Research Articles
Vol. 12 No. 4 (2025)
In vitro antimitotic and cytotoxic potentials of aqueous and organic extracts of Rhamnus alaternus: Correlation with chemical composition and in vivo acute oral toxicity evaluation
MPCAE Laboratory - Materials, Natural Substances, Environment and Modeling Team (MSNEM), Polydisciplinary Faculty of Taza (FPT), Sidi Mohamed Ben Abdellah University (USMBA) of Fez, B.P.: 1223 Taza-Gare. Taza, Morocco
MPCAE Laboratory - Materials, Natural Substances, Environment and Modeling Team (MSNEM), Polydisciplinary Faculty of Taza (FPT), Sidi Mohamed Ben Abdellah University (USMBA) of Fez, B.P.: 1223 Taza-Gare. Taza, Morocco
MPCAE Laboratory - Materials, Natural Substances, Environment and Modeling Team (MSNEM), Polydisciplinary Faculty of Taza (FPT), Sidi Mohamed Ben Abdellah University (USMBA) of Fez, B.P.: 1223 Taza-Gare. Taza, Morocco
Abstract
Rhamnus alaternus L. is a shrub known for its beneficial digestive, hepatoprotective, antioxidant, cytotoxic and antihyperglycemic properties. The aim of this study was to evaluate, in vitro, the antimitotic and cytotoxic activities of aqueous (decocted, infused and macerated) and organic (methanolic, methanolic macerated, chloroform, ethyl acetate and petroleum ether) extracts of R. alaternus using two assays: the Lepidium sativum phytotest and the brine shrimp lethality test. Principal component analysis (PCA) was employed to analyze correlations between chemical composition and the two biological activities. An in vivo study of the acute oral toxicity of the most in vitro active plant extracts was carried out on mice. The results of the antimitotic activity study showed that decocted and ethyl acetate extract were the most active, with IC50 values of 25.403 ± 0.153 mg/mL and 6.050 ± 0.037 mg/mL, respectively. Similarly, these two extracts possess the most potent cytotoxic effect with LC50 values of 7.420 ± 0.135 mg/mL and 0.355 ± 0.004 mg/mL, respectively. Correlation analysis between chemical composition and the two in vitro activities revealed a significantly positive correlation. The acute toxicity study of the decocted, methanolic
extract and ethyl acetate extract revealed that all three extracts caused no mortality in mice and consequently, the LD50 was estimated to be ≥ 5000 mg/kg. The results obtained suggest that R. alaternus leaves may have anticancer properties, particularly the decocted, methanolic and ethyl acetate extracts, in addition to demonstrating safety.
References
- 1. Stewart BW, Kleihues P, editors. World cancer report. Vol. 57. Lyon: IARC Press; 2003. p. 182-85.
- 2. Maamri A. Données épidémiologiques sur le cancer dans le monde et au Maroc Revue bibliographique. Ann Sci Santé. 2015;1(1):20-29.
- 3. Bray F, Laversanne M, Sung H, Ferlay J, Siegel RL, Soerjomataram I, et al. Global cancer statistics 2022: GLOBOCAN estimates of incidence and mortality worldwide for 36 cancers in 185 countries. CA Cancer J Clin. 2024;74(3):229-63. https://doi.org/10.3322/caac.21834
- 4. Soerjomataram I, Bray F. Planning for tomorrow: Global cancer incidence and the role of prevention 2020-2070. Nat Rev Clin Oncol. 2021;18(10):663-72. https://doi.org/10.1038/s41571-021-00514-z
- 5. Ferlay J, Colombet M, Soerjomataram I, Mathers C, Parkin DM, Piñeros M, et al. Estimating the global cancer incidence and mortality in 2018: GLOBOCAN sources and methods. Int J Cancer. 2019;144(8):1941-53. https://doi.org/10.1002/ijc.31937
- 6. Organisation mondiale de la Santé (OMS). Observatoire de la santé mondiale. Cancer. 2025 [cited 2025 Jun 12]. Available from: https://www.who.int/news-room/fact-sheets/detail/cancer
- 7. Dangou JM, Sambo BH, Moeti M, Diarra-Nama AJ. Cancer prevention and control in the WHO African region: A plea for action. Afr J Cancer. 2009;1(1):56-60. https://doi.org/10.1007/s12558-008-0008-6
- 8. Ministry of Health, Lalla Salma Foundation – Cancer Prevention and Treatment. National Cancer Prevention and Control Plan 2020-2029. Rabat (Morocco): Ministry of Health; 2020. Available from: https://www.sante.gov.ma/Documents/2021/03/Plan_National_de_Prevention_et_de_Controle_du_Cancer_2020-2029_VF.pdf?csf=1&e=ejDjaj
- 9. Khiari H, Mallekh R, Hsairi M. Strategies of the Maghreb countries in the fight against cancer. Tunis Med. 2021;99(1):148-57.
