illnesses such as stomach aches, bodily aches, and physical relapse. This study was conducted to identify the phytochemical compounds and evaluate the antibacterial and cytotoxic activity of A. philippinensis. The leaf extracts were subjected to qualitative phytochemical screening, in vitro testing for antibacterial activity using agar well-diffusion assay against selected gram-positive and gram-negative bacteria and cytotoxic activity using 3-(4,5-dimethylthiazol-2-y-l)-2,5-diphenyltetrazolium bromide (MTT) cell viability assay against human colorectal carcinoma cells (HCT 116). The results of the phytochemical screening of the leaf extracts showed that leaf extracts contain flavonoids, tannins, alkaloids, and steroids; however, there is an absence of saponins, anthraquinones, and cyanogenic glycosides. The agar well-diffusion assay also revealed that leaf extracts have inhibitory effects on all test organisms but are relatively lower compared to the control (Chloramphenicol). In contrast, aqueous leaf extract showed no inhibitory effects. Furthermore, the MTT cell viability assay demonstrated that leaf extracts inhibited the growth of HCT116 effectively with a half-maximal inhibitory concentration (IC50) of 141.57 ppm than the aqueous extract with an IC50 of 506.66 ppm. This study demonstrated that the use of A. philippinensis has a pharmacological basis as an ethnomedicine and could be vital in drug discovery.

Keywords

• Agar well-diffusion
• IC50
• MTT assay
• Medicinal plant

References

1. Abubakar, M.A., Zulkifli, R.M., Hassan, W.N.A.W., Shariff, A.H.M., Malek, N.A.N.N., Zakaria, Z., & Ahmad, F. (2015). Antibacterial properties of Persicaria minor (Huds.) ethanolic and aqueous-ethanolic leaf extracts. Journal of Applied Pharmaceutical Science, 5(Suppl 2), 50–56. https://doi.org/10.7324/JAPS.2015.58.S8
2. Aguinaldo, A.M., Espeso, E.I., Guevara, B.Q. & M.G. Nonato. (2005). Phytochemistry. In: Guevara B.Q. [ed.] A Guidebook to Plant Screening: Phytochemical and Biological. University of Santo Tomas, Manila, Philippines.
3. Al Farraj, D.A., Ragab Abdel Gawwad, M., Mehmood, A., Alsalme, A., Darwish, N.M., Al-Zaqri, N., & Warad, I. (2020). In-vitro antimicrobial activities of organic solvent extracts obtained from Dipcadi viride (L.) Moench. Journal of King Saud University - Science, 32(3), 1965–1968. https://doi.org/10.1016/j.jksus.2020.01.007
4. Alonso-Castro, A.J., Domínguez, F., Ruiz-Padilla, A.J., Campos-Xolalpa, N., Zapata-Morales, J.R., Carranza-Alvarez, C., & Maldonado-Miranda, J.J. (2017). Medicinal plants from north and central america and the caribbean considered toxic for humans: The other side of the coin. Evidence Based Complementary and Alternative Medicine, 2017, 9439868. https://doi.org/10.1155/2017/9439868
5. Amna, U., Halimatussakdiah, Wahyuningsih, P., Saidi, N., & Nasution, R. (2019). Evaluation of cytotoxic activity from Temurui (Murraya koenigii [Linn.] Spreng) leaf extracts against HeLa cell line using MTT assay. Journal of Advanced Pharmaceutical Technology and Research, 10(2), 51–55. https://doi.org/10.4103/japtr.JAPTR_373_18
6. Bhuyan, D.J., Vuong, Q.V., Chalmers, A.C., van Altena, I.A., Bowyer, M.C., & Scarlett, C.J. (2017). Phytochemical, antibacterial and antifungal properties of an aqueous extract of Eucalyptus microcorys leaves. South African Journal of Botany, 112, 180–185. https://doi.org/10.1016/j.sajb.2017.05.030
7. Borah, P.J., Borah, D., Das, U., Das, T.J., & Sarma, R. (2021). A review on ethnopharmacological utility, traditional knowledge and phytochemistry of Aristolochia species in assam, india. Notulae Scientia Biologicae, 13(3), 1 18. https://doi.org/10.15835/nsb13311027
8. Carag, H., and Buot Jr, I.E. (2017). A checklist of the orders and families of medicinal plants in the Philippines. Sylvatrop, The Technical Journal of Philippine Ecosystems and Natural Resources, 27(1 & 2), 40–83.

