|Year : 2019 | Volume
| Issue : 3 | Page : 173-177
Acute toxicity study of Cholewin Plus tablet: A herbal formulation in rodent
Manish P Deshmukh1, Rangnath R Chikane1, Shyam Bhutada2, Anil Anjankar1
1 Central Preclinical Research Facility, Datta Meghe Institute of Medical Sciences (Deemed University) (DMIMS), Wardha, Maharashtra, India
2 Department of Samhita Siddhanta, Mahatma Gandhi Ayurved College, Hospital and Research Centre, Salod(H), Wardha, Maharashtra, India
|Date of Submission||20-Dec-2019|
|Date of Decision||06-Jan-2020|
|Date of Acceptance||13-Jan-2020|
|Date of Web Publication||12-Feb-2020|
Mr. Manish P Deshmukh
Central Preclinical Research Facility, Datta Meghe Institute of Medical Sciences (Deemed University) (DMIMS), Wardha, Maharashtra
Source of Support: None, Conflict of Interest: None
Background: Cholewin Plus tablets are used for the treatment of hyperlipidemia and coronary artery disease. Aim: The aim of this study was to determine acute oral toxicity of Cholewin Plus tablet in vivo in Wistar rats. Materials and Methods Rats were orally administrated single dose of 5, 300, and 2000 mg/kg Cholewin Plus tablet and the controlled group received vehicle only orally. There were five rats in each group. All animals were killed after 14 days of treatment and histopathology was performed. A total of four parameters were observed throughout the study: cage side observation, the effect on body weight, the effect on hematological parameter, and histopathology. Result: No mortality and morbidity were noted after 14 days of treatment. In general, behavior, food and water consumption, hematological studies, and histopathology showed no significant changes. Conclusion: The oral administration of Cholewin Plus tablet did not show any toxic effect on the animal at the dose of 2000 mg/kg body weight. Therefore, it is a safe remedy for human use.
Keywords: Acute toxicity, Cholewin Plus tablet, single dose
|How to cite this article:|
Deshmukh MP, Chikane RR, Bhutada S, Anjankar A. Acute toxicity study of Cholewin Plus tablet: A herbal formulation in rodent. J Indian Sys Medicine 2019;7:173-7
|How to cite this URL:|
Deshmukh MP, Chikane RR, Bhutada S, Anjankar A. Acute toxicity study of Cholewin Plus tablet: A herbal formulation in rodent. J Indian Sys Medicine [serial online] 2019 [cited 2020 Apr 4];7:173-7. Available from: http://www.joinsysmed.com/text.asp?2019/7/3/173/278141
| Introduction|| |
Herbal formulations, which have attained widespread acceptability as therapeutic agents in India, include lipid-lowering, antidiabetic, hepatoprotective, nootropic, and immunomodulatory agents.,, Cholewin Plus tablet is a polyherbal lipid-lowering formulation containing 11 ingredients of herbal origin. It is the best polyherbal medicine for cholesterol formulated using medicinal plants as recommended by Ayurveda. It is designed to maintain cardiac function as well as to promote hypolipidemia to optimize cardiac health. It works to protect the heart and to strengthen the heart muscles. It regulates cholesterol and triglyceride levels and is also helpful to stop coronary artery blockages. The key ingredients and their roles in the tablet are as follows.
Arjun: Arjun (Terminalia arjuna Rob) is a cardioprotective herb. It prevents oxidative damage to the heart and stabilizes blood pressure.
Guggul and Katuki: Research shows that Guggul (Balsamodendron mukul Hook. ex Stocks) and Katuki (Picrorhiza kurroa Royle ex Benth.) prevent liver from releasing excessive amounts of low-density lipoprotein (LDL) cholesterol into the bloodstream.
Kachnar, Methi, and Taj: Kachnar (Bauhinia variegata L. Benth), Methi (Trigonella foenum-graecum L.), and Taj (Cinnamomum cassia (L.) J. Presi) are useful to maintain healthy blood flow and quality, and they also help in keeping healthy cholesterol levels.,,
Lashuna: Lashuna (Allium sativum L.) has fibrinolytic, antithrombotic, and anti-hyperlipidemic actions.