- 10. Nani S, Benallal M, Hassoune S, Kissi D, Maaroufi A. Involvement of general practitioners in the province Benimellal (Morocco) in screening for cervical cancer. Pan Afr Med J. 2013;14:152. https://doi.org/10.11604/pamj.2013.14.152.2570
- 11. Bisol Â, de Campos PS, Lamers ML. Flavonoids as anticancer therapies: A systematic review of clinical trials. Phytother Res. 2020;34(3):568-82. https://doi.org/10.1002/ptr.6551
- 12. Ouedraogo B, Bayala B, Kaboré B, Zongo PFI, Ouedraogo E, Coulibaly LL, et al. Antioxidant and cytotoxic properties of Combretum paniculatum and Fadogia agrestis extracts on prostate and cervical cancers cell lines. Int J Biol Chem Sci. 2024;17(6):2158 69. https://doi.org/10.4314/ijbcs.v17i6.2
- 13. Belhouala K, Pandiella A, Benarba B. Synergistic effects of medicinal plants in combination with spices from Algeria: Anticancer, antiangiogenic activities, and embrytoxicity studies. J Ethnopharmacol. 2024;118187. https://doi.org/10.1016/j.jep.2024.118187
- 14. Boukef K. Rhamnus alaternus. Essaydali. 2001;81:34-35.
- 15. Ben Ammar R, Kilani S, Bouhlel I, Skandrani I, Naffeti A, Boubaker J, et al. Antibacterial and cytotoxic activities of extracts from (Tunisian) Rhamnus alaternus (Rhamnaceae). Ann Microbiol. 2007;57:453-60. https://doi.org/10.1007/BF03175089
- 16. Ben Ammar R, Kilani S, Bouhlel I, Ezzi L, Skandrani I, Boubaker J, et al. Antiproliferative, antioxidant, and antimutagenic activities of flavonoid-enriched extracts from (Tunisian) Rhamnus alaternus L.: Combination with the phytochemical composition. Drug Chem Toxicol. 2008;31(1):61-80. https://doi.org/10.1080/01480540701688725
- 17. Boussahel S, Speciale A, Dahamna S, Amar Y, Bonaccorsi I, Cacciola F, et al. Flavonoid profile, antioxidant and cytotoxic activity of different extracts from Algerian Rhamnus alaternus L. bark. Pharmacogn Mag. 2015;11(Suppl 1):S102.
- 18. Bouhazama H, Lamchouri F, Toufik H. Phytochemical investigation and quantification of bioactive compounds of Rhamnus alaternus (L.). Trop J Drug Res. 2024;1(1):19-29. https://doi.org/10.26538/tjdr/v1i1.3
- 19. Bouabid K, Lamchouri F, Toufik H, Sayah K, Cherrah Y, Faouzi MEA. Phytochemical screening and in vitro evaluation of alpha amylase, alpha glucosidase and beta galactosidase inhibition by aqueous and organic Atractylis gummifera L. extracts. Plant Sci Today. 2018;5(3):103-12. https://doi.org/10.14719/pst.2018.5.3.393
- 20. Boulfia M, Lamchouri F, Senhaji S, Lachkar N, Bouabid K, Toufik H. Mineral content, chemical analysis, in vitro antidiabetic and antioxidant activities, and antibacterial power of aqueous and organic extracts of Moroccan Leopoldia comosa (L.) Parl. bulbs. Evid Based Complementary Altern Med. 2021;2021(1):9932291.
