Acute and Subacute Oral Toxicity Assessment of The Polysaccharides Extracted from Clinacanthus nutans Leaves: A Preclinical Model for Drug Safety Screening
Main Article Content
Abstract
Emerging investigations have indicated that many plant polysaccharides may be beneficial for treating metabolic diseases. To date, the therapeutic efficacy and potential toxicity of polysaccharides extracted from Clinacanthus nutans (C. nutans) remain unexplored. This study investigated the in vivo acute and subacute oral toxicological profiles of the highest doses of C. nutans bioactive polysaccharides (CNBP) extracted from the leaves using conventional toxicity methods. The total of 39 healthy 8–10 weeks male Sprague-Dawley rats (n = 3) were randomly assigned to control (C), acute (A) and subacute (SA) groups receiving 125, 250, 500, 1,000, 2,000 or 3,000 mg/kg/day of CNBP extract, respectively. The acute group received a single dose of CNBP extract, whereas the subacute group received daily single doses of CNBP extract for 14 days. Oral administration of up to 3,000 mg/kg CNBP extract caused no abnormal signs of toxicity during 14 days. However, daily administration of 500 mg/kg or higher doses of CNBP extract for 14 days induced a mild degree of toxicity in the liver, characterised by elevated alkaline phosphatase levels with C (163 ± 9 U/L) vs. SA500 (222 ± 49 U/L), SA1000 (223 ± 29 U/L), SA2000 (238 ± 33 U/L) and SA3000 (252 ± 18 U/L). CNBP extracts exhibit therapeutic potential, exemplified by diuretic, natriuretic, anti-hypertensive, anti-tachycardia, reno-protective and cholesterol-lowering properties. Precautions should be taken when administering the extracts at higher doses and for longer durations.
Article Details

This work is licensed under a Creative Commons Attribution 4.0 International License.
References
Aliyu A, Shaari M R, Ahmad Sayuti N S, Reduan M F H, Sithambaram S, Noordin M M, Khozirah Shaari K and Hamzah H. (2020). Subacute oral administration of Clinacanthus nutans ethanolic leaf extract induced liver and kidney toxicities in ICR mice. Molecules 25(11): 2631. https://doi.org/10.3390/molecules25112631
Asyura S N, Hamzah H, Shaari R M, Sithambaram S and Mustapha N M. (2016). Blood profiles and histopathological changes of liver and kidney tissues from male Sprague Dawley rats treated with ethanol extracts of Clinacanthus nutans leaf. Journal of Clinical Toxicology 6: 329. https://doi.org/10.4172/2161-0495.1000329
Azemi A K, Mokhtar S S and Rasool A H G. (2021). Clinacanthus nutans: Its potential against diabetic vascular diseases. Brazilian Journal of Pharmaceutical Sciences 56: e18838. https://doi.org/10.1590/s2175-97902020000118838
Ba?aran N, Pasl? D and Ba?aran A A. (2022). Unpredictable adverse effects of herbal products. Food and Chemical Toxicology 159: 112762. https://doi.org/10.1016/j.fct.2021.112762
Benalaya I, Alves G, Lopes J and, Silva L R. (2024). A review of natural polysaccharides: Sources, characteristics, properties, food, and pharmaceutical applications. International Journal of Molecular Sciences 25: 1322. https://doi.org/10.3390/ijms25021322
Chia T Y, Gan C Y, Gurjeet K, Pike-See C, Vikneswaran M, Ashfaq A, Bader A, Sulaiman M A A, Muhammad H S, Selvamani N N, Mohammed H A and Edward J J. (2023). Acute and subacute oral toxicity assessment of the polysaccharides extracted from Clinacanthus nutans leaves: A preclinical model for drug safety screening. Authorea preprints.
