Protective Effects of Citrullus Lanatus Seed Extract on Diclofenac-Induced Splenic Oxidative Stress and Histopathological Alterations in Adult Male Wistar Rats

Authors

Mbah, Chikodili Adolphus

Department of Anatomy, Faculty of Basic Medical Sciences, David Umahi Federal University of Health Sciences, Ebonyi State (Nigeria)

Elemuo, Chukwuebuka Stanley

Department of Anatomy, Faculty of Basic Medical Sciences, Chukwuemeka Odumegwu Ojukwu University, Anambra State (Nigeria)

Udeonu Somtochukwu Jennifer

Department of Anatomy, Faculty of Basic Medical Sciences, Nnamdi Azikiwe University Awka, Nnewi Campus, Anambra State (Nigeria)

Ezejindu, Nnadozie Cosmas

Department of Public Health, Faculty of Allied Health Sciences, David Umahi Federal University of Health Sciences, Ebonyi State (Nigeria)

Onuigbo, Ogochukwu Nancy

Department of Information Systems and Technology, Faculty of Computing, National Open University of Nigeria, Enugu State (Nigeria)

Odo Jude Emeka

Department of Humanities, School of General Studies, State University of Medical and Applied Sciences Igbo-Eno, Enugu State (Nigeria)

Ofoego, Uzozie Chikere

Department of Anatomy, Faculty of Basic Medical Sciences, Nnamdi Azikiwe University Awka, Nnewi Campus, Anambra State (Nigeria)

Article Information

DOI: 10.51244/IJRSI.2026.1306000064

Subject Category: Social science

Volume/Issue: 13/6 | Page No: 925-938

Publication Timeline

Submitted: 2026-05-16

Accepted: 2026-05-21

Published: 2026-06-22

Abstract

Diclofenac, a widely used non-steroidal anti-inflammatory drug (NSAID), has been associated with oxidative stress and tissue injury following prolonged administration. Citrullus lanatus (watermelon) seeds contain bioactive phytochemicals with antioxidant properties that may protect against oxidative damage. This study investigated the effect of ethanolic seed extract of Citrullus lanatus on oxidative stress markers and splenic histology in diclofenac-induced splenic injury in male Wistar rats. Twenty-five adult male Wistar rats were randomly assigned into five groups (A–E; n = 5). Group A served as the control; Group B received 100 mg/kg body weight of diclofenac; Group C received 300 mg/kg body weight of C. lanatus extract; while Groups D and E received diclofenac concurrently with 150 mg/kg and 300 mg/kg body weight of the extract, respectively. Treatments were administered orally for 28 days. Body weight, relative spleen weight, serum malondialdehyde (MDA), and superoxide dismutase (SOD) levels were assessed, while splenic histology was evaluated using hematoxylin and eosin staining. Diclofenac administration resulted in reduced body weight gain, elevated MDA levels, decreased SOD activity, and severe splenic alterations characterized by hemorrhage and fibrosis. Treatment with C. lanatus extract reduced MDA levels and significantly improved SOD activity compared with the diclofenac-only group. Histological examination revealed preservation of normal splenic architecture in rats treated with the extract alone and attenuation of splenic lesions in co-treated groups. The findings suggest that ethanolic seed extract of Citrullus lanatus exerts protective effects against diclofenac-induced splenic injury, possibly through its antioxidant activity and enhancement of endogenous antioxidant defenses.

Keywords

Diclofenac; Citrullus lanatus; spleen; oxidative stress; malondialdehyde; superoxide dismutase

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References

1. Hirsa, M., Fichna, J., & Tarasiuk-Zawadzka, A. (2025). Phytotherapy with Fruit Seed Extracts as a Promising Approach for the Treatment of Inflammation. Current nutrition reports, 14(1), 100. https://doi.org/10.1007/s13668-025-00695-4 [Google Scholar] [Crossref]

