Advanced Search

Publications > Journals > Journal of Exploratory Research in Pharmacology > Article Full Text


Treatment with Hibiscus sabdarrifa L Anthocyanins Improve Hematological Parameters in Rats Exposed to Cadmium

  • Osuvwe C. Orororo1,*  and
  • Samuel O. Asagba2
 Author information
Journal of Exploratory Research in Pharmacology 2022;():-

doi: 10.14218/JERP.2022.00003


Background and objectives

Cadmium (Cd) is toxic to blood cells and other tissues of the body. This study examined the influence of Hibiscus sabdarrifa anthocyanins (HSA) pretreatment on selected blood parameters in rats administered Cd.


Forty Wistar rats were randomized into eight groups with five rats in each group. The rats were handled in two experimental periods: a five-day acute study and 15-day chronic toxicity study. The experimental groups were the control, Cd, HSA and HSA+Cd groups.


Compared with the healthy control, Cd administration significantly increased the counts of white blood cells (WBC), but decreased red blood cells (RBC), platelets, packed cell volume (PCV), hemoglobin (Hb), mean corpuscular volume (MCV), mean corpuscular hemoglobin concentration (MCHC), and mean corpuscular hemoglobin (MCH) in rats (p < 0.05 for all). In contrast, treatment with HSA significantly reduced the hematological toxicity of Cd in rats by decreasing the counts of WBC, but increasing RBC, PCV, Platelets, Hb, MCV, MCHC, and MCH values in rats.


The results indicate that HSA treatment ameliorates the Cd-caused hematological toxicity in rats and may be valuable for intervention of Cd poisoning.


Cadmium, Hematological parameters, Hibiscus sabdarrifa anthocyanins, Oxidative stress.


Cadmium (Cd) application in industrials in many developing countries has caused environmental pollution, leading to acute and chronic Cd poisoning.1 According to Okorie et al.,2 Cd (and its compounds) is presently regarded as one of the major hazardous pollutants in the world. Cd remains in the environment for a long time and has highly toxic effects. Cd can be inhaled and ingested by the general population, especially workers in industrials, and its accumulation can cause acute and chronic intoxications. Ingestion of Cd-contaminated agricultural products is a major route for human intoxications.3 Cd presents in cigarettes, batteries, paints and other industrial products.4

After ingestion or inhalation, Cd mainly accumulates in the liver, kidneys, and reproductive organs and causes its toxicity. In the liver, Cd causes hepatocyte swelling, necrosis, and degeneration.57 Cd can induce strong oxidative stress and membrane lipid peroxidation that mediate its toxicity in different organs. These effects on the liver and those on other organs of the body have been attributed to Cd-induced oxidative stress, which has been reported as the major means of Cd toxicity.811 Particularly, Cd-toxicity can damage the function and structure of bone and blood cells and inhibit their functional development.12,13 After entering the blood circulation, Cd can bind to red blood cell membranes, where it triggers the production of metallothioneins and increases the generation of reactive oxygen species, thereby damaging the antioxidant defense system and red blood cells.14,15

Previous studies have shown that natural antioxidants can counteract the Cd-induced oxidative stress and in this regard, plant pigments such as anthocyanins and other polyphenols have shown great potentials.16,17 Blood parameters are good indicators of the physiological and nutritional status of animals and reflect the changes in the metabolic processes.18 These parameters can also measure how well nutrients and other food additives are metabolized by living organisms, and the relationship between chemicals, drugs, and various plant extracts and blood functions.19,20

Hibiscus sabdariffa L. (Hs) (Malvaceae) is also known as roselle and is an annual, herbaceous, subshrub with typically red stems and flowers. The plant is largely grown in the tropics and subtropics, such as Thailand, Sudan, West India, Egypt, Mexico and China. In Nigeria, Thailand, Mexico, Egypt and Sudan, roselle juice, made by water-extraction from the calyces, is a soft drink loved and consumed by many people.17 Given its excellent safety profile, this study aimed to investigate the influence of Hibiscus sabdarrifa anthocyanins (HSA) pretreatment on selected blood parameters in rats administered Cd.

Materials and methods


All reagents and chemicals used were standard and analytical grade products.

Plant materials

Fresh calyces of Hibiscus sabdariffa L. were bought from local dealers in Warri, Delta State, and were dried at room temperature under continuous air-flow.

Extraction and purification of HSA

The extraction and purification of HSA were done as described previously.17,21,22

Experimental animals

Forty (40) male Wistar rats weighing 188–192 g were used for the study. They were acclimatized in standard laboratory cages with temperature of 25 ± 2 °C and a 12-hour light/dark cycle for seven days. Standard laboratory and animal care guidelines were applied.

