|Year : 2017 | Volume
| Issue : 3 | Page : 108-113
Sub-chronic safety evaluation of aqueous extract of Alangium salvifolium (L.f.) Wangerin leaves in rats
Bhupinder Kapoor, Gagandeep Kaur, Mukta Gupta, Reena Gupta
Department of Pharmacology, School of Pharmaceutical Sciences, Lovely Professional University, Phagwara, Punjab, India
|Date of Web Publication||10-Jul-2017|
School of Pharmaceutical Sciences, Lovely Professional University, Phagwara, Punjab
Source of Support: None, Conflict of Interest: None
Conventionally, the juice and extract of Alangium salvifolium leaves have been used for the treatment of diabetes, wound healing, dog bite, and as a poultice in rheumatism. To carry out the sub-chronic toxicity and thereafter safety evaluation of A. salvifolium leaves. The aqueous extract of A. salvifolium leaves was administered orally at the doses of 200, 400, and 800 mg/kg/day for 90 days. All the animals were observed daily for general behavior, changes in body weight, food, and water consumption. At the end of the treatment period, biochemical and hematological parameters were analyzed; and the animals were sacrificed for histopathological examination of heart, lungs, liver, and kidney. The general behavior and water intake were normal in all the rats. The increase in body weight was observed in female rats of all the groups while body weight was decreased in high dose group animals of both sexes. Hematological parameters were not disturbed by the continuous use of extract. A significant decrease in glucose level was observed in intermediate- and high-dose group animals while urea and creatinine level were significantly high in animals of high-dose group. Although histopathological examination of most of the organs exhibited no structural changes, some tubal damage in kidneys was observed in high-dose group animals. The high dose of extract has shown mild signs of toxicity on kidney function test, but no toxic response was observed on hematological and liver biochemical parameters. The extract also exhibited hypoglycemic potential.
Keywords: Alangium salvifolium, phytoconstituents, sub-chronic toxicity
|How to cite this article:|
Kapoor B, Kaur G, Gupta M, Gupta R. Sub-chronic safety evaluation of aqueous extract of Alangium salvifolium (L.f.) Wangerin leaves in rats. J Adv Pharm Technol Res 2017;8:108-13
|How to cite this URL:|
Kapoor B, Kaur G, Gupta M, Gupta R. Sub-chronic safety evaluation of aqueous extract of Alangium salvifolium (L.f.) Wangerin leaves in rats. J Adv Pharm Technol Res [serial online] 2017 [cited 2022 May 23];8:108-13. Available from: https://www.japtr.org/text.asp?2017/8/3/108/210027
| Introduction|| |
The use of medicinal plants, either as an extract pure compound or as a derivative, is being used for various therapeutic purposes. The World Health Organisation in 2008 reported that 80% of Asian population uses herbal medicinal products for their primary health care and the data are same for developed countries also. As a result of inadequate knowledge regarding mode of action, potential side effects, interactions with other pharmaceuticals; the indiscriminate use of herbal products can result is serious toxic effects., Some extremely toxic phytoconstituents such as digoxin and strychnine have been extracted for various therapeutic purposes, hence careful investigation is required before use of such medicinal plants. Since safety continues to be a major issue with the use of herbal medicines, it is important that the relevant regulatory measures are to be taken to protect public health by ensuring that all herbal medicines are safe and of suitable quality; and have passed the standard quality and toxicity studies.
Alangium salvifolium (Syn. Alangium lamarckii; Family: Alangiaceae) is a deciduous, thorny, armed or unarmed plant which is cultivated extensively in India due to its various therapeutic uses.,,, This plant is commonly known as Ankol or Ankola in Hindi and Ankolam, Aranji in Tamil and Malayalam The widespread uses of this plant are due to various phytoconstituents such as flavanoids, terpenoids, phenolic glycosides, tannins, steroids, β-carboline, and quinolizidine alkaloids such as alangine, alangicin, marckindine, tubulosine deoxytubulosine, emetine, ankorine, and alangimarckine.,, Leaf juice is applied externally and taken internally in case of dog bite. Leaves are also used as a poultice in rheumatic pains, wound healing, diabetes, and hemorrhages.