Ingredients present in Cholewin Plus Tablet (each film-coated tablet contains)
In this study, the acute toxicity of Cholewin Plus tablet, a polyherbal formulation, was investigated to assess its safety and tolerability profile in short-term treatment.
| Materials and Methods|| |
The supplier provided the test container containing a medicinal herb identified as Cholewin Plus tablet. The herbal remedy, Cholewin Plus tablet, was manufactured by Gayatri Ayupharma Pvt. Ltd., 234/2/2, Patel Estate, Nikol Road, Ahmedabad, Gujarat, India. The test substance was stored according to the manufacturer’s storage condition in a cool and dark place.
Adult healthy Wistar albino rats, weighing between 130 and 170g, were obtained from the animal house of the Central Preclinical Research Facility, Datta Meghe Institute of Medical Sciences, Wardha, Maharashtra, India. Nulliparous nonpregnant female rats were selected for acute toxicity study. The animals were housed in 37cm × 23cm × 16cm polypropylene cages with five rats per cage and were maintained at a constant temperature on a standard 12:12 light/dark cycle. Food and water were provided ad libitum. They were allowed to acclimatize for 7 days to the laboratory conditions before the experiment., The experiment was conducted according to the ethical norms approved by Institutional Animal Ethics Committee (Registration No. 571/02/a/CPCSEA), Central Preclinical Research Facility, Datta Meghe Institute of Medical Sciences.
Acute Toxicity Study
Female Wistar albino rats, weighing between 130 and 170g, maintained under standard laboratory conditions were used for the acute toxicity test according to the Organisation for Economic Co-operation and Development (OECD) Guidelines 420. A total of 20 female rats were selected for the study and were divided into four groups having five animals in each group. Animals were kept overnight fasting prior to drug administration by oral gavage. After administration of drug, sample food was provided after 3–4h. All animals were observed individually at least once for the first 30min after dosing and thereafter every 1h for the next 4h. Observation was done daily for the next 14 days every 24h. Daily observation on the changes in skin and fur, eyes and mucous membrane, respiratory rates, autonomic effects such as salivation, lacrimation, perspiration, piloerection, urinary incontinence and defection, and the effects on the central nervous system such as drowsiness, tremors, and convulsions were noted.,,
Preparation of test substance
The contents of Cholewin Plus film-coated tablet were insoluble in water. Suspension of the test substance was prepared by triturating the tablet and was suspended in 1% gum acacia solution as a suspending agent and 0.1% Tween 80 as a wetting agent. The suspension was then stored at 4°C for up to 24h. All other chemicals and solvents used in this study were of analytical grade.
Dosage of test substance on animals
Single-dose toxicity study was performed according to OECD Guidelines 420. In order to study any possible toxic effect, four groups of rats (all female) were used in this experiment. The acute toxicity of Cholewin Plus tablet was studied at different dose concentrations. Group I was administered a mixture of 1% gum acacia and 0.1% Tween 80. Groups II, III, and IV were administered test drug suspension at a dose of 5, 300, and 2000 mg/kg, respectively. The maximum volume of gum acacia suspension was not more than 2mL/100g body weight. Animals fasted prior to conducting the experiment only of food but not of water, which was withheld overnight. Following 24h of fasting, the animals were weighed and the test substance was administered. After the test substance was administered, food was withheld for further 2–4h.
Cage Side Observation and Body Weight Measurement
Animals were observed individually after drug administration and special attention was given during the first 4h and every 12h daily thereafter, for a total of 14 days. All observations were systematically recorded, with individual records being maintained for each animal. Cage side observation included evaluation of skin and fur, eyes, respiratory effects; autonomic effects such as salivation, diarrhea, and urination; and the central nerve effects including tremors and convulsions; changes in the level of activity; gait and posture; reactivity to handling or sensory stimuli; and altered strength., Individual body weight was measured and recorded prior to the administration of the test drug. After 2 weeks, the change in body weight of animals was calculated and recorded.
