TOXICITY TEST OF ROSELLE CALYX EXTRACT ON OSTEOBLASTS CELLS USING MTT ASSAY

Evie Lamtiur Pakpahan

doi:10.32509/mirshus.v5i2.112

Evie Lamtiur Pakpahan Department of Orthodontic, Faculty of Dentistry, University of Prof. Dr. Moestopo (Beragama), Jakarta

DOI: https://doi.org/10.32509/mirshus.v5i2.112

Keywords: Roselle Extract, Osteoblast, MTT Assay

Abstract

Background: Roselle calyx are the part of the roselle plant that is most often used as an extract because it has a lot of bioactive content. Roselle calyx contain dietary fiber, polyphenols, vitamin C, iron, zinc, calcium, magnesium, and potassium, often used as raw materials for herbal medicines. Polyphenols as a whole also have benefits for bone remodeling. Polyphenols can maintain bone integrity by reducing oxidative stress, reducing inflammation through signaling pro-inflammatory and modulation of osteoblastogenesis or osteoclastogenesis through osteoimmunological actions. Methods: Cytotoxicity was evaluated using MTT assay, and data were analyzed using the Shapiro-Wilk normality followed by one-way ANOVA and Tukey’s HSD post hoc test. Results: The result showed that osteoblast cell viability after treatment with roselle calyx extract was 93.95% at 15%, 99.60% at 20%, 108.87% at 25%, and a concentration of 30% had the highest cell viability percentage of 136.69%. Conclusion: Roselle calyx extract is non-toxic to osteoblast cells and has the potential as an alternative natural material to inhibit relapse in orthodontic treatment.

References

Almutary, A., & Sanderson, B. J. S. (2016). The MTT and Crystal Violet Assays: Potential Confounders in Nanoparticle Toxicity Testing. International Journal of Toxicology, 35(4), 454–462. https://doi.org/10.1177/1091581816648906

Cao, L., Wang, J., Zhang, Y., Tian, F., & Wang, C. (2022). Osteoprotective effects of flavonoids: Evidence from in vivo and in vitro studies (Review). Molecular Medicine Reports, 25(6), 1–9. https://doi.org/10.3892/mmr.2022.12716

Fotakis, G., & Timbrell, J. A. (2006). In vitro cytotoxicity assays: Comparison of LDH, neutral red, MTT and protein assay in hepatoma cell lines following exposure to cadmium chloride. Toxicology Letters, 160(2), 171–177. https://doi.org/10.1016/j.toxlet.2005.07.001

Hadi, A. F. N., Aghniya, S. N., Haidar, G. A., Sihombing, W. S. M., Sutedjo, A., & Alhasyimi, A. A. (2024). Post-Orthodontic Relapse Prevention through Administration of a Novel Synthetic Carbonated Hydroxyapatite–Chitosan Hydrogel Derived from Blood Cockle Shell (Anadara granosa L.). Dentistry Journal, 12(1). https://doi.org/10.3390/dj12010018

Ikbal, M., Hasanuddin, H., Mude, A. H., Marlina, E., & Ruslin, M. (2025). Plant-based extracts and biocompatible materials in dental tissue regeneration: An editorial review. Journal of Dentomaxillofacial Science, 10(3), 165–171. https://doi.org/10.15562/jdmfs.v10i3.2115

Jabeur, I., Pereira, E., Barros, L., Calhelha, R. C., Soković, M., Oliveira, M. B. P. P., & Ferreira, I. C. F. R. (2017). Hibiscus sabdariffa L. as a source of nutrients, bioactive compounds and colouring agents. Food Research International, 100, 717–723. https://doi.org/10.1016/j.foodres.2017.07.073

Kamiloglu, S., Sari, G., Ozdal, T., & Capanoglu, E. (2020). Guidelines for cell viability assays. Food Frontiers, 1(3), 332–349. https://doi.org/10.1002/fft2.44

Komala, O., & Rosyanti, R. (2013). Uji Efektivitas Antibakteri Ekstrak Etanol dan Ekstrak Air Kelopak Bunga Rosella (Hibiscus sabdariffa L) Terhadap Bakteri Streptococcus pneumoniae. 3(1).

Krishnan, V., & Davidovitch, Z. (2006). Cellular, molecular, and tissue-level reactions to orthodontic force. American Journal of Orthodontics and Dentofacial Orthopedics, 129(4), 469.e1-469.e32. https://doi.org/10.1016/j.ajodo.2005.10.007

Kusparmanto, F. S., Pakpahan, E. L., Yuditha, S., & Kusparmanto, L. (2024). Study in the Potential of Chemical Compounds of Rosella Flower (Hibiscus Sabdariffa Linn) Extract As a Prevention of Relapse After Orthodontic Treatment. Moestopo International Review on Social, Humanities, and Sciences, 4(1), 78–87. https://doi.org/10.32509/mirshus.v4i1.71

Li, J., Li, Y., Li, M., Lin, L., Qi, J., Xu, J., Zhang, L., Fang, P., & Tao, A. (2022). Novel Insights into Anthocyanin Synthesis in the Calyx of Roselle Using Integrated Transcriptomic and Metabolomic Analyses. International Journal of Molecular Sciences, 23(22), 13908. https://doi.org/10.3390/ijms232213908

Mahadevan, N., Shivali, & Kamboj, P. (2009). Hibiscus sabdariffa Linn. - An overview.

