|Year : 2013 | Volume
| Issue : 2 | Page : 81-86
The effect of Solenostemma argel on anemia related parameters in Albino Rats and Rabbits
HM Osman1, ME Shayoub2, EM Babiker3
1 Department of Biochemistry, University of National Ribat, Sudan
2 Department of Pharmaceutics, Faculty of Pharmacy, University of Khartoum, Sudan
3 Department of Zoology, Faculty of Science, University of Khartoum, Sudan
|Date of Web Publication||10-Sep-2013|
H M Osman
Department of Biochemistry, Faculty of Pharmacy, University of the National Ribat. Khartoum
Source of Support: None, Conflict of Interest: None
Background: The plant has been used for treatment of number of diseases in folklore medicine but not used for anemia. The present study is aimed to investigate its effect on anemia and related blood indices and body weights (BWs). Materials and Methods: Two sets of Albino rats each of four groups of six animals were used in this study. In each set one group for the control and the three groups for experimental doses. Two groups of rabbits each of five animals were also used for control and experiment. In one set of Albino rats, leaves of Solenostemma argel extracted in ethanol were used to test for its effect alone although in the second set, Albino rats were used to test for anemia. However, the rabbits were tested for the effect of S. argel on blood indices and BW. Blood parameters were measured by Sysmex, electrolytes by spectrophotometer, and flame photometer and BW by Mettler balance. Measurements were conducted after 30 days. Results: The present study revealed significant increases (P < 0.05) in mean cell hemoglobin concentration (MCHC) and packed cell volume (PCV) in rats provided with 600 mg/kg and PCV in the group provided with 300 mg/kg, whereas no significant changes in the group provided with 150 mg/kg. For rabbits significant increase (P < 0.05) was restricted in hemoglobin (Hb) and mean cell volume (MCV). However, no significant changes (P > 0.05) occurred in BWs of experimental animals. In the meantime, significant decreases (P < 0.01) in red blood cell counts, Hb, MCHC, PCV and Fe as well as increase in MCV occurred in those treated with Aluminum chloride (AlCl 3 ) alone (Al) group or Aluminum chloride co-administered with 600 mg/ kg of S. argel extract (AlS) group. The results also showed significant increase (P < 0.01) in PCV and MCHC and significant decrease in MCV in group treated with AlCl 3 co-administered with plant when compared to those provided with AlCl 3 alone. Moreover, administration of plant extracts alone (S) group lead to significant increase (P < 0.05) in MCHC, PCV, K, P, and Ca when compared to the control group (CG). Conclusion: The present study concludes that S. argel is of high nutritional values and of significance in mitigating anemia.
Keywords: Aluminum chloride, blood parameters, body weights, medicinal plant, nutritional anemia, Solenostemma argel
|How to cite this article:|
Osman H M, Shayoub M E, Babiker E M. The effect of Solenostemma argel on anemia related parameters in Albino Rats and Rabbits. Saudi J Health Sci 2013;2:81-6
| Introduction|| |
The plant is indigenous to northern Sudan between Berber and Abu Hamad, and found in North Africa.  It belongs to the Asclepiadaceae family. It is an erect shrub, reaches a maximum height of 100 cm, with many velvety, pubescent branches from the base. It is known locally as "Hargal" and widely used in traditional folkloric medicine as antispasmodic,  anti-inflammatory,  anti-rheumatic as well as anti-syphilitic agent. , Smoke inhalation and infusion of the whole Solenostemma argel is also used in treatment of hypercholesterolemia, diabetes mellitus, cold, cough, jaundice and measles. Moreover, it is described for treating gastrointestinal cramp, urinary tract infection and the disturbance of the menstrual cycle. 
The plant also has antimicrobial activity,  it possesses insecticidal effect and hence was used to combat insect pests.  In this context, it was used against mosquito species, the causative agent of malaria in Sudan. ,, Moreover, it was reported to have antimicrobial properties, as well as antibacterial and antioxidant activity. , Moreover, chemical investigations, chromatographic screening and phytochemical as well as tissue culture studies of S. argel leaves, stems, and flowers revealed the presence of numerous biochemical ingredients such as pyrgene glycosides, flavonoids, kaempferol, quercetin, rutin, flavonols, flavanones, chalcones and alkaloids. ,, Of these, pyrgene glycosides were reported to reduce cell proliferation. 