- 21. Lachkar N, Lamchouri F, Toufik H. In vitro antimitotic and hypoglycemic effect study and acute toxicity assessment of the aqueous and organic extracts of Chamaerops humilis L. var. argentea Andre. BioMed Res Int. 2022;2022(1):4303506. https://doi.org/10.1155/2022/4303506
- 22. Mahtal A, Lamchouri F, Toufik H. Mineral composition, phenolic content and antibacterial activity of aqueous and organic extracts of wild Morchella esculenta (L.) Pers. from the province of Taza, Morocco. Biochem Res Int. 2024;2024(1):6769243. https://doi.org/10.1155/bri/6769243
- 23. Senhaji S, Lamchouri F, Boulfia M, Lachkar N, Bouabid K, Toufik H. Mineral composition, in vitro inhibitory effects of α-amylase, α-glucosidase, β-galactosidase enzymes and antibacterial activity of Ajuga iva subsp. pseudoiva (DC.) Bric. Biointerface Res Appl Chem. 2021;12(2):2373-91. https://doi.org/10.33263/BRIAC122.23732391
- 24. Senhaji S, Lamchouri F, Boulfia M, Bouabid K, Lachkar N, Akabli T, et al. Cell growth inhibition, toxicity assessment, and correlation between chemical composition of aqueous and organic extracts of Ajuga iva subsp. pseudoiva (DC.) Bric. and their biological activities. Biointerface Res Appl Chem. 2022;13(1):58. https://doi.org/10.33263/BRIAC131.058
- 25. Lachkar N, Lamchouri F, Bouabid K, Senhaji S, Boulfia M, Akabli T, et al. In vitro study of the antimitotic power and in vivo acute toxicity of aqueous and organic extracts of the aerial part of Haloxylon scoparium Pomel. and evaluation of the correlation between the chemical profile and their biological activities. Plant Sci Today. 2023;10(1):242-51. https://doi.org/10.14719/pst.2001
- 26. Gagiu F, Nistor C, Danciu F, Cacoveanu A, Luputiu G, Ban C, et al. Mitodepressant substances: A new bio test and its application to some alpha alkylphenyl acetic derivatives. Ann Pharm Fr. 1972;31:363-67.
- 27. Meyer BN, Ferrigni NR, Putnam JE, Jacobsen LB, Nichols DJ, McLaughlin JL. Brine shrimp: A convenient general bioassay for active plant constituents. Planta Med. 1982;45(5):31-34.
- 28. Organisation for Economic Co-operation and Development. Test No. 423: Acute oral toxicity – acute toxic class method. OECD guidelines for the testing of chemicals. Section 4. Paris: OECD Publishing; 2001.
- 29. Paier CRK, Maranhão SSA, Carneiro TR, Lima LM, Rocha DD, Santos RDS, et al. Natural products as new antimitotic compounds for anticancer drug development. Clinics. 2018;73(suppl 1):e813s. https://doi.org/10.6061/clinics/2018/e813s
- 30. Kobayakawa J, Sato-Nishimori F, Moriyasu M, Matsukawa Y. G2-M arrest and antimitotic activity mediated by casticin, a flavonoid isolated from Viticis fructus (Vitex rotundifolia Linne fil.). Cancer Lett. 2004;208(1):59-64. https://doi.org/10.1016/j.canlet.2004.01.012
- 31. Du G, Lin H, Wang M, Zhang S, Wu X, Lu L, et al. Quercetin greatly improved therapeutic index of doxorubicin against 4T1 breast cancer by its opposing effects on HIF-1α in tumor and normal cells. Cancer Chemother Pharmacol. 2010;65:277-87. https://doi.org/10.1007/s00280-009-1032-7
- 32. Gupta PS, Patel S. In vitro antimitotic and cytotoxic potential of plant extracts: A comparative study of Mucuna pruriens, Asteracantha longifolia and Sphaeranthus indicus. Future J Pharm Sci. 2020;6:1-9. https://doi.org/10.1186/s43094-020-00137-8
- 33. Ouzid Y, Kaci-Boudiaf MN, Zeghouini A, Madi AO, Smail-Saadoun N, Houali K. Antimitotic and genotoxic effect of methanolic extracts of leaves of Peganum harmala L. on the meristematic cells of Allium cepa L. Bioagro. 2023;35(2):97-104. http://www.doi.org/10.51372/bioagro352.2
- 34. Mbayo MK, Kalonda EM, Muya RK, Tshisand PT, Kanangila AB, Maseho FM, et al. Test d'activité antimitotique et étude chimique préliminaire de quelques Euphorbiaceae du Katanga méridional (RDC). Phytotherapie. 2018;16(2):74. https://doi.org/10.1007/s10298-016-1060-5
- 35. Tir R, Dutta PC, Badjah‐Hadj‐Ahmed AY. Effect of the extraction solvent polarity on the sesame seeds oil composition. Eur J Lipid Sci Technol. 2012;114(12):1427-38. https://doi.org/10.1002/ejlt.201200129
- 36. Waghulde S, Kale MK, Patil V. Brine shrimp lethality assay of the aqueous and ethanolic extracts of the selected species of medicinal plants. Proceedings. 2019;41(1):47. https://doi.org/10.3390/ecsoc-23-06703
- 37. Mazid MA, Datta BK, Nahar L, Sarker SD. Study of antibacterial activity and toxicity in brine shrimp of two Polygonum species. Ars Pharmaceutica. 2008;49(2):127-34.
