E-ISSN:2456-3110

Research Article

Holoptelea Integrifolia

Journal of Ayurveda and Integrated Medical Sciences

2024 Volume 9 Number 10 OCTOBER
Publisherwww.maharshicharaka.in

An experimental evaluation of bark of Chirbilva (Holoptelea integrifolia Planch.) for Hypercholesterolemia in Albino Rats

Katoch V1*, Kumar Jha P2, Kumar V3
DOI:10.21760/jaims.9.10.4

1* Vinod Katoch, Associate Professor, Dravyaguna, Abhilashi Ayurvedic Medical College and Research Institute, Mandi, Himachal Pradesh, India.

2 Prashant Kumar Jha, Professor, Quality Control Laboratory, ALN Rao Memorial Ayurvedic Medical College, Koppa, Karnataka, India.

3 Vikas Kumar, Associate Professor, Department of Samhita Siddhant, Abhilashi Ayurvedic Medical College and Research Institute, Mandi, Himachal Pradesh, India.

Hypercholesterolemia is an emerging problem affecting population now a days and responsible for CVD, HTN, Diabetes. Not following Dincharya, Ritucharya, smoking, consuming alcohol, lack of exercise, stress, intake of fast food are one of common cause for hypercholesterolemia. Hypercholesterolemia can be compared in Ayurveda as Medodusti which is due to intake of excessive Sleshmavradhaka Ahara (heavy food) leads to Agnimandhya (diminished digestive fire) and in turn Medodusti that is abnormal accumulation of Meda (Cholesterol). Chirbilva in Sushruta Samhita mentioned as Kaphamedovishoshaka. Hence to trace out its hypercholesteremic effect present study is taken.

Keywords: Hypercholesterolemia, Medodusti, Chirbilva, Holoptelea integrifolia

Corresponding Author How to Cite this Article To Browse
Vinod Katoch, Associate Professor, Dravyaguna, Abhilashi Ayurvedic Medical College and Research Institute, Mandi, Himachal Pradesh, India.
Email: 		Katoch V, Kumar Jha P, Kumar V, An experimental evaluation of bark of Chirbilva (Holoptelea integrifolia Planch.) for Hypercholesterolemia in Albino Rats. J Ayu Int Med Sci. 2024;9(10):26-32.
Available From
https://jaims.in/jaims/article/view/3703

Manuscript Received Review Round 1 Review Round 2 Review Round 3 Accepted
2024-09-11 2024-09-21 2024-10-01 2024-10-14 2024-10-24
Conflict of Interest Funding Ethical Approval Plagiarism X-checker Note
None Nil Yes 12.63

© 2024by Katoch V, Kumar Jha P, Kumar Vand Published by Maharshi Charaka Ayurveda Organization. This is an Open Access article licensed under a Creative Commons Attribution 4.0 International License https://creativecommons.org/licenses/by/4.0/ unported [CC BY 4.0].

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Vinod K et al. An experimental evaluation of bark of Chirbilva

Introduction

Global Health observatory data W.H.O risk of heart disease and stoke due to raised cholesterol levels. Globally ischemic heart disease is due to high cholesterol.[1]

Overall, raised cholesterol is estimated to cause 2.6 million deaths (4.5% of total) and 29.7 million disability adjusted life years (DALYS), or 2.0% of total DALYS. According to Journal of Clinical Lipidology April 2020 Rates of atherosclerotic cardiovascular disease is increasing in India as compared to western countries also found in younger age with hypercholesterolemia. CVD is an important cause of mortality and morbidity in India too. According to data of Registrar General of India, age-adjusted CVD is more in southern and eastern India. Studies reported the various factors. In real sense fast moving life-style and getting away from nature have invited number of physiological problems in general human being. Industrialization, heavy work load bringing the stress, changed dietary habits, lack of exercise, good consumption of beverages, fast food and lower intake of fruit and vegetables etc. have added fuel with physiological problems especially when high total serum cholesterol related problems are considered. World Health Organization has set the goal to reduce the global death rate by 2% a year up to 2015.[2] The goal is attributed with lowering of high total serum cholesterol as this is directly related with cardiovascular diseases. Various combination of statins are suggested as safe and potent medication for the problem throughout globe, but adverse effects are also reported with use as Australian Adverse Drug Reactions Advisory Committee had received 22 reports of paraesthesia associated with simvastatin.[3] Symptoms most frequently involved the face, scalp, tongue, and limbs and ranged from hypoaesthetic to hyperaesthetic sensations, although 4 cases of more serious neurological damage had also been reported. To avoid such side-effects, the whole world is looking towards alternative medicine and Ayurveda comes ahead amongst different traditional system of medicines with long lists of medicine for the purpose. The problem In Ayurvedic classical texts, a condition described by name of Medo Dhatu Dusti very much resembles hypercholesterolemia and this is resulted from imbalance of energy consumption and intake.