The leaves of this plant; however, are characterized by having a high percentage of carbohydrates (64.8), slightly low percentage of protein (15), low percentage of crude fiber (6.5), crude oil (1.6), about 7.7% as ash, and 4.4% as moisture. This in addition to lower percentage of minerals namely: potassium (0.54%), calcium (0.06%), magnesium (0.03%), sodium (0.01%), cupper (0.0001%), ferrous (0.002%), manganese (0.002%), and lead (0.001). 
On the other hand, aluminum (Al), although in its ionic form has no known biological role, but when accumulates in the body it can induce several clinical disorders such as neurotoxicity,  hepatotoxicity,  bone diseases and anemia.  It also has a direct effect on hematopoiesis  and its high levels in serum of hemodialysis patients were associated with impaired erythropoiesis and iron (Fe)-deficiency anemia. , Furthermore, Aluminum is known to disrupt cellular functions by perturbing Fe homeostasis. 
Al vaccines for children have been linked to a variety of serious autoimmune and inflammatory conditions.  In addition to this, Al one of the environmental xenobiotics that induces free radical-mediated cytotoxicity and reproductive toxicity. 
The present paper is aimed to find out the effect of the ethanolic leaves extract of S. argel on the status of blood parameters, Al-induced anemia as well as on the body weights (BWs) of albino rats and rabbits.
| Materials and Methods|| |
The leaves of S. argel were bought from local market, and placed in ethanol (80%) for 3 days in Soxhlet apparatus. Then, the ethanol extract was dried in Rotary Evaporator apparatus, weighed and dissolved in distilled water to give the final concentration of 150 mg extract/kg of BW, 300 mg extract/kg of BW and 600 mg extract/kg of BW.
For measuring the effect of S. argel on blood parameters and BWs, Albino rats (n = 24, average 235 g) and rabbits (n = 10, local breed, average 530 g) were used. Albino rats were divided into four groups of six animals; first group to act as a control group (CG) and the other three groups to act as experimental groups and denoted (G 1 , G 2 and G 3 ). Similarly, rabbits were divided, but into two groups of five animals; one to act as a control (RG 1 ) and the other as an experimental group (RG 2 ). The control group of Albino rats (CG) was provided with normal diet concentrate (dried meat, milk powder, oil, and flour in some water) without S. argel although the experimental groups of Albino rats were provided, in addition to the concentrate of normal diet, with doses of 150, 300 and 600 of S. argel leaves extract for the three experimental groups of Albino rats (G 1 , G 2 and G 3 ) respectively. The doses were administered orally by Gavage daily for 30 days. For the rabbits, the control group (RG 1 ) was provided with clover leaves only, whereas the experimental group (RG 2 ) was fed with both leaves of clover and of S. argel (2.5 BW) daily for 30 days. For these groups, BW of each animal before and after being experimented was measured using the Mettler sensitive balance (number 202845).
For of Al-induced anemia, only albino rats (n = 24, average BW = 284 g) were used. The rats were also divided into four groups each of six. The 1 st group denoted (Al) was provided with Aluminum chloride (AlCl 3 ) (0.5 mg/kg BW).  The 2 nd group denoted (AlS) was provided with AlCl 3 and 10 min later with S. argel extract (600 mg/kg BW). The 3 rd group denoted (S) was provided with S. argel extract (600 mg/kg BW) alone. The 4 th group denoted (C) was provided with the physiological normal saline and used to act as a control group. All experimental doses were administered orally by Gavage and were given daily for 30 days. These groups were measured for the status of blood as well as for electrolytes.