2. Nikolakakis, E., Ofrydopoulou, A., Shiels, K., Saha, S. K., & Tsoupras, A. (2026). In Vitro Antioxidant, Anti-Platelet and Anti-Inflammatory Natural Extracts of Amphiphilic Bioactives from Organic Watermelon Juice and Its By-Products. Metabolites, 16(1), 81. https://doi.org/10.3390/metabo16010081 [Google Scholar] [Crossref]

3. Messaoudi, S., Tebibel, S., Beladjila, A. K., Touhami, F. K., & Kabouche, Z. (2019). Anti-hyperlipidemic, anti-inflammatory and antioxidant activities of Citrullus lanatus. World Journal of Environmental Biosciences, 8(1), 100–106. [Google Scholar] [Crossref]

4. Sorokina, M., McCaffrey, K. S., Deaton, E. E., Ma, G., Ordovás, J. M., Perkins-Veazie, P. M., Steinbeck, C., Levi, A., & Parnell, L. D. (2021). A Catalog of Natural Products Occurring in Watermelon-Citrullus lanatus. Frontiers in nutrition, 8, 729822. https://doi.org/10.3389/fnut.2021.729822 [Google Scholar] [Crossref]

5. Bamidele, T. O., Sunday, H. G., Mathew, A., Ombugadu, J., & Maryam, A. (2021). Evaluation of the Phytochemicals, Nutritional and Anti-nutritional Compositions of Fresh, Sprouted and Toasted Citrullus lanatus (Watermelon) Seed Extracts. Asian Journal of Biochemistry, Genetics and Molecular Biology, 7(3), 11–19. https://doi.org/10.9734/ajbgmb/2021/v7i330174 [Google Scholar] [Crossref]

6. Collins, J. K., Wu, G., Perkins-Veazie, P., Spears, K., Claypool, P. L., Baker, R. A., Clevidence, B. A. (2007). Watermelon consumption increases plasma arginine concentrations in adults. Nutrition, 23(3), 261–266. https://doi.org/10.1016/j.nut.2007.01.005 [Google Scholar] [Crossref]

7. Nissar, J., Sidiqi, U. S., Dar, A. H., & Akbar, U. (2025). Nutritional composition and bioactive potential of watermelon seeds: A pathway to sustainable food and health innovation. Sustainable Food Technology, 3(2), 375–395. https://doi.org/10.1039/D4FB00335G [Google Scholar] [Crossref]

8. Alfaro, R. A., & Davis, D. D. (2023). Diclofenac. In StatPearls. StatPearls Publishing. https://www.ncbi.nlm.nih.gov/books/NBK557879/ [Google Scholar] [Crossref]

9. Gan, T. J. (2010). Diclofenac: An update on its mechanism of action and safety profile. Current Medical Research and Opinion, 26(7), 1715–1731. https://doi.org/10.1185/03007995.2010.486301 [Google Scholar] [Crossref]

10. Elnashar, A. A., Kamal, H. I., Magdy, M. A., Gamal, T., Hamid, L., Ragab, S. F. M., Khodier, S. A., Mohamed, A., Kamel, M. M., El-Shahawy, A. A., & Yehia, D. A. Y. (2024). Review on diclofenac toxicities in different organs. Ain Shams Journal of Forensic Medicine and Clinical Toxicology, 42(1), 10–24. https://doi.org/10.21608/AJFM.2024.333661 [Google Scholar] [Crossref]

11. Thai, P. N., Ren, L., Xu, W., Overton, J., Timofeyev, V., Nader, C. E., Haddad, M., Yang, J., Gomes, A. V., Hammock, B. D., Chiamvimonvat, N., & Sirish, P. (2023). Chronic diclofenac exposure increases mitochondrial oxidative stress, inflammatory mediators, and cardiac dysfunction. Cardiovascular Drugs and Therapy, 37(1), 25–37. https://doi.org/10.1007/s10557-021-07253-4 [Google Scholar] [Crossref]

12. Yoo, S., Noh, J. H., Lee, H. S., Lee, S. H., Choi, E., Kim, D. I., Min, S. E., Han, K. H., & Kim, S. K. (2025). Toxicity of diclofenac sodium salt after two weeks of daily intramuscular administration in cynomolgus monkeys. Toxicological research, 41(3), 279–290. https://doi.org/10.1007/s43188-025-00281-4 [Google Scholar] [Crossref]