Other reported therapeutic uses of the plant include antiulcer, antidiarrheal, hepatoprotective, anticancer, analgesic and anti-inflammatory. Despite potential therapeutic uses of the plant, no data is available on safety or toxicity; hence, the present sub-chronic toxicity study was designed to evaluate the safety profile of aqueous extract of leaves.
| Materials and Methods|| |
Plant material, extraction, and preliminary phytochemical screening
A. salvifolium leaves were procured from the local market of Lucknow and was identified by Dr. S. Rajan, Department of Botany, Government College of Ooty, Tamil Nadu (specimen voucher no. LSPS/04/H/001/2013). Leaves were shade dried, crushed, and extracted by continuous hot extraction using water as solvent. After filtration, the extract was concentrated by rotary evaporator and stored in a desiccator. Qualitative analysis of the extract was carried out to identify the various phytoconstituents such as alkaloids, phenols, flavonoids, tannins, saponins, and glycosides.
The present study was carried out after the approval from Institutional Animal Ethical Committee and Sprague-Dawley rats were procured from National Institute of Pharmaceutical Education and Research, Mohali. The protocol approval number of the study was 954/PO/ac/06/CPCSEA/Nov-2012/I. The study was performed as per guidelines of Organization for Economic Co-operation and Development (408) for oral sub-chronic toxicity studies in rodents. The animals were acclimatized to environmental conditions for 7 days before the start of the study.
Sub-chronic toxicity study
The rats were randomly divided into four groups: control group, low-, medium-, and high-dose group (Group I–IV respectively); and each group contained 16 animals (8 males and 8 females). Rats were administered with aqueous extract by oral gavage once daily for 90 consecutive days. The control group received 2 ml/100 g of distilled water while Group II, III, and IV were administered with dose of 200, 400, and 800 mg/kg/day, respectively.
During the treatment period, all animals were observed once daily for the changes in eyes, skin, fur, occurrence of secretions, food intake, water intake, and mortality. At the end of the study, the blood was collected from the anesthetized animals by retro-orbital puncture for biochemical and hematological analysis. The animals were euthanized by cervical dislocation, and histopathological examination of kidneys, lungs, heart, and liver was carried out.
Hematological and biochemical analysis
Complete blood count and biochemical analysis were performed after the completion of the study. Lipid profile, liver function test, and kidney function test were carried out to evaluate the effect of the extract on important organ systems.
At the end of the study, the animals were sacrificed by cervical dislocation and subjected to gross and histopathological alterations. In the gross examination, changes in organ size, shape, and any visible lesions were recorded.
Results are shown as mean ± standard deviation. The statistical significance of the differences among the treatments and control was verified using one-way analysis of variance. P < 0.05 was considered statistically significant.
| Results|| |
During the 90-day observation period, all the groups (control as well as treatment) did not show any sign of morbidity or mortality. No unusual behavior or physical changes were observed in animals.
Animals body weight changes and food consumption
A marked reduction in the body weight of male animals was observed in intermediate and high dose groups from 9th week onward, but the data were not statistically significant. In contrast, significant increase in the body weight was observed in female rats in all dose groups with respect to control group. Changes in body weight also supported the less food consumption in male animals only in intermediate and high dose group from 10th week onward [Figure 1], [Figure 2], [Figure 3], [Figure 4].
|Figure 1: Effect of aqueous extract of Alangium salvifolium leaves on body weight of male rats (g/rat)|
Click here to view
|Figure 2: Effect of aqueous extract of Alangium salvifolium leaves on body weight of female rats (g/rat)|
Click here to view
|Figure 3: Effect of aqueous extract of Alangium salvifolium leaves on food consumption of male rats|
Click here to view
|Figure 4: Effect of aqueous extract of Alangium salvifolium leaves on food consumption of female rats|
Click here to view
Treatment group animals showed no significant changes in the hemoglobin and red blood cells level in both male and female animals in comparison to control group animals [Table 1].