Number of tissues in any human body is rich in Sneha (oil) and they are Medodhatu, Vasa and Majjadhatu. They have Snehatva as common feature but they differ in their site and function.[4] Meda’s is present mainly in Udara, but some are also present in Mamsa and Asthi. Medas present in small Asthi is called as Sarakta Medas and those present in large Asthi, is termed as Majja. Pure Meda present in Mamsa is known as Vasa.[5] It is Medodhatu which is having significant role in developing many metabolic disorders like Medoroga etc. There are two types of Medodhatu by name of Poshaka and Poshya. Poshaka Medodhatu is mobile in nature which is circulated in whole body along with Rasarakta Dhatu. Its purpose is to provide nutrition to Poshya Medodhatu and recent researches have proved that cholesterol is circulated with blood, means Gatiyukta. Poshya Medo Dhatu is having immobile nature, which is stored in Medodharakala. The site of Medodharakala is Udara, Anuasthi, Sphika, Stana and Gala. Such conditions are result of derangement of metabolism of Medas. Excessive intake of Slesma Vardhaka Ahara produces Agnimandya. This Agnimandya brings Ama Dosa resulting in Ama Annarasa which is circulated in body. This vitiates Dosa, Dhatu, Srotasa, Bhutagni and Dhatwagni. Excessive vitiation occurs in Medodhatwagni which brings more production of Sama Medasa. This Sama Medasa circulates in body which brings Posaka and Posaka Dhatus in Sama condition. Due to this Medo Dhatu Dusti occurs. Medodushti is abnormal accumulation of Meda Dhatu in body. It includes several numbers of other Medovikaras, which are collectively known as Medoroga. Various formulations and plants are suggested for Medoroga viz., Musta, Haridra, Daruharidra, Vacha, Agnimantha, Agaru, Ashoka, Triphala, Trikatu, Chirabilva, Shimsapa etc. Reference regarding Chiravilva is found in Sutrasthana 38/8-9 of Sushruta Samhita as Kaphamedovishoshana. Less references are found regarding hypocholesterolemic effect of Chirbilva (Holoptlea integrifolia). So, present work is taken for study.

Materials and Methods

Pharmacological Study[6-10]
Animals Procurement and Standard Setup for Experiment

1. Male Wister Strain Albino rats weighing 150-250g were used for the study.


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Vinod K et al. An experimental evaluation of bark of Chirbilva

2. The animals were obtained from the animal house attached to the A.L.N Rao Memorial Ayurvedic Medical College, with Reg. No. -191/ CPCSEA, IAEC Approval no: A.E.D.G.03/11.
3. They were maintained on “Amrut” brand animal pellet feed of Pranav Agro Industries and plain tap water given ad libitum.
4. Animals were exposed to natural day and night cycles with ideal laboratory conditions in terms of ambient temperature (22 ± 3ºC) and humidity (50-60%).
5. The experiments were carried out after obtaining permission from “Institutional Animal Ethics Committee”.

Pilot Study

Maximum dose of cholesterol powder in rat is around 25g/kg/day. Average weight of rat is around 200 gm. Hence, dose of cholesterol comes as:25×200/1000 = 5mg A pilot study was conducted to fix the dose of the rat. For that 5mg of cholesterol powder was mixed with 5ml of vanaspati ghee by making it in lukewarm state, the 5ml of saturated solution was administered with the help of gastric tube, but on the next day the rat was not survived because of excess dose of vanaspati ghee. Therefore, dose of ghee was reduced to 2ml and was mixed with same 5mg of cholesterol powder in lukewarm state, it was observed that this particular dose was suitable for rat and easily digestible.

Autopsy Finding

On autopsy, for 5mg cholesterol in 5 ml vanaspati ghee, it was found that ghee regurgitated back and entered the airways of rat and to lungs, due to these, rat by choking.

Dose Fixation and Schedule

As, per FDA approved The Jounal of Korean Oriental Medicine formula for conversion of Human dose to rat dose is:

Rat dose (mg/kg) = HED (mg/kg) × Conversion factor Whereas, Conversion factor for rat is 6.17 (As per reference for specified weight of rats) Conversion of the dose obtained above to dose in mg/kg/day by multiplying with suitable conversion factor based on the average weight of the animal. Dose of Kashaya for human being = 2pala = 2×48 =96ml/60kg body weight So, HED = 1.6ml/kg body weight

This can be calculated as per formula that is:

Animal dose = 1.6 × 6.17 = 9.872/kg body weight As, the average weight of rat taken is in grams so, 9.872 × 210/ 1000 = 2.07ml/210 gm body weight of rat 2ml of freshly prepared Kashaya was given for experiment.

Dose fixation of standard drug:

Human dose of Atorvastatin is 10-80