Mariod, A. A., Tahir, H. E., & Mahunu, G. K. (2021). Chapter 5 - Composition of Hibiscus sabdariffa calyx, pigments, vitamins. In A. A. Mariod, H. E. Tahir, & G. K. Mahunu (Eds.), Roselle (Hibiscus sabdariffa) (pp. 69–75). Academic Press. https://doi.org/https://doi.org/10.1016/B978-0-12-822100-6.00002-1

Meeran, N. (2012). Biological response at the cellular level within the periodontal ligament on application of orthodontic force - An update. Journal of Orthodontic Science, 1(1), 2–10. https://doi.org/10.4103/2278-0203.94769

Min Seung, L., So Hyun, L., Ah-Ran, Y., Chi Yeon, H., & Insug, K. (2021). Carfilzomib in Combination with Bortezomib Enhances Apoptotic Cell Death in B16-F1 Melanoma Cells. Biology, 10(2), 153.

Mosmann, T. (1983). Rapid colorimetric assay for cellular growth and survival: Application to proliferation and cytotoxicity assays. Journal of Immunological Methods, 65(1–2), 55–63. https://doi.org/10.1016/0022-1759(83)90303-4

Mumpuni, N. C., Triwahyuni, I. E., Lestari, P. E., Kedokteran, F., Universitas Jember, G., Mulut, B. P., & Mikrobiologi, B. (2021). Efektivitas Ekstrak Bunga Rosella (Hibiscus Sabdariffa L.) Sistemik Terhadap Penyembuhan Ulser Pada Tikus Wistar (Rattus Norvegicus).

Nicolin, V., De Tommasi, N., Nori, S. L., Costantinides, F., Berton, F., & Di Lenarda, R. (2019). Modulatory effects of plant polyphenols on bone remodeling: A prospective view from the bench to bedside. In Frontiers in Endocrinology (Vol. 10). Frontiers Media S.A. https://doi.org/10.3389/fendo.2019.00494

Nurnasari, E., & Khuluq, A. D. (2018). Potensi Diversifikasi Rosela Herbal (Hibiscus sabdariffa L.) untuk Pangan dan Kesehatan. Buletin Tanaman Tembakau, Serat & Minyak Industri, 9(2), 82. https://doi.org/10.21082/btsm.v9n2.2017.82-92

Obouayeba, A. P., Djyh, N. B., Diabate, S., Djaman, A. J., N’Guessan, J. D., Kone, M., & Kouakou, T. H. (2014). Phytochemical and antioxidant activity of roselle (Hibiscus Sabdariffa L.) petal extracts. Research Journal of Pharmaceutical, Biological and Chemical Sciences, 5(2), 1453–1465.

Osei-Kwarteng, M., Gweyi-Onyango, J. P., Komla Mahunu, G., Tahir, H. E., & Apaliya, M. T. (2021). Chapter 6 - Hibiscus sabdariffa: protein products, processing, and utilization. In A. A. Mariod, H. E. Tahir, & G. K. Mahunu (Eds.), Roselle (Hibiscus sabdariffa) (pp. 77–89). Academic Press. https://doi.org/https://doi.org/10.1016/B978-0-12-822100-6.00012-4

Prasetyoputri, A., Rahmawati, S. I., Atikana, A., Izzati, F. N., Hapsari, Y., Septiana, E., Bustanussalam, & Putra, M. Y. (2021). A Mini Review on the Antibacterial Activity of Roselle (Hibiscus sabdariffa L.) Phytochemicals. IOP Conference Series: Materials Science and Engineering, 1192(1), 012017. https://doi.org/10.1088/1757-899x/1192/1/012017

Proffit, W. R., Fields, H. W., & Larson, B. E. (2019). Contemporary orthodontics (6th ed.). Elsevier.

Ramesh, P., Jagadeesan, R., Sekaran, S., Dhanasekaran, A., & Vimalraj, S. (2021). Flavonoids: Classification, Function, and Molecular Mechanisms Involved in Bone Remodelling. Frontiers in Endocrinology, 12(November), 1–22. https://doi.org/10.3389/fendo.2021.779638

Riaz, G., & Chopra, R. (2018). A review on phytochemistry and therapeutic uses of Hibiscus sabdariffa L. In Biomedicine and Pharmacotherapy (Vol. 102, pp. 575–586). Elsevier Masson SAS. https://doi.org/10.1016/j.biopha.2018.03.023

Sari, F., Nurkhasanah, & Bachri, M. S. (2016). Acute toxicity test of rosella (Hibiscus sabdariffa l .) Calyx ethanolic extract on Sprague dawley rats. Traditional Medicine Journal, 21(April), 12–18.

Sharma, A. R., Lee, Y., Bat-ulzii, A., Chatterjee, S., Bhattacharya, M., Chakraborty, C., & Lee, S. (2023). Flavonoids for Bone Loss.

Suniarti, D. F., Suwandi, T., Putri, S. A., & Kurnia, D. (2022). Potential of Hibiscus sabdariffa L. Calyx (Rosella) extract as antibacterial agent in dental disease: Phytochemical and chemical components profiling. Journal of Advanced Pharmaceutical Technology and Research, 13(3), 202–206. https://doi.org/10.4103/japtr.japtr_64_22

Sura, S., Majeed, B. D. S., & Ghani, B. A. (2018). Effect of Topical Application of Flavanoids Extract of Hibiscus Sabdariffa on Experimentally Induced Bone Defect. In J Bagh College Dentistry (Vol. 30, Issue 1).

Tsolakis, I. A., Christopoulou, I., Sitaras, S., Lyros, I., Rontogianni, A., Dalampira, M., & Tsolakis, A. I. (2023). Molecular and Biological Aspects of Orthodontic Tooth Movement: Possibilities for Bioengineering Intervention: A Narrative Review. Bioengineering, 10(11). https://doi.org/10.3390/bioengineering10111275

Weaver, C. M., Alekel, D. L., Ward, W. E., & Ronis, M. J. (2012). Flavonoid Intake and Bone Health. Journal of Nutrition in Gerontology and Geriatrics, 31(3), 239–253. https://doi.org/10.1080/21551197.2012.698220