Ethical approval

The study was approved by the Ethic Committee of the Faculty of Science, Delta State University, Abraka (SCI/2015/7b). All applicable international, national, and/or institutional guidelines for the care and use of animals were followed (Supplementary File 1).

Experimental design and treatment of animals

The animals were randomized into eight groups with five rats per group and were handled in two experimental periods: a five-day acute study and 15-day chronic toxicity study. Four groups were used for the 5-day acute toxicity study while the other four groups were used for the 15-day chronic toxicity study. The animals were treated as shown in Table 1.

Table 1

Experimental design

Acute study – 5 days
Control (food and water only)Cd (3 mg/kg body weight)HSA (3 g/kg body weight)HSA Pre-Cd High Dose; HSA (3 g/kg body weight) for 5 consecutive days and a single dose of Cd (5 mg/kg body weight)
Chronic study – 15 days
Control (feed and water only)Cd (3 mg/kg body weight)HSA (3 g/kg body weight)HSA Pre-Cd (HSA 3 g/ kg body weight for 10 consecutive days and Cd 3 mg/kg body weight for 5 days)

Cd and anthocyanins were administered to the animals orally in the morning daily. When the experimental period elapsed, the animals were euthanized. Their blood samples were taken and processed for hematological assays.

Biochemical assay (estimation of hematological parameters)

Platelets count, hemoglobin concentration (Hb), red blood cell count (RBC), white blood cell count (WBC), packed cell volume (PCV), mean corpuscular hemoglobin concentration (MCHC), mean corpuscular hemoglobin (MCH) and mean corpuscular volume (MCV) were assayed using an auto-analyzer.

Analysis of data

The Statistical Package for Social Sciences software was employed and the difference among the groups was analyzed by repeated analysis of variance. A p-value of <0.05 was considered statistically significant. Results are presented as mean ± SD.


Compared with the controls, pretreatment with HSA did not significantly change the values of any hematological parameter, except for significantly increasing the values of RBC and platelets in both models of rats (Table 2). Secondly, administration with Cd caused acute hematological toxicity by significantly increasing the counts of WBC, but decreasing the values of MCH, PCV, platelets, MCV, RBC, Hb, and MCHC in rats.

Table 2

Influence of H sabdariffa L Anthocyanin on hematological parameters/indices of rats exposed to Cd

Acute ExposureWBC (104/mm3)449 ± 0.16785± 0.17*453± 0.22638± 0.29*#
RBC (106/mm3)450 ±0.38203± 0.29*518± 0.46*360 ±0.23*#
Hb (g/dl)10.08±0.1905.02±0.27*12.01 ± 0.4208.25± 0.47*#
PLT (103/mm3)22.01±0.1115.22± 0.09*41.11± 0.38*24.45± 0.22#
PCV (%)40.92±0.1032.53±0.17*44.60± 0.2239.44± 0.29#
MCV (fL)69.14 ± 2.5663.17 ± 1.45*69.74 ± 1.3665.11 ± 1.12*
MCH (pg)19.10 ± 0.6220.13 ± 1.23*17.34 ± 1.24*18.05 ± 1.42#
MCHC (g/L)323.42 ± 4.16301.10 ± 3.22*327.45 ± 4.01311.35 ± 3.66*#
Chronic ExposureWBC (104/mm3)438 ±0.12780±0.32*444±0.20530±0.19*#
RBC (106/mm3)446± 5.45233±5.32*523± 4.47*410± 4.29*#
Hb (g/dl)11.12± 0.1904.92±0.27*11.89± 0.4209.95±0.47*#
PLT (103/mm3)24.0± 0.1114.8± 0.09*43.3±0.37*25.4± 0.22#
PCV (%)44.2± 0.2430.8± 0.27*43.8±0.2338.5±0.32*#
MCV (fL)76.12 ± 1.2668.34 ± 2.53*75.19 ± 1.8470.22 ± 2.42*#
MCH (pg)19.75 ± 2.3415.34 ± 2.31*19.45 ± 2.4117.51 ± 2.23*#
MCHC (g/L)314.22 ± 3.62299.15 ± 3.26*315.15 ± 3.12308.45 ± 3.36*#

Pretreatment with HSA significantly mitigated the Cd-changed values of these hematological parameters in both models of rats, except for insignificantly increasing MCH values in the acute toxicity model of rats.