9. Das, A.K., Islam, M.N., Faruk, M.O., Ashaduzzaman, M., & Dungani, R. (2020). Review on tannins: Extraction processes, applications and possibilities. South African Journal of Botany, 135, 58–70. https://doi.org/10.1016/j.sajb.2020.08.008
10. Demetillo, M.T., Baguio, M.L., Uy, M.M., & Nuñeza, O.M. (2017). Phytochemical analysis and cytotoxic activity of ant plant (Myrmecodia tuberosa Jack.). J Bio Env Sci, 2017(6), 85–91.
11. De Padua, L.S., Lugod, G.C., & Pancho, J.V. (1981). Handbook of Philippine Medicinal Plants. University of the Philippines-Los Baños.
12. Dubey, A., Yadav, P., Peeyush, Verma, P., & Kumar, R. (2022). Investigation of proapoptotic potential of Ipomoea carnea leaf extract on breast cancer cell line. Journal of Drug Delivery and Therapeutics, 12(1), 51–55. https://doi.org/10.22270/jddt.v12i1.5172
13. Eloff, J.N. (1999). The antibacterial activity of 27 southern African members of the Combretaceae. South African Journal of Science, 95(3), 148–152.
14. Ezemokwe, G.C., Aguiyi, J.C., & Chollom, F.P. (2020). The antibacterial activity of aqueous and ethanolic leaf extracts of Balanites aegyptiaca (l.) del plant on some selected clinical human pathogens. Journal of Advances in Microbiology, 20(10), 51–66. https://doi.org/10.9734/jamb/2020/v20i1030290
15. Lawal, F., & Tshikalange T.E. (2020). Chapter 43: Terminalia prunioides. Elsevier.
16. Garcia, J.S., Borja, N., Sevilla, J.B., Villanueva, J.D., & Peyraube, N. (2018). An ethnobotanical study of medicinal plants and perceptions on plant biodiversity conservation in Leyte, Philippines. Journal of Human Ecology, 7(1), 26–42.
17. Gutiérrez-Venegas, G., Gómez-Mora, J.A., Meraz-Rodríguez, M.A., Flores-Sánchez, M.A., & Ortiz-Miranda, L.F. (2019). Effect of flavonoids on antimicrobial activity of microorganisms present in dental plaque. Heliyon, 5(12), e03013 e03018. https://doi.org/10.1016/j.heliyon.2019.e03013
18. International Agency for Reseach on Cancer. (2002). IARC Monographs on the Evaluation of Carcinogenic Risks to Humans, Volume 82, Traditional Herbal Medicines, Some Mycotoxins, Naphthalene and Styrene. IARC Press, Lyon, France.
19. Kaci, F.N., Rüzgar, D., Görmez, A., & Efe, D. (2021). The Evaluation of cytotoxic and antibacterial activity of the ethanol extract of Punica granatum L. peels. Journal of the Institute of Science and Technology, 11(3), 2319–2327. https://doi.org/10.21597/jist.875449
20. Kosasih, K., Sumaryono, W., & Supriono, A. (2019). Cytotoxicity of ethyl acetate extract of Cantigi (Vaccinium varingiaefolium (Blume) Miq. young leaves on Artemia salina L. larvae, MCF-7, T47D, and vero cell lines. Journal of Pharmacognosy and Phytochemistry, 8(4), 24–33.
21. Kuete, V. (2014). Health effects of alkaloids from African medicinal plants: Toxicological survey of African medicinal plants. Elsevier
22. Lerma-Herrera, M.A., Beiza-Granados, L., Ochoa-Zarzosa, A., López-Meza, J.E., Navarro-Santos, P., Herrera-Bucio, R., Aviña-Verduzco, J., & García-Gutiérrez, H.A. (2022). Biological activities of organic extracts of the Genus Aristolochia: A review from 2005 to 2021. Molecules, 27(12), 1–19. https://doi.org/10.3390/molecules27123937
23. Madjos, G., & Ramos, K. (2021). Ethnobotany, systematic review and field mapping on folkloric medicinal plants in the Zamboanga Peninsula, Mindanao, Philippines. Journal of Complementary Medicine Research, 12(1), 21. https://doi.org/10.5455/jcmr.2021.12.01.05
24. Marjoni, M.R., Zulfisa, A. (2017). Antioxidant activity of methanol extract / fractions of senggani leaves (Melastoma candidum D. Don). Pharmaceutica Analytica Acta. 8(8), 1–6. https://doi.org/10.4172/2153-2435.1000557
25. Mathew, L.S., Mtewa, A.G., Ajayi, C.O., Deyno, S., Weisheit, A., Tolo, C.U., Denge, A.L., & Ogwang, P.E. (2020). Ethnopharmacology, pharmacology and phytochemistry of. East Africa Science Journal, 2(1), 23–28.