Food and Water Consumption
The amount of food and water consumed was measured daily from the quantity of food and water supplied and the amount remaining after 24h for 2 weeks of the study period.,
The change in body weight was recorded on day 1 before dosing as well as on day 8 and day 15 after dosing that is weekly basis.,,
During the study period, all animals were observed daily for any mortality. If any dead animal was found, histopathology was performed of that animal and it was observed for any abnormal sign on measure organs such as kidney, liver, and heart. After completion of the study, all animals were killed using anesthesia, and histopathology was performed and the results were recorded.,
| Results|| |
Acute Toxicity Study
No mortality and morbidity or any toxicity or any sign of behavioral changes were observed throughout the 14-day period after single oral administration of Cholewin Plus tablet suspension up to the dose of 2000 mg/kg body weight. The morphological characteristics such as fur, skin, eyes, and nose appeared normal. During observation no tremors, convulsion, salivation, diarrhea, lethargy, or unusual behaviors such as self-mutilation and walking backward were observed; gait and posture, grip strength, and sensory stimuli all were found to be normal. Observations were recorded and were reported in [Table 1].
|Table 1: Acute study with dose of 2000 mg/kg/day on female rats for 14 days|
Click here to view
Effect on Body Weight
The weight variation between test and treatment groups was noted. A slight increase in the weight was noted on day 1 and day 15.
The change in body weight was recorded and reported in [Table 2] and [Chart 1].
|Table 2: Change in body weight at day 1, 8, and 15 at a dose of 2000 mg/kg|
Click here to view
No abnormal clinical sign was observed throughout the acute toxicity study.
Effect on Hematological Parameter
After compilation of acute toxicity study, blood samples were collected in heparinized vials. These samples were provided for determination hemoglobin, total red blood cell (RBC) and white blood cell (WBC) count, platelets, and percentage of reticulocytes and hematocrit count. The change in hematological parameter was recorded and noted in [Table 3]. No significant changes were observed in blood cell count.
No abnormalities were detected in pathological examinations of the tissues during microscopic examination of internal organs in comparative histology of tissues of control and test animals. Cholewin Plus tablet does not affect the histology of vital organs, namely, lungs, heart, spleen, liver, and kidneys.
| Discussion|| |
Nowadays a number of polyherbal-based formulations are used abundantly worldwide compared to allopathic medicines for the treatment of different types of ailments. In India, different types of Ayurveda-based herbal formulations are used for the treatment of lipid disorder as well as to maintain cardiac function. The test formulation was the mixture of this herbal plant formulated in the form of a tablet. These tablets help maintain healthy cholesterol level, increase good cholesterol level, and prevent coronary artery diseases., In this study, acute toxicity of herbal product was analyzed in female Wistar rats as a single-dose study. The acute toxicity study was conducted for 14 days at the dose of 5, 300, and 2000 mg/kg/day administered daily in all groups. The results showed that there was no morbidity and mortality. This proves that the drug could be safely administered for acute treatments up to a maximum dose level of 2000 mg/kg/day for the oral route. The change in body weight has been used as an indicator of the adverse effect of test formulation. During the experimental period, no remarkable changes were observed in animal behavior, body weight, and organ weight at all dose levels in treated rats as compared to a controlled group. It can be inferred that test substance is nontoxic at the dose administered. The analysis of animals’ blood parameters can be used for risk evaluation in humans as a change in the hematological system has a higher predictive value for human toxicity of test compound. In this study, all hematological parameters that were observed showed normal value for the animal blood sample. So we can predict that this herbal formulation is safe for humans also.
| Conclusion|| |
The result strongly supports that the herbal Cholewin Plus tablet formulation is safe and well tolerated at the tested oral doses of 5, 300, and 2000 mg/kg/day for 14 days as no deleterious changes were observed in animal macroparameters, behavior and habits, hematology, and histopathological parameters.
Financial support and sponsorship
Conflicts of interest
There are no conflicts of interest.
| References|| |
Benzie IFF, Wachtel-Galor S. Herbal Medicine: Biomolecular and Clinical Aspects. Boca Raton, FL: CRC Press; 2011.
Ishtiaq S, Akram M, Kamran SH, Hanif U, Afridi MSK, Sajid-Ur-Rehman, et al
. Acute and sub-acute toxicity study of a Pakistani polyherbal formulation. BMC Complement Altern Med 2017;17:387.
Barnes J, Anderson LA, Phillipson JD, editors. Herbal Medicines: A Guide for Healthcare Professionals. London: Pharmaceutical Press; 2002.