13. Bronte, V., & Pittet, M. J. (2013). The spleen in local and systemic regulation of immunity. Immunity, 39(5), 806–818. https://doi.org/10.1016/j.immuni.2013.10.010 [Google Scholar] [Crossref]

14. Lewis, S. M., Williams, A., & Eisenbarth, S. C. (2019). Structure and function of the immune system in the spleen. Science immunology, 4(33), eaau6085. https://doi.org/10.1126/sciimmunol.aau6085 [Google Scholar] [Crossref]

15. Gabr, A., Mohamed, A. M., Abou Khalil, N. S., & Sayed, A. E.-D. H. (2025). The protective effect of Chlorella vulgaris against diclofenac toxicity in Clarias gariepinus: Haemato-immunological parameters and spleen histological features as outcome markers. Frontiers in Immunology, 16, Article 1566496. https://doi.org/10.3389/fimmu.2025.1566496 [Google Scholar] [Crossref]

16. National Research Council. (2011). Guide for the care and use of laboratory animals (8th ed.). The National Academies Press. https://doi.org/10.17226/12910 [Google Scholar] [Crossref]

17. Lorke, D. (1983). A new approach to practical acute toxicity testing. Archives of Toxicology, 54(4), 275–287. https://doi.org/10.1007/BF01234480 [Google Scholar] [Crossref]

18. Parasuraman, S., Raveendran, R., & Kesavan, R. (2010). Blood sample collection in small laboratory animals. Journal of Pharmacology & Pharmacotherapeutics, 1(2), 87–93. https://doi.org/10.4103/0976-500X.72350 [Google Scholar] [Crossref]

19. Misra, H. P., & Fridovich, I. (1972). The role of superoxide anion in the autooxidation of epinephrine and a simple assay for superoxide dismutase. The Journal of Biological Chemistry, 247(10), 3170–3175. https://doi.org/10.1016/S0021-9258(19)45228-9 [Google Scholar] [Crossref]

20. Abbas, S. S., Schaalan, M. F., Bahgat, A. K., & El-Denshary, E. S. (2014). Possible potentiation by certain antioxidants of the anti-inflammatory effects of diclofenac in rats. The Scientific World Journal, 2014, 731462. https://doi.org/10.1155/2014/731462 [Google Scholar] [Crossref]

21. Daniel, A. O., Imafidon, K. E., & Obayuwana, O. (2021). Nephrotoxic and in vivo antioxidant effects of Citrullus lanatus seed extract. Biomedical Journal of Scientific & Technical Research, 33(5). DOI: 10.26717/BJSTR.2021.33.005473. [Google Scholar] [Crossref]

22. Eke, R., Ejiofor, E., Oyedemi, S., Onoja, S., & Omeh, N. (2021). Evaluation of nutritional composition of Citrullus lanatus Linn. (watermelon) seed and biochemical assessment of the seed oil in rats. Journal of Food Biochemistry, 45(6), e13763. https://doi.org/10.1111/jfbc.13763 [Google Scholar] [Crossref]

23. Azwanida, N. N. (2015). A review on the extraction methods used in medicinal plants: Principles, strength, and limitations. Medicinal & Aromatic Plants, 4(3), 196. https://doi.org/10.4172/2167-0412.1000196 [Google Scholar] [Crossref]

24. Ohkawa, H., Ohishi, N., & Yagi, K. (1979). Assay for lipid peroxides in animal tissues by thiobarbituric acid reaction. Analytical Biochemistry, 95(2), 351–358. https://doi.org/10.1016/0003-2697(79)90738-3 [Google Scholar] [Crossref]

25. Izak-Shirian, F., Najafi-Asl, M., Azami, B., Heidarian, E., Najafi, M., Khaledi, M., & Nouri, A. (2022). Quercetin ameliorates diclofenac-induced renal injury by attenuating oxidative stress and inflammation. European Journal of Inflammation, 20, 1–12. https://doi.org/10.1177/1721727X221086530 [Google Scholar] [Crossref]