- 38. Kamanja IT, Mbaria JM, Gathumbi PK, Mbaabu M, Kabasa JD, Kiama SG. Cytotoxicity of selected medicinal plant extracts using the brine shrimp lethality assay from Samburu County, Kenya. J Med Res. 2018;4(5):249-55.
- 39. Afoulous S, Ferhout H, Raoelison EG, Valentin A, Moukarzel B, Couderc F, et al. Helichrysum gymnocephalum essential oil: Chemical composition and cytotoxic, antimalarial and antioxidant activities, attribution of the activity origin by correlations. Molecules. 2011;16(10):8273-91.
- 40. Tuy-On T, Itharat A, Maki P, Thongdeeying P, Pipatrattanaseree W, Ooraikul B. In vitro cytotoxic activity against breast, cervical, and ovarian cancer cells and flavonoid content of plant ingredients used in a selected Thai traditional cancer remedy: Correlation and hierarchical cluster analysis. Evid Based Complement Altern Med. 2020;2020.
- 41. Rajabi S, Ramazani A, Hamidi M, Naji T. Artemia salina as a model organism in toxicity assessment of nanoparticles. DARU J Pharm Sci. 2015;23:1-6. https://doi.org/10.1186/s40199-015-0105-x
- 42. Tahraoui A, Israili ZH, Lyoussi B. Acute and sub-chronic toxicity of a lyophilised aqueous extract of Centaurium erythraea in rodents. J Ethnopharmacol. 2020;132(1):48-55. https://doi.org/10.1016/j.jep.2010.07.038
- 43. Ouahchia C, Cherif HS, Hamaidi-Chergui F, Marzen L, Deradji S, Hemma R, et al. Toxicité aiguë et subaiguë des extraits méthanoliques d’Inula viscosa L. (Dittrichia viscosa L.). Agrobiologia. 2017;7(2):562-73.
- 44. Duke JA. The green pharmacy: New discoveries in herbal remedies for common diseases and conditions from the world's foremost authority on healing herbs. Emmaus (PA): Rodale Books; 1997.
- 45. Amtaghri S, Farid O, Lahrach N, Slaoui M, Eddouks M. Antihyperglycemic effect of Rhamnus alaternus L. aqueous extract in streptozotocin-induced diabetic rats. Cardiovasc Hematol Disord Drug Targets. 2022;22(4):245 55. https://doi.org/10.2174/1871529X23666230123123317
- 46. Ouchtati S, Seridi R, El-Shora H. Influence of Rhamnus alaternus L leaf extract on normal and alloxane-induced diabetic rats. Biosci Res. 2019;16(2):2009-16.
- 47. Bouabid K, Lamchouri F, Toufik H, Boulfia M, Senhaji S, Faouzi MEA. In vivo anti-diabetic effect of aqueous and methanolic macerated extracts of Atractylis gummifera. Bangladesh J Pharmacol. 2019;14(2):67-73. https://doi.org/10.3329/bjp.v14i2.38870
Downloads
Download data is not yet available.