Hematological parameters can be used for evaluation of nutritional status of individuals and toxicity profile of drugs, chemicals, and food products.9,23 In this study, we found that Cd administration significantly increased the counts of WBC, but decreased the values of Hb, platelets, RBC, PCV, MCV, MCH, and MCHC in rats, consistent with previous observations.6,9,24,25 Similarly, Cd administration has been reported to reduce the values of Hb, PCV, and platelets in rats,10 although this study did not detect the reduced values of MCH and MCV in rats. However, the reduced values of MCV and MCHC by Cd were also reported by İhsan et al.26

Mechanistically, the Cd-induced anaemia may be mediated by intravascular hemolysis, direct damage of Cd to erythrocytes, and iron depletion in the liver, spleen and other tissues.27,28 In addition, Ladokun et al.,29 have noted that the decreased counts of RBC can be explained by a corresponding decrease in MCV and MCH, which determine the size of RBCs. Thus, the decreased values of MCV and MCH by Cd exposure may have resulted in a decrease in RBC counts and Hg contents in rats. The results also agree with a previous study that found that Cd induced microcytic hypochromic anemia in rats.30

The insignificant changes in PCV, WBC, and Hb values and significant changes in RBC and platelets values caused by administration of HSA are similar to some previous reports, but not in agreement with others. Olatunji et al.31 and Famurewa et al.32 indicated that treatment with H sabdariffa calyx aqueous extract failed to significantly change the values of Hb, RBC, platelets, and PCV in rats. On the other hand, Ejere et al.33 reported that treatment with H sabdariffa calyces significantly increased the values of these parameters in rats. Nevertheless, we found that treatment with HSA significantly increased the counts of RBC, pointing to the ability of HSA to enhance the RBC development. This, together with increased values of PCV and Hb, suggests that HSA has blood boosting properties. It is possible that administration of HSA may accelerate RBC development or limit RBC destruction in rats. The therapeutic effect of HSA indicates that HSA may have potentials as a health promoting molecule in ameliorating Cd toxicity.

Our findings were also consistent with previous reports that treatment with HS extracts elevates the levels of Hb, RBC, and PCV.34,35 Interestingly, these studies have highlighted the importance of antioxidant anthocyanins, flavonoids, and polyphenolic acid in the medicinal effects of Hibiscus sabdariffa.36 Therefore, our findings provide new evidence to demonstrate that HSA has potent antioxidant activity to mitigate the Cd-hematologic toxicity.

Future directions

The development of red and white blood cells is regulated with great precision in healthy humans, but the number of leukocytes greatly rapidly increases during infection. Chronic Cd intoxication commonly induces anemia, which may be due to intravascular hemolysis or to the direct damaging effect of Cd on erythrocytes. The exact mechanism underlying the hematologic toxicity of Cd remains to be examined. We are also interested in further investigating how HSA treatment significantly increases WBC counts in rats and whether HSA treatment can also promote the development maturation of other types of blood cells.


In this study, our data indicated that Cd administration significantly increased the counts of WBC, but decreased the values of other hematologic parameters we tested in rats, and pretreatment with HSA significantly increased RBC and Hb values in healthy rats and mitigated the Cd-altered hematological parameters we tested in rats. These results suggest that HSA may be valuable for the intervention of Cd-mediated hematologic toxicity. Our findings also indicated that anthocyanins had potent antioxidant activity against oxidative stress in the pathogenesis of many diseases.t

Supporting information

Supplementary material for this article is available at https://doi.org/10.14218/JERP.2022.00003 .

Supplementary File 1

ARRIVE Checklist.








Hibiscus sabdarrifa anthocyanins


mean corpuscular hemoglobin


mean corpuscular hemoglobin concentration


mean corpuscular volume


packed cell volume


red blood cells


white blood cells




Ethical statement

Ethical approval for the study was obtained from the Ethic Committee of the Faculty of Science, Delta State University, Abraka (SCI/2015/7b). All applicable international, national, and/or institutional guidelines for the care and use of animals were followed.

Data sharing statement

The data used to support the findings of this study are available from the corresponding author



Conflict of interest

The authors declare no competing interests.

Authors’ contributions

OOC and ASO designed the experiment, OOC carried out the laboratory work, OOC and ASO wrote and edited the article.