|Table 1: Effect of aqueous extract of Alangium salvifolium leaves on hematological parameters|
Click here to view
Blood glucose levels were significantly decreased in both male and female rats of intermediate-dose group (P< 0.05) and high-dose group (P< 0.01). Significant reduction in the cholesterol and triglyceride levels were reported at the dose of 400 mg/kg and 800 mg/kg (at P < 0.01) [Table 2]. Serum glutamic oxaloacetic transaminase (SGOT) level was significantly reduced at the dose of 200 mg/kg (P< 0.05) and 800 mg/kg (P< 0.05) in male rats. No significant change was observed in the serum glutamate pyruvate transaminase (SGPT) level of both control and treated rats. Total serum protein test measures the total amount of protein including globulin and albumin in the blood. Total serum protein levels were significantly increased at the dose of 400 mg/kg (P< 0.05) and 800 mg/kg (P< 0.01) in female rats but no significant change was observed in male rats. All the treatment groups exhibited similar level of alkaline phosphatase as compared to control group [Table 3]. Serum urea and creatinine level were significantly increased in both male and female rats of high-dose group (P< 0.05) in comparison to control group animals. No significant change was observed in the uric acid level in all groups [Table 4].
|Table 2: Effect of aqueous extract of Alangium salvifolium on glucose and lipid profile|
Click here to view
|Table 3: Effect of aqueous extract of Alangium salvifolium on liver function test|
Click here to view
|Table 4: Effect of aqueous extract of Alangium salvifolium on kidney function test|
Click here to view
All the animals were subjected to necropsy after the completion of 13-week study period. For histopathological examination, the heart, kidneys, liver, and lungs were isolated and examined [Figure 5], [Figure 6], [Figure 7], [Figure 8].
|Figure 5: (a-b) Effect of aqueous extract of Alangium salvifolium leaves on microscopy of heart|
Click here to view
|Figure 6: (a-b) Effect of aqueous extract of Alangium salvifolium leaves on microscopy of lungs|
Click here to view
|Figure 7: (a-b) Effect of aqueous extract of Alangium salvifolium leaves on microscopy of kidneys|
Click here to view
|Figure 8: (a-b) Effect of aqueous extract of Alangium salvifolium leaves on microscopy of liver|
Click here to view
| Discussion|| |
Plants and plant-derived products have been used for prevention and control of diseases from ancient times. Medicines, functional foods, and nutritional supplements are increasingly being used worldwide. These medicines are perceived as to be safe or free from toxic effects, but a number of adverse effects are reported from these agents and scientific validation of herbal products is essential.
In present investigation, the aqueous extract of A. salvifolium leaves was evaluated for sub-chronic toxicity study after oral administration. Conventionally, this plant is used for various therapeutic purposes due to a number of bioactive phytoconstituents such as alkaloids, flavonoids, steroids, and glycosides. The reported therapeutic actions of the plant include antidiabetic,, anticancer, antioxidant, larvicidal, pesticidal, and skin disorders.,
After oral administration of aqueous extract for 13 weeks, none of the treatment group had shown any toxic effect. The first symptom observed is the change in body weight of animals; as it is a sensitive indicator for toxic responses to drugs, to hormonal variations, liver disorders, and decreased absorption of essential nutrients,, In contrast to male animals, significant increase in body weight of female animals was seen during the study [Figure 1] and [Figure 2].
Hematopoietic system is a sensitive target for toxic compounds and is an important index of physiological and pathological conditions in man and animals. No significant change suggested the nontoxic nature of extract to the hematopoietic system.
High dose group animals showed a significant decrease in blood glucose level in comparison to control and other treatment groups. It may be attributed to the hypoglycemic activity of the extract. This reduction in blood glucose level can be correlated with the decreased level of triglyceride in intermediate and high dose groups.
Significant reduction in the blood cholesterol and triglyceride levels at the dose of 400 mg/kg and 800 mg/kg suggested the hypolipidemic activity of the extract which indicates potential benefits of extract against cardiovascular diseases. Serum cholesterol and proteins are mainly regulated by liver and any change in serum concentrations of these parameters suggest some alterations in liver functions. SGOT level was significantly reduced at the dose of 200 mg/kg (<0.05) and 800 mg/kg (<0.01) in male rats. Both control and treated rats exhibited comparable SGPT level. Total serum protein levels (albumin and globulin) were significantly increased at the dose of 400 mg/kg (<0.05) and 800 mg/kg (<0.01) in female rats, but no significant change was observed in male rats. The elevated total serum protein level indicates dehydration, high protein and high-calorie diet, multiple myeloma, chronic liver disease, and chronic infections. To see the effect of extract on cellular damage, alkaline phosphatase level was measured and showed no significant change at all dose levels in both male and female animals as compared to control.