Chia T Y, Gan C Y, Murugaiyah V, Hashmi S F, Fatima T, Ibrahim L, Mohammed H A, Farhan K A, Edward J J and Ashfaq A. (2021a). A narrative review on the phytochemistry, pharmacology and therapeutic potentials of Clinacanthus nutans (Burm. f.) lindau leaves as an alternative source of future medicine. Molecules 27(1): 139. https://doi.org/10.3390/molecules27010139
Chia T Y, Murugaiyah V, Khan N A K, Sattar M A, Abdulla M H, Edward J J, Ahmad A, Hassan Z, Gurjeet K, Mei H Y, Ahmad F U and Akhtar S. (2021b). Inhibition of L-NAME induced hypertension by combined treatment with apocynin and catalase: The role of Nox 4 expression. Physiological Research 70: 13–26. https://doi.org/10.33549/physiolres.934497
Chia T Y, Murugaiyah V, Sattar M A, Khan N A K, Ahmad A, Abdulla M H, Edward J J, Mei H Y, Akhtar S and Ahmad F U. (2020). The restorative effect of apocynin and catalase in L-arginine-induced hypotension on normotensive subjects: The role of oxidative stress. Physiological Research 69(6): 1051–1066. https://doi.org/10.33549/physiolres.934426
Debelo N, Afework M, Debella A, Makonnen E, Ergete W and Geleta B. (2015). Histopathological and biochemical assessment of chronic oral administration of aqueous leaf extract of Thymus serrulatus in mice. International Journal of Clinical and Experimental Pathology 5: 258. https://doi.org/10.4172/2161-0681.1000258
Farsi E, Esmailli K, Shafaei A, Moradi Khaniabadi P, Al Hindi B, Khadeer Ahamed M B, Doblin S, Munavvar A S, Zhari I, Amin M S A M and Aman S A M. (2016). Mutagenicity and preclinical safety assessment of the aqueous extract of Clinacanthus nutans leaves. Drug and Chemical Toxicology 39: 461–473. https://doi.org/10.3109/01480545.2016.1157810
Haida Z and Hakiman M. (2019). A review of therapeutic potentials of Clinacanthus nutans as source for alternative medicines. Sains Malaysiana 48: 2683–2691. https://doi.org/10.17576/jsm-2019-4812-09
Ismail I and Ilya I M. (2017). Nephroprotective effect of Clinacanthus nutans against cisplatin-induced human kidney cell (PCS-400–010). Planta Medica International Open 4(S 01): S1–S202. https://doi.org/10.1055/s-0037-1608407
Kamarudin F and Gan C Y. (2016). Molecular structure, chemical properties and biological activities of Pinto bean pod polysaccharide. International Journal of Biological Macromolecules 88: 280–287. https://doi.org/10.1016/j.ijbiomac.2016.04.003
Khoo L W, Foong Kow A S, Maulidiani M, Lee M T, Tan C P, Shaari K, Chau Ling Tham C L and Abas F. (2018). Hematological, biochemical, histopathological and 1H-NMR metabolomics application in acute toxicity evaluation of Clinacanthus nutans water leaf extract. Molecules 23(9): 2172. https://doi.org/10.3390/molecules23092172
Liao Y -H, Jones S A, Forbes B, Martin G P and Brown M B. (2005). Hyaluronan: Pharmaceutical characterization and drug delivery. Drug Delivery 12: 327–342. https://doi.org/10.1080/10717540590952555
Liaskou E, Wilson D V and Oo Y H. (2012). Innate immune cells in liver inflammation. Mediators of Inflammation 2012: 1–-22. https://doi.org/10.1155/2012/949157
Mas’ ulun M J, Mustika A and Qurnianingsih E (2021). The effect of Dandang Gendis extract (Clinacanthus nutans) on kidney histopathological features of diabetic rats model. Health Notions 5: 19–22. https://doi.org/10.33846/hn50104
Meneguin A B, Silvestre A L P, Sposito L, de Souza M P C, Sabio R M, Araujo V H S, Cury B S F and Chorilli M. (2021). The role of polysaccharides from natural resources to design oral insulin micro-and nanoparticles intended for the treatment of Diabetes mellitus: A review. Carbohydrate Polymers 256: 117504. https://doi.org/10.1016/j.carbpol.2020.117504
Nsagha D S, Ayima C W, Nana-Njamen T and Assob J C N. (2020). The role of traditional, complementary/alternative medicine in primary healthcare, adjunct to universal health coverage in Cameroon: A review of the literature. American Journal of Epidemiology and Infectious Disease 8(1): 37–47.