  1. Rani A, Kumar A, Lal A, Pant M. Cellular mechanisms of cadmium-induced toxicity: a review. Int J Environ Health Res 2014;24(4):378-399 View Article PubMed/NCBI
  2. Okorie E, Olorunfemi CI. Monitoring the distribution of cadmium in sediment samples from Obajana stream in North central Nigeria. Int J Biol Chem Sci 2014;8(4):1948-1954 View Article
  3. Anetor JI. Rising Environmental cadmium levels in developing countries: threat to genome stability and health. J Environ Anal Toxicol 2012;2(4):1-9 View Article
  4. Nordberg GF, Nogawa K, Nordberg M, Friberg LT. Foreword: Metals—A new old environmental problem and Chapter 23: Cadmium. Handbook on the Toxicology of Metals. 3rd ed. Academic Press; 2011:446-451
  5. El-Sokkary GH, Awadalla EA. The protective role of vitamin C against cerebral and pulmonary damage induced by cadmium chloride in male adult albino rat. Open Neuroendocrinol J 2011;4:1-8 View Article
  6. Ibrahim MA, Almaeen AH, El-Moneim MA, Tammam HG, Khalifa AM, Nasibe MN. Cadmium-induced hematological, renal, and hepatic toxicity: The amelioration by spirulina platensis. Saudi J Forensic Med Sci 2018;1(1):5-13 View Article
  7. Kouadria M, Djemli S, Tahraoui A. Hepatoprotective effect of Zinc and Magnesium against subchronic Cadmium toxicity on biochemical, histopathological and neurobehavioral parameters in Wistar rats. J Anim Behav Biometeorol 2020;8:63-73 View Article
  8. Vesey DA. Transport pathways for cadmium in the intestine and kidney proximal tubule: focus on the interaction with essential metals. Toxicol Lett 2010;198(1):13-19 View Article PubMed/NCBI
  9. Andjelkovic M, Buha Djordjevic A, Antonijevic E, Antonijevic B, Stanic M, Kotur-Stevuljevic J, et al. Toxic Effect of Acute Cadmium and Lead Exposure in Rat Blood, Liver, and Kidney. Int J Environ Res Public Health 2019;16(2):E274 View Article PubMed/NCBI
  10. Innih SO, Eluehike N, Francis B. Effects of aqueous extract of Cyperusesculentus (tiger nut) on antioxidant status and hematological indices in the heart of cadmium-induced wistar rats. Niger J Exp Clin Biosci 2021;9(1):17-24 View Article
  11. Mafulul SG, Okoye ZSC. Protective effect of pre-supplementation with selenium on cadmium-induced oxidative damage to some rat tissues. Int J Biol Chem Sci 2012;6(3):1128-1138 View Article
  12. Bodo M, Balloni S, Lumare E, Bacci M, Calvitti M, Dell’Omo M, et al. Effects of sub-toxic Cadmium concentrations on bone gene expression program: results of an in vitro study. Toxicol In Vitro 2010;24(6):1670-1680 View Article PubMed/NCBI
  13. Radhakarishan MV. Immunological effect of cadmium in heteropneus fossils bloch. Global Vet 2010;4(6):544-547
  14. McCarty MF. Zinc and multi-mineral supplementation should mitigate the pathogenic impact of cadmium exposure. Med Hypotheses 2012;79(5):642-648 View Article PubMed/NCBI
  15. Olajide JE, Momoh S, Achimugu OJ, Jegede ER. Effects of Tremaorientalis Leaves Extract on Hematological Parameters of Cadmium Induced Toxicity in Wistar Rats. IOSR J Biotech Biochem 2020;6(1):1-7 View Article
  16. Orororo OC, Asagba SO, Tonukari NJ, Okandeji OJ, Mbanugo JJ. Hibiscus sabdarrifa L. anthocyanins-induced changes in reproductive hormones of cadmium-exposed rats. Inter J Sci Res Pub 2018;8(4):308-311 View Article
  17. Orororo OC, Asagba SO, Tonukari NJ, Okandeji OJ, Mbanugo JJ. Effects of Hibiscus Sabdarrifa L. anthocyanins on cadmium-induced oxidative stress in Wistar rats. J Appl Sci Environ Management 2018;22(4):465-470 View Article
  18. Hounkpatin ASY, Edorh PA, Guédénon P, Alimba CG, Ogunkanmi A, Dougnon TV, et al. Haematological evaluation of Wistar rats exposed to chronic doses of cadmium, mercury and combined cadmium and mercury. Afri J Biotech 2013;12(23):3731-3737 View Article