26. Moradi, A., Abolfathi, M., Javadian, M., Heidarian, E., Roshanmehr, H., Khaledi, M., & Nouri, A. (2021). Gallic acid exerts nephroprotective and antioxidant effects against diclofenac-induced renal injury in rats. Archives of Medical Research, 52(4), 380–388. https://doi.org/10.1016/j.arcmed.2020.12.005 [Google Scholar] [Crossref]

27. Olukayode, A. G., & Clara, T. F. (2021). Review of studies published on the medicinal importance of different parts of Citrullus lanatus in the last ten years. Journal of Biological Research and Biotechnology, 19(2), 1458–1468. https://doi.org/10.4314/br.v19i2.10 [Google Scholar] [Crossref]

28. Zamuz, S., Munekata, P. E. S., Gullón, B., Rocchetti, G., Montesano, D., & Lorenzo, J. M. (2021). Citrullus lanatus as source of bioactive components: An up-to-date review. Trends in Food Science & Technology, 111, 208–222. https://doi.org/10.1016/j.tifs.2021.03.002 [Google Scholar] [Crossref]

29. Varışlı, B., Çağlayan, C., Kandemir, F. M., Gür, C., Ayna, A., Genç, A., & Taysı, S. (2023). Chrysin mitigates diclofenac-induced hepatotoxicity by modulating oxidative stress, apoptosis, autophagy and endoplasmic reticulum stress in rats. Molecular Biology Reports, 50(1), 433–442. https://doi.org/10.1007/s11033-022-07928-7 [Google Scholar] [Crossref]

30. Okafor, I. J., & Elemuo, C. O. (2018). Histological evaluation of ethanolic extract of watermelon seed on the kidney of alloxan-induced diabetic Wistar rat. International Journal of Medical Science and Applied Biosciences, 3(1), 100–106. http://www.casirmediapublishing.com/wp-content/uploads/2019/09/Pages-64-72-2018-3099.pdf [Google Scholar] [Crossref]

31. Eze, C. U., & Ofoego, U. U. (2022). Effects of watermelon rind extract against potassium bromate-induced damage on the liver and haematological parameters of adult Wistar rats. World Journal of Pharmaceutical and Medical Research, 8(11), 25–32. [Google Scholar] [Crossref]

32. Nnamani, O. E., Ukoha, U., & Ofoego, U. C. (2022). Effect of Citrullus lanatus (watermelon) on semen and testis against acetaminophen-induced toxicity in Wistar rats. International Journal of Innovative Science and Research Technology, 7(11), 1121–1128. [Google Scholar] [Crossref]

33. Del Rio, D., Stewart, A. J., & Pellegrini, N. (2005). A review of recent studies on malondialdehyde as a toxic molecule and biological marker of oxidative stress. Nutrition, Metabolism and Cardiovascular Diseases, 15(4), 316–328. https://doi.org/10.1016/j.numecd.2005.05.003 [Google Scholar] [Crossref]

34. Ofoego, U. C., Ekwujuru, E. U., Ireka, M. I., & Ofoego, A. N. (2020). Ameliorative effect of Aframomum melegueta (alligator pepper) against paraquat-induced testicular damage. World Journal of Pharmaceutical Research, 9(5), 2105–2124. https://doi.org/10.20959/wjpr20205-17442 [Google Scholar] [Crossref]

35. Burke, A., Smyth, E. M., & Fitzgerald, G. A. (2006). Analgesic-antipyretic agents: Pharmacotherapy of gout. In L. L. Brunton, J. S. Lazo, & K. L. Parker (Eds.), Goodman & Gilman's the pharmacological basis of therapeutics (11th ed., pp. 671–716). McGraw-Hill Medical. [Google Scholar] [Crossref]

36. Owumi, S. E., & Dim, U. J. (2019). Biochemical alterations in diclofenac-treated rats: Effect of selenium on oxidative stress, inflammation, and hematological changes. Toxicology Research and Application, 3, 1–11. https://doi.org/10.1177/2397847319874359 [Google Scholar] [Crossref]

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