26. Miya, G.M., Oriola, A.O., Payne, B., Cuyler, M., Lall, N., & Oyedeji, A.O. (2023). Steroids and fatty acid esters from Cyperus sexangularis leaf and their antioxidant, anti-inflammatory and anti elastase properties. Molecules, 28(8), 1 14. https://doi.org/10.3390/molecules28083434
27. Naik, S.K., Mohanty, S., Padhi, A., Pati, R., & Sonawane, A. (2014). Evaluation of antibacterial and cytotoxic activity of Artemisia nilagirica and Murraya koenigii leaf extracts against mycobacteria and macrophages. BMC Complementary and Alternative Medicine, 14, 1–10. https://doi.org/10.1186/1472-6882-14-87
28. Nishino, C., Enoki, N., Mori, A., Kobayashi, K., Fukushima, M., & Tawata, S. (1987). Antibacterial Activity of Flavonoids against Staphylococcus epidermidis, a Skin Bacterium. Agricultural and Biological Chemistry, 51(1), 139 143. https://doi.org/10.1271/bbb1961.51.139
29. Ogunmefun, O.T. (2018). Phytochemicals-God’s Endowment of Curative Power in Plants. https://doi.org/10.5772/intechopen.77423
30. Ogbole, O.O., Segun, P.A., & Adeniji, A.J. (2017). In vitro cytotoxic activity of medicinal plants from Nigeria ethnomedicine on Rhabdomyosarcoma cancer cell line and HPLC analysis of active extracts. BMC Complementary and Alternative Medicine, 17(1), 1–10. https://doi.org/10.1186/s12906-017-2005-8
31. Ozen, T., Bora, N., Yenigun, S., & Korkmaz, H. (2020). An investigation of chemical content, enzyme inhibitory propert, antioxidant and antibacterial activity of Aristolochia bodamae Dingler (develiotu) (Aristolochiaceae) root extracts from Samsun, Turkey. Flavour and Fragrance Journal, 35(3), 270–283. https://doi.org/10.1002/ffj.3559
32. Promega Corporation. (2012). CellTiter 96® Non-Radioactive Cell Proliferation Assay Instructions for Use of Products G4000 AND G4100. Promega Corporation Technical Bulletin.
33. Rajput, A., Sharma, R., Bharti, R. (2022). Pharmacological activities and toxicities of alkaloids on human health. Materials Today: Proceedings, 48(5), 1407 1415. https://doi.org/10.1016/j.matpr.2021.09.189
34. Raju, Renjith, Prakash, T., Rahul, R., Poonangadu, S.S., Senthil Kumar, S., Sonaimuthu, P.T. Chua, J.M., & T. Capili, J. (2021). Phytochemical analysis of three common medicinal plants (Gliricidia sepium, Melothria pendula, and Pithecellobium dulce) in the Philippines. Scholars Academic Journal of Biosciences, 9(3), 84 88. https://doi.org/10.36347/sajb.2021.v09i03.004
35. Saxena, M., Saxena, J., Nema, R., Singh, D., Gupta, A. (2013). Phytochemistry of medicinal plants. Journal of Phamacognosy and Phytochemistry, 1(6), 13 14. https://doi.org/10.1007/978-1-4614-3912-7_4
36. Taid, T.C., Rajkhowa, R.C., & Kalita, J.C. (2014). A study on the medicinal plants used by the local traditional healers of Dhemaji district, Assam, India for curing reproductive health related disorders. Pelagia Research Library Advances in Applied Science Research, 5(1), 296–301.
37. Tan, Z., Deng, J., Ye, Q., Zhang, Z. (2022). The antibacterial activity of natural-derived flavonoids. Current Topics in Medicinal Chemistry, 22(12),1009-1019.
38. Thakur, L., Ghodasra, U., Patel, N., & Dabhi, M. (2011). Novel approaches for stability improvement in natural medicines. Pharmacognosy Reviews, 5(9), 48 54. https://doi.org/10.4103/0973-7847.79099
39. Tong, Z., He, W., Fan, X., & Guo, A. (2022). Biological function of plant tannin and its application in animal health. Frontiers in Veterinary Science, 8(January), 1 7. https://doi.org/10.3389/fvets.2021.803657
40. Ventola, C.L., (2015). The antibiotic resistance crisis: part 1: causes and threats. Pharm Therapeutics, 40(4), 277–283.
41. Wiart, C. (2006). Medicinal plant of Asia and the Pacific. Taylor and Francis Group.
42. World Health Organization. (2023, March 24). https://www.who.int/news-room/fact-sheets/detail/antimicrobial-resistance