Kidney function test was performed to see the effect of the extract on urea, uric acid, and creatinine levels as these are considered as important markers for kidney dysfunction. Serum urea and creatinine levels were increased significantly in both male and female animals of high dose group in comparison to control group. Serum uric levels were not altered in any of the treatment group animals.
No remarkable structural changes were observed on histopathological examination of heart, lungs, and liver [Figure 5], [Figure 6] and [Figure 8]. Minimal tubal damage in kidneys was observed in high dose group of both sexes [Figure 7] which can be also correlated with significant changes in kidney function tests. It can be concluded that the plant should be taken cautiously in case of patients with kidney dysfunction.
| Conclusion|| |
It is concluded that the aqueous extract of A. salvifolium did not induce any toxicological effects on the majority of the parameters. The extract showed hypoglycemic activity and, therefore, can be of therapeutic value in the management of diabetes. The hypoglycemic effect of the plant can be correlated to the reduced triglyceride and cholesterol levels as they may decrease insulin resistance. Significant alterations in kidney function test parameters were observed which was further supported by histopathological analysis. All these findings suggested antidiabetic and hypolipidemic potential of the plant up to a dose of 400 mg/kg, but it has to be further supported by chronic toxicity study.
Financial support and sponsorship
Conflicts of interest
There are no conflicts of interest.
| References|| |
Araújo MC, Barcellos NM, Vieira PM, Gouveia TM, Guerra MO, Peters VM, et al.
Acute and sub chronic toxicity study of aqueous extract from the leaves and branches of Campomanesia velutina
(Cambess) O. Berg. J Ethnopharmacol 2017;201:17-25.
Chan TY. Aconitum alkaloid content and the high toxicity of aconite tincture. Forensic Sci Int 2012;222:1-3.
Reena G, Sanjiv D, Bhupinder K. Sub-chronic toxicity study of aqueous extract of Clerodendrum phlomidis
Leaves. Int J Drug Dev Res 2012;4:197-207.
Jordan SA, Cunningham DG, Marles RJ. Assessment of herbal medicinal products: Challenges, and opportunities to increase the knowledge base for safety assessment. Toxicol Appl Pharmacol 2010;243:198-216.
Kumar PP, Prabhakara MC, Satyavathi K, Kumar AS. Evaluation of cardiac activity of some traditionally used backyard Indian medicinal plants. Res J Pharm Biol Chem Sci 2010;1:641-4.
Kumar R, Pate DK, Prasad SK, Sairam K, Hemalatha S. Antidiabetic activity of alcoholic leaves extract of Alangium lamarckii
Thwaites on streptozotocin-nicotinamide induced type 2 diabetic rats. Asian Pac J Trop Med 2011;4:904-9.
Sharma AK, Agarwal V, Kumar R, Balasubramaniam A, Mishra A, Gupta R. Pharmacological studies on seeds of Alangium salvifolium
Linn. Acta Pol Pharm 2011;68:897-904.
Porchezhian E, Ansari SH, Ahmad S. Analgesic and anti-inflammatory effects of Alangium salvifolium
. Pharm Biol 2001;39:65-6.
Jain S, Sinha A, Bhakuni DS. The biosynthesis of β-carboline and quinolizidine alkaloids of Alangium lamarckii
. Phytochemistry 2002;60:853-9.
Karigar AA, Shariff WR, Sikarwar MS. Wound healing property of alcoholic extract of leaves of Alangium salvifolium
. J Pharm Res 2010;3:267-9.
Selin-Rani S, Senthil-Nathan S, Revathi K, Chandrasekaran R, Thanigaivel A, Vasantha-Srinivasan P, et al.
Toxicity of Alangium salvifolium
Wang chemical constituents against the tobacco cutworm Spodoptera litura
Fab. Pestic Biochem Physiol 2016;126:92-101.