  19. Majid T, Mohsen T, Abas AG, Sayed AT. Performance, immunity, serum biochemical and haematological parameters in broiler chicks fed dietary thyme as alternative for an antibiotic growth promoter. Afri J Biotech 2010;9(40):6819-6825 View Article
  20. Lodia S, Kansala L. Antioxidant activity of rubiacordifolia against lead toxicity. Int J Pharmacol Sci Res 2012;3(7):2224-2232 View Article
  21. Hong V, Wrolstad RE. Use of HPLC separation/photodiode array detection for characterization of anthocyanins. J Agric Food Chem 1990;38:708-715 View Article
  22. Ologundudu A, Ologundudu AO, Ololade IA, Obi FO. Effect of Hibiscus sabdariffa anthocyanins on 2,4-dinitrophenylhydrazine-induced hematotoxicity in rabbits. Afri J Biochem Res 2010;3(4):140-144
  23. Vinodini NA, Pratik KC, Kunal SVB, Rashmi KS, Nayanatara AK. Effectiveness of Moringaoleifea extract in attenuating the toxic effect on platelet count: An experiment on Cadmium exposed rats. Pharmacog J 2019;11(4):689-693 View Article
  24. Onwuka FC, Erhabor O, Eteng MU, Umoh IB. Ameliorative effect of cabbage extract on cadmium-induced changes on hematology and biochemical parameters of albino rats. J Toxico Environ Health Sci 2010;2(2):11-16
  25. Embugushiki RE, Mafulul SG, Okoye ZSC. Protective effect of carrot juice pretreatment on cadmium-induced oxidative cytotoxic damage to some rat tissues. J Pharm Bio Sci 2013;7(6):55-62
  26. İhsan K, Mehmet F, Aydınb İÜ. Partial protective effects of melatonin on cadmium-induced changes in hematological characteristics in rats. Biotechnic & Histochemistry 2021;4(2):15-23 View Article
  27. Taqa GA, Mamdoh JK. Protective effect of injectable iron on cadmium-induced anemia in rats. J Edu Sci 2009;22(3):30-35 View Article
  28. Min KS, Ueda H, Tanaka K. Involvement of intestinal calcium transporter 1 and metallothionein in cadmium accumulation in the liver and kidney of mice fed a low-calcium diet. Toxicol Lett 2008;176(1):85-92 View Article PubMed/NCBI
  29. Ladokun O, Ojezele M, Arojojoye O. Comparative study on the effects of aqueous extracts of viscum album (mistletoe) from three host plants on hematological parameters in albino rats. Afr Health Sci 2015;15(2):606-612 View Article PubMed/NCBI
  30. Ashour TH. Preventative effects of caffeic Acid phenyl ester on cadmium intoxication induced hematological and blood coagulation disturbances and hepatorenal damage in rats. ISRN Hematol 2014;2014:764754 View Article PubMed/NCBI
  31. Olatunji LA, Adebayo JO, Akinola OB, Olatunji VA, Adekoya A, Badaki OJ, et al. Haematological effects of aqueous extract of Hibiscus sabdariffa petals in rats. Trop J Health Sci 2005;12:44-45 View Article
  32. Famurewa AC, Kanu SC, Ogugua VN, Nweke ML. Protective effect of pretreatment of rats with calyx extract of Hibiscus sabdariffa against carbon tetrachloride-induced hematotoxicity. J Biol Sci 2015;15(3):138-143 View Article
  33. Ejere VC, Nnamonu EI, Chukwuka CO, Ugwu GC, Ejim AO, Asogwa CN. Effects of aqueous extract of Hibiscus sabdariffa calyces on haematological characteristics of Rattus novergicus. Animal Res Int 2013;10(3):1809-1816
  34. Olusola AO, Olusola AO, Bada SO, Obi FO. Comparative study on the effect of Hibiscus sabdariffa calyx anthocyanins and ascorbate on 2,4-dinitrophenylhydrazine-induced damage in rabbits. Am J Biochem 2012;2:1-6 View Article
  35. Agbai EO, Nwanegwo CO. Effect of methanolic extract of Hibiscus sabdariffa on some hematological parameters in levodopa-induced anemia. J Biol Sci Bioconserv 2013;5:44-53
  36. Da-Costa-Rocha I, Bonnlaender B, Sievers H, Pischel I, Heinrich M. Hibiscus sabdariffa L. - a phytochemical and pharmacological review. Food Chem 2014;165:424-443 View Article PubMed/NCBI
  • Journal of Exploratory Research in Pharmacology
  • eISSN 2572-5505

Treatment with Hibiscus sabdarrifa L Anthocyanins Improve Hematological Parameters in Rats Exposed to Cadmium

Osuvwe C. Orororo, Samuel O. Asagba
  • Reset Zoom
  • Download TIFF