Nwokocha C, Owu D, Kinlocke K, Murray J and Delgoda R. (2021). Possible mechanism of action of the hypotensive effect of Peperomia pellucida and interactions between human cytochrome P450 enzymes. Medicinal and Aromatic Plants 1: 4. https://doi.org/10.4172/2167-0412.1000105
Organisation for Economic Co-operation and Development (OECD). (2008). OECD-407. Guidelines for testing chemicals-repeated dose 28-day oral toxicity study in rodents, 1–13. https://ntp.niehs.nih.gov/iccvam/suppdocs/feddocs/oecd/oecdtg407-2008.pdf (accessed on 22 February 2023).
Organisation for Economic Co-operation and Development (OECD). (2001). OECD-423. Guidelines for testing chemicals-acute oral toxicity for acute toxic class method, 1–14. https://ntp.niehs.nih.gov/iccvam/suppdocs/feddocs/oecd/oecd_gl423.pdf (accessed on 21 February 2023).
Posadzki P, Watson L K and Ernst E. (2013). Adverse effects of herbal medicines: An overview of systematic reviews. Clinical Medicine 13(1): 7–12. https://doi.org/10.7861/clinmedicine.13-1-7
Sarega N, Imam M U, Esa N M, Zawawi N and Ismail M. (2016a). Effects of phenolicrich extracts of Clinacanthus nutans on high fat and high cholesterol diet-induced insulin resistance. BMC Complementary and Alternative Medicine 16: 1–11. https://doi.org/10.1186/s12906-016-1049-5
Sarega N, Imam M U, Ooi D -J, Chan K W, Md Esa N, Zawawi N and Ismail M. (2016b). Phenolic rich extract from Clinacanthus nutans attenuates hyperlipidemia-associated oxidative stress in rats. Oxidative Medicine and Cellular Longevity 2016: 137908. https://doi.org/10.1155/2016/4137908
Shafie M H, Yusof R and Gan C Y. (2019). Deep eutectic solvents (DES) mediated extraction of pectin from Averrhoa bilimbi: Optimization and characterization studies. Carbohydrate Polymers 216: 303–311. https://doi.org/10.1016/j.carbpol.2019.04.007
Staines S S. (2011). Herbal medicines: Adverse effects and drug-herb interactions. Journal of the Malta College of Pharmacy Practice 17: 38-42.
Tan H F and Gan C Y. (2016). Polysaccharide with antioxidant, ?-amylase inhibitory and ACE inhibitory activities from Momordica charantia. International Journal of Biological Macromolecules 85: 487–496. https://doi.org/10.1016/j.ijbiomac.2016.01.023
Vasvani S, Kulkarni P and Rawtani D. (2020). Hyaluronic acid: A review on its biology, aspects of drug delivery, route of administrations and a special emphasis on its approved marketed products and recent clinical studies. International Journal of Biological Macromolecules 151: 1012–1029. https://doi.org/10.1016/j.ijbiomac.2019.11.066
Wang C, He Y, Tang X and Li N. (2020). Sulfation, structural analysis, and anticoagulant bioactivity of ginger polysaccharides. Journal of Food Science 85: 2427–2434. https://doi.org/10.1111/1750-3841.15338
Widjaja S S and Ichwan M. (2021). Enhanced cytotoxic effects of Clinacanthus nutans and doxorubicin in combination toward breast cancer cell lines. Journal of Advanced Pharmaceutical Technology and Research 12: 152. https://doi.org/10.4103/japtr.JAPTR_251_20
Zakaria Z A, Rahim M H A, Mohtarrudin N, Kadir A A, Cheema M S, Ahmad Z, Ching S M and Tohid S F M. (2016). Acute and sub-chronic oral toxicity studies of methanol extract of Clinacanthus nutans in mice. African Journal of Traditional, Complementary and Alternative Medicines 13(2): 210–222. https://doi.org/10.4314/ajtcam.v13i2.25