Both blood parameters and electrolytes were measured. For blood parameters about 2 ml of blood sample was collected by hematocrit capillary tube from retro-orbital plexus of each rat, although about 2 ml from jugular vein of each rabbit. Subsequently, each blood sample was placed in ethylene diamine tetra-acetic acid (EDTA) tube and used immediately for measurement of blood parameters using automated coagulating Sysmex apparatus of the type 8999. The blood parameters included: Hemoglobin (Hb), mean cell volume (MCV), red blood cells count (RBCs), white blood cells count (WBC), mean cell hemoglobin concentration (MCHC), platelets (PLT), lymphocytes (LYM) and packed cell volume (PCV).
For electrolytes, additional 2 ml of blood sample was taken only from each rat of those treated with aluminum alone (Al), with aluminum and plant (AlS), with plant alone (S) and from the control group (C). Each sample was then placed in a plane tube, left to coagulate, centrifuged at 1000 rpm and the supernatant (serum) was employed in measuring Na, K, Ca, P and Fe. For Na and K, aliquot for each element was placed in a cuvette and measured using flame photometer apparatus, and the same carried out for measuring Ca, P and Fe using Spectrophotometers apparatus of the type (30122). Both instruments were adjusted to zero by blanks brought with the kit for each element.
Student t- test was used for paired comparison of mean values (mean ± SD) between the control and the experimental groups in both rats and rabbits, between the control (C) and Al- treated groups (Al) as well as between Al-treated and those provided with Al co-administered with the plant (AlS). The significance is taken at P < 0.05 and P < 0.01.
| Results|| |
The results of blood parameters in rats and rabbits are shown in [Table 1] and [Table 2], respectively. For the rats, the results showed only PCV and MCHC in group (G 3 ) and PCV in group (G 2 ) that had increased significantly (P < 0.05) [Figure 1], whereas for the rabbits, only Hb and MCV values had increased significantly (P < 0.05) [Figure 2].
|Figure 1: Mean cell hemoglobin concentration in control group (1) and three experimental groups of rats provided daily with 150 mg/kg (2), 300 mg/kg (3) and 600 mg/kg (4) respective doses of S. argel leaves extract daily for 30 days in captivity|
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|Figure 2: Mean cell volume in control group of rabbits (1) and experimental one (2) fed with leaves of S. argel (2.5 g/kg) mixed with their clover feed daily for 30 days in captivity|
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|Table 1: Mean ± SD values of blood parameters in control group of rats (CG) and three experimental groups (G1, G2 and G3) provided daily with the respective doses of S. argel leaves extract daily for 30 days in captivity|
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|Table 2: Mean ± SD values of blood parameters in control group (RG1) and experimental group of rabbit (RG2) fed with leaves of S. argel (2.5 g/kg) mixed with their clover feed daily for 30 days in captivity|
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Mean values of BWs of rats and rabbits are shown in [Table 3] and [Table 4], respectively. The results showed that there were no significant changes (P > 0.05) observed in the BWS of the two groups of animals.
|Table 3: Mean ± SD values of initial body weight (IBW) and final body weight (FBW) of control group of rats (CG) and three experimental groups (G1, G2 and G3) provided daily with the respective doses of S. argel leaves extract for 30 days in captivity|
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|Table 4: Mean ± SD values of initial body weight (IBW) and final body weight (FBW) of control rabbits (RG1) and experimental rabbits (RG2) fed with leaves of S. argel (2.5 g/kg) daily for 30 days in captivity|
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The results of blood parameters in group (Al), group (AlS), group (S) and group (C) are shown in [Table 5]. The results showed RBCs, Hb, PCV and MCHC of group (Al) and group (AlS) were each significantly low when compared to those in group (C) and group (S) whereas in these two groups each pair was not significantly different (P > 0.05) [Figure 3] and [Figure 4]. On the other hand, WBC count, PLT and Lym of group (Al) were not significantly affected (P > 0.05) when each compared to that in group (C), group (AlS) and group(S) and only MCHC and PCV of group (S) were significantly higher (P < 0.05) than those in the control group (C).