Jain S, Yadav PP, Gill V, Vasudeva N, Singla N. Terminalia arjuna
a sacred medicinal plant: Phytochemical and pharmacological profile. Phytochem Rev 2009;8:491-502.
Elmahdi B, El-Bahr SM, Abdelghany AM. Effect of dietary supplementation of fenugreek (Trigonella foenum-graecum L.
) on selected biochemical parameters of rats fed high cholesterol diet: Profiles of lipid and lipoproteins. Sylwan 2014;158:399-420.
Tomar R, Kaur G, Sannd R, Singh H, Sarkar B. A review on guggulu formulations used in Ayurveda. Annals Ayurvedic Med 2014;3:96-113.
Sagia A, Khan MI, Eva EO, Mahbub MI. Comparison of lipid lowering effect of aqueous extract of cinnamon (Cinnamomum cassiae
) with that of rosuvastatin on experimentally induced hypercholesterolaemic rats. TAJ 2018;31:52-61.
OECD. Available from: http://www.oecd.org/document/22/0,2340,en_2649_34377_1916054_1_1_1_1,00.htm. [Last accessed on 2019 Sept 15].
British Toxicology Society Working Party on Toxicity. Special report: A new approach to the classification of substances and preparations on the basis of their acute toxicity. Human Toxicol 1984;3:85-92.
OECD. Guidance Document on the Recognition, Assessment and Use of Clinical Signs as Humane Endpoints for Experimental Animals Used in Safety Evaluation. Environmental Health and Safety Monograph Series on Testing and Assessment No. 19. Available from: http://www.oecd.org/document/22/0,2340,en_2649_34377_1916054_1_1_1_1,00.html. [Last accessed on 2019 Sept 15].
Londhe VP. Role of garlic (Allium sativum
) in various diseases: An overview. Angiogenesis 2011;12:13.
Thackaberry EA. Vehicle selection for nonclinical oral safety studies. Expert Opin Drug Metab Toxicol 2013;9:1635-46.
Harizal SN, Mansor SM, Hasnan J, Tharakan JK, Abdullah J. Acute toxicity study of the standardized methanolic extract of Mitragyna speciosa Korth in rodent. J Ethnopharmacol 2010;131:404-9.
Kumarnsit E, Keawpradub N, Nuankaew W. Acute and long-term effects of alkaloid extract of Mitragyna speciosa on food and water intake and body weight in rats. Fitoterapia 2006;77:339-45.
Ali R, Ali R, Jaimini A, Nishad DK, Mittal G, Chaurasia OP, et al
. Acute and sub acute toxicity and efficacy studies of hippophae rhamnoides based herbal antioxidant supplement. Indian J Pharmacol 2012;44:504-8.
] [Full text]
Teo S, Stirling D, Thomas S, Hoberman A, Kiorpes A, Khetani V. A 90-day oral gavage toxicity study of D-methylphenidate and D,L-methylphenidate in Sprague-Dawley rats. Toxicology 2002;179:183-96.
Patrick-Iwuanyanwu KC, Amadi U, Ayalogu EO, Charles IA. Evaluation of acute and sub-chronic oral toxicity study of baker cleansers bitters: A polyherbal drug on experimental rats. Excli J 2012;11:632-40.
Kuruvilla A. Herbal formulations as pharmacotherapeutic agents. Indian J Exp Biol 2002;40:7-11.
Karthikeyan K, Bai BR, Gauthaman K, Sathish KS, Devaraj SN. Cardioprotective effect of the alcoholic extract of Terminalia arjuna
bark in an in vivo model of myocardial ischemic reperfusion injury. Life Sci 2003;73:2727-39.
Chander R, Singh K, Khanna AK, Kaul SM, Puri A, Saxena R, et al
. Antidyslipidemic and antioxidant activities of different fractions of Terminalia arjuna
stem bark. Indian J Clin Biochem 2004;19:141-8.
Joshi CS, Priya ES, Venkataraman S. Acute and subacute toxicity studies on the polyherbal antidiabetic formulation diakyur in experimental animal models. J Health Sci 2007;53:245-9.
[Table 1], [Table 2], [Table 3], [Table 4]