Ahad HA, Yesupadam MR, Padmaja S, Swamy K. Phytochemical and anti-inflammatory evaluation of Alangium lamarckii
root extract. Pharm Sin 2011;2:119-26.
Mohanty P, Panda S, Mishra S, Panda P, Jaliwala Y, Milind P. Study of antiulcer activity of roots of Alangium salvifolium
Linn. in pylorus ligated rats. Int Res J Pharm 2011;2:190-2.
Zahan R, Alam MB, Islam MS, Sarker GC, Chowdhury NS, Hosain SB, et al
. Anticancer activity of Alangium salvifolium
flowers in Ehrlich ascites carcinoma bearing mice. Int J Curr Res 2011;7:254-62.
Chander TR, Reddy YN. Evaluation of hepatoprotective activity with leaf extract of Alangium salvifolium
Wang on CCl4
induced rats. Int J Pharm Technol 2014;5:6039-50.
Nahar L, Zahan R, Mosaddik A, Islam S, Haque A, Fazal A, et al
. Antioxidant and antitumor activity of chloroform extract of Alangium salvifolium
flowers. Phytopharmacology 2012;2:123-34.
Tran MH, Nguyen HD, Kim JC, Choi JS, Lee HK, Min BS. Phenolic glycosides from Alangium salviifolium
leaves with inhibitory activity on LPS-induced NO, PGE(2), and TNF-alpha production. Bioorg Med Chem Lett 2009;19:4389-93.
Zhang Z, Liu R, Pu X, Sun Y, Zhao X. Evaluation of the sub-chronic toxicity of a standardized flavonoid extract of safflower in rats. Regul Toxicol Pharmacol 2017;85:98-107.
Izzo AA, Di Carlo G, Borrelli F, Ernst E. Cardiovascular pharmacotherapy and herbal medicines: The risk of drug interaction. Int J Cardiol 2005;98:1-14.
Zahan R, Mosaddik MA, Barman RK, Wahed MI, Haque ME. Antibacterial and antidiarrhoeal activity of Alangium salvifolium
Wang flowers. Mol Clin Pharmacol 2012;2:34-43.
Prakash NU, Bhuvaneswar S, Prethy S, Rajalakshmi N, Saranya M, Ruth J, et al
. Studies on antimicrobial, antioxidant, larvicidal, pesticidal activity and phytochemistry of leaves of Alangium salvifolium
(Lf) wang. Int J Pharm Pharm Sci 2013;5:86-9.
Mukinda JT, Eagles PF. Acute and sub-chronic oral toxicity profiles of the aqueous extract of Polygala fruticosa
in female mice and rats. J Ethnopharmacol 2010;128:236-40.
Lee MY, Shin IS, Seo CS, Kim JH, Han SR, Shin HK. Subchronic oral toxicity studies of the traditional herbal formula Bangpungtongseong-san in Crl: CD (SD) rats. J Ethnopharmacol 2012;144:720-5.
Tan PV, Mezui C, Enow-Orock G, Njikam N, Dimo T, Bitolog P. Teratogenic effects, acute and sub chronic toxicity of the leaf aqueous extract of Ocimum suave
Wild (Lamiaceae) in rats. J Ethnopharmacol 2008;115:232-7.
Bitzur R, Cohen H, Kamari Y, Shaish A, Harats D. Triglycerides and HDL cholesterol: Stars or second leads in diabetes? Diabetes Care 2009;32 Suppl 2:S373-7.
Bidhe RM, Ghosh S. Acute and subchronic (28-day) oral toxicity study in rats fed with novel surfactants. AAPS J 2004;6:7-16.
Zaias J, Mineau M, Cray C, Yoon D, Altman NH. Reference values for serum proteins of common laboratory rodent strains. J Am Assoc Lab Anim Sci 2009;48:387-90.
Gnanamani A, Sudha M, Deepa G, Sudha M, Deivanai K, Sadulla S. Haematological and biochemical effects of polyphenolics in animal models. Chemosphere 2008;72:1321-6.
[Figure 1], [Figure 2], [Figure 3], [Figure 4], [Figure 5], [Figure 6], [Figure 7], [Figure 8]
[Table 1], [Table 2], [Table 3], [Table 4]