|Figure 3: Red blood cells in control group (1) and three experimental groups of rats; treated with AlCl3 0.5 mg/kg (2), with AlCl3 plus S. argel (3) and with S. argel 600 mg/kg alone (4) daily for 30 days|
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|Figure 4: Hemoglobin concentration in control group (1) and three experimental groups of rats; treated with AlCl3 0.5 mg/kg (2), with AlCl3 plus S. argel (3) and with S. argel 600 mg/kg alone (4) daily for 30 days|
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|Table 5: Mean ± SD values of blood parameters in control group© and three experimental groups; treated with AlCl3 0.5 mg/kg (Al), with AlCl3 plus S. argel (AlS) and with S. argel 600 mg/kg alone (S) daily for 30 days|
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The results showing the mean values of electrolytes; namely: Na, K, Ca, P and Fe are shown in [Table 6]. The results revealed the levels of K, Ca and P in group (S) to increase significantly (P < 0.05) when each compared to those in group (C). On the other hand, the level of K and P in group (AlS) and K, Ca and P of group (S) were found significantly higher (P < 0.01) when each compared to that in group (Al).
|Table 6: Mean ± SD values of blood electrolytes in the control group© and three experimental groups; treated with AlCl3 0.5 mg/kg (Al), with AlCl3 plus S. argel (AlS) and with S. argel alone 600 mg/kg (S)|
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For Fe concentration, it was significantly reduced (P < 0.01) in group (Al) and group (AlS) when each compared to its levels in group (C). Moreover, the level of Fe was found significantly higher (P < 0.01) in group (S) when compared to its levels in group (Al) but not differed from the control group.
| Discussion|| |
The blood parameters; RBCs count, Hb, PCV, MCHC, MCV together with the level of Fe are indicative indices of anemia and could be used to indicate nutritional values of ingested diets as well. , Therefore, occurrence of anemia is attributable to lowered values of such indices.
However, the observed increase in PCV and MCHC in tested rats of this study might be due to stimulating effect of the S. Argel on erythropoietin, a hormone leading to red blood production.  Moreover, the increase in PCV appeared to be dose- dependent and to some extent for MCHC.
In the meantime, the increase of Hb and MCV in rabbits following intake of S. argel, could be due to high nutritional values of this plant particularly in vitamins and minerals such as Fe and copper.  In this context, Fe is reported to be necessary for the formation of the heme part of the hemoglobin and copper for the absorption of Fe from the gastrointestinal tract. 
These results, probably were expected as phytochemical screening of leaves of this shrub had revealed the presence of plenty of biochemical nutritive components as well as minerals.  Unexpected to occur, however, was that such finding not to be reflected in marked increase in BWs of the experimental animals. However, insignificant changes in BWs were probably owing to the fact that the animals were only fed for a month and progressed in weight with slightly slow rate of increase. On the other hand, persistence of feeding for prolonged periods might lead to overwhelmed increase in the BWs of experimental animals.
Similar to its toxic effects reported elsewhere Martínez et al.,  Vota et al.,  Tomljenovic and Shaw  Newairy et al.,  Al administration by rats in the present study had also induced significant reductions in the blood indices of anemia. However, the significant increase in MCV in Al-treated group could be attributed to reduced level of RBCs count in the same group where MCV is usually calculated by dividing PCV by RBCs count. In this context, several mechanisms have been proposed for the Al-induced anemia, but the exact mechanism is unknown. The proposed mechanisms appear to involve reduction in heme biosynthesis and Fe metabolism in rats , moreover, the increased Al levels in serum of patients with chronic renal failure on hemodialysis were reported to associate with impaired erythropoiesis and Fe metabolism.  Other studies conducted by Farina et al.,  Vittori et al.,  Willhite et al.,  confirmed AlCl 3 involvement in reduction of RBCs synthesis in bone marrow.
In the present study, uptake of ethanolic extract of S. argel co-administered with AlCl 3 seems to mitigate Al-induced anemia and to raise the same values of blood indices of anemia but not to significant levels except for MCHC, MCV and PCV when compared with those of (Al) alone. As inferred from other reports, two mechanisms could be suggested for the prevention of Al toxicity by S. argel: First, the plant inhibited or reduced Al absorption from intestine and the second: Aluminum overload might modulate gastrointestinal Fe absorption and hinder the cellular uptake of Al.  Moreover, insignificant changes of WBCs, PLT and LYM prior and after treatment with Al toxicity could confirm further that the effect of Al was associated with Fe metabolism.
On the other hand, concurrent fluctuated levels of Fe with administration of AlCl 3 alone and with S. argel could furtherly confirm the claim of the mitigative effect of S. argel against anemia. Moreover, having other electrolytes unaffected by Al toxicity taken with their levels being improved by S. argel, alone or mixed with AlCl 3 would probably support the claim that the plant was effective as anti-inflammatory agent,  against adverse effect that might be caused by Al and hence enhanced absorption of these minerals via intestinal tract.
| References|| |
|1.||Innocenti G, Dall'Acqua S, Minesso P, Budriesi R, Micucci M, Chiarini A. Evaluation of muscarinic M3-receptor antagonism of Solenostemma argel leaves. Planta Med 2010;76:634. |
|2.||Perrone A, Plaza A, Hamed A, Pizza C, Piacente Solenostemma argel: A Rich source of very unusual pregnane and 14, 15-secopregnane glycosides with antiproliferative activity . Curr Org Chem 2008;12:1648-60. |
|3.||Tigani SE, Ahmed SS. Solenostemma argel tissue culture for production of secondary metabolites. J Gene Engineering Biotechnol 2009;19:23. |
|4.||ElKamali HH, Khalid SA. The most common herbal remedies in central Sudan. Fitoterapia 1996;4:301. |
|5.||Mohamed EZ, Amani SA, Mounerah R, Reham M. Antimicrobial activities of Saudi Arabian desert plants. Phytopharmacology 2012;2:106-13. |
|6.||Awad KT, Khalid OA, Tagelsir IM, Sidahmed O. Argel (Solennostemma argel Del. Hayenne) applications for control of the date palm green scale insect (Asterolicanium phoenicis Rao) and yield enhancement. ARPN J Agricult Biological Sci 2012;7:380-5. |
|7.||Feiha MH, Awad KT, Hatim GM, Omar AA. Water extracts of Hargal plant (Solenostemma argel, Del Hyne) and Usher (Calotropis procera Ail) leaves as natural insecticides against mosquito larvae. J Sci Tech 2009;10:67-76. |
|8.||Stangeland T, Alele PE, Katuura E, Lye KA. Plants used to treat malaria in Nyakayojo sub-county, western Uganda. J Ethnopharmacol 2011;137:154-66. |
|9.||AbouZid SF, Mohamed AA. Survey on medicinal plants and spices used in Beni-Sueif, Upper Egypt. J Ethnobiol Ethnomed 2011;7:18. |
|10.||Shafek RE, Michael HN. Antibacterial and antioxidant activity of two new kaempferol glycosides isolated from Solenostemma argel stem extract. Asian J plant Sci 2012;11:143-7. |
|11.||Mahalel UA. Antibacterial sensitivity for some chemically diverse steroidal glycosides in vitro. J Agric Soc Sci 2012;8:24-8. |
|12.||Plaza A, Perrone A, Balestrieri ML, Felice F, Balestrieri C, Hamed AI, et al. New unusual pregnane glycosides with antiproliferative activity from Solenostemma argel. Steroids 2005;70:594-603. |
|13.||Murwan KS, Murwa AM. Chemical composition, minerals, protein fractionation, and anti-nutrition factors in leaf of Hargel plant (Solenostemma argel). Eur J Sci Res 2010;43:430-4. |
|14.||Campbell A. The potential role of aluminium in Alzheimer's disease. Nephrol Dial Transplant 2002;17 Suppl 2:17-20. |
|15.||Geyikoglu F, Turkez H, Bakir TO, Cicek M. The genotoxic, hepatotoxic, nephrotoxic, haematotoxic and histopathological effects in rats after aluminium chronic intoxication. Toxicol Ind Health 2012;15. [Epub ahead of print]. |
|16.||Cannata-Andía JB, Fernández-Martín JL. The clinical impact of aluminium overload in renal failure. Nephrol Dial Transplant 2002;17:9-12. |
|17.||Willhite CC, Ball GL, McLellan CJ. Total allowable concentrations of monomeric inorganic aluminum and hydrated aluminum silicates in drinking water. Crit Rev Toxicol 2012;42:358-442. |
|18.||Martínez Mdel P, Bozzini C, Olivera MI, Dmytrenko G, Conti MI. Aluminum bone toxicity in immature rats exposed to simulated high altitude. J Bone Miner Metab 2011;29:526-34. |
|19.||Vota DM, Crisp RL, Nesse AB, Vittori DC. Oxidative stress due to aluminum exposure induces eryptosis which is prevented by erythropoietin. J Cell Biochem 2012;113:1581-9. |
|20.||Lemire J, Mailloux R, Auger C, Whalen D, Appanna VD. Pseudomonas fluorescens orchestrates a fine metabolic-balancing act to counter aluminium toxicity. Environ Microbiol 2010;12:1384-90. |
|21.||Tomljenovic L, Shaw CA. Mechanisms of aluminum adjuvant toxicity and autoimmunity in pediatric populations. Lupus 2012;21:223-30. |
|22.||Yousef MI, Salama AF. Propolis protection from reproductive toxicity caused by aluminium chloride in male rats. Food Chem Toxicol 2009;47:1168-75. |
|23.||Fahid A. Camel's milk protects against aluminium chloride-induced normocytic normocromic anaemia, lipid peroxidation and oxidative stress in erythrocytes of white albino rats. Am J Biochem Biotechnol 2009;5:127-36. |
|24.||Weber DK, Danielson K, Wright S, Foley JE. Hematology and serum biochemistry values of dusky-footed wood rat (Neotoma fuscipes). J Wildl Dis 2002;38:576-82. |
|25.||Hewitt CD, Innes DJ, Savory J, Wills MR. Normal biochemical and hematological values in New Zealand white rabbits. Clin Chem 1989;35:1777-9. |
|26.||Bagchi K. Iron deficiency anaemia - An old enemy. East Mediterr Health J 2004;10:754-60. |
|27.||Janus J, Moerschel SK. Evaluation of anemia in children. Am Fam Physician 2010;81:1462-71. |
|28.||Debeljak N, Sytkowski AJ. Erythropoietin and erythropoiesis stimulating agents. Drug Test Anal 2012;4:805-12. |
|29.||Salah A, Njunda L. Effect of Ruellia praetermissa extract on erythropoiesis in pregnant women. Global J Res Med Plants Indigen Med 2012;8:309-14. |
|30.||Newairy AS, Salama AF, Hussien HM, Yousef MI. Propolis alleviates aluminium-induced lipid peroxidation and biochemical parameters in male rats. Food Chem Toxicol 2009;47:1093-8. |
|31.||Ganchev T, Dyankov E, Zacharieva R, Pachalieva I, Velikova M, Kavaldjieva B. Influence of aluminium on erythropoiesis, iron metabolism and some functional characteristics of erythrocytes in rats. Acta Physiol Pharmacol Bulg 1998;23:27-31. |
|32.||Han J, Han J, Dunn MA. Effect of dietary aluminum on tissue nonheme iron and ferritin levels in the chick. Toxicology 2000;142:97-109. |
|33.||Farina M, Rotta LN, Soares FA, Jardim F, Jacques R, Souza DO, et al. Hematological changes in rats chronically exposed to oral aluminum. Toxicology 2005;209:29-37. |
|34.||Vittori D, Nesse A, Pérez G, Garbossa G. Morphologic and functional alterations of erythroid cells induced by long-term ingestion of aluminium. J Inorg Biochem 1999;76:113-20. |
|35.||Cannata JB, Gómez Alonso C, Fernández Menéndez MJ, Fernández Soto I, McGregor S, Menéndez-Fraga P, et al. Iron uptake in aluminium overload: In vivo and in vitro studies. Nephrol Dial Transplant 1991;6:637-42. |
[Figure 1], [Figure 2], [Figure 3], [Figure 4]
[Table 1], [Table 2], [Table 3], [Table 4], [Table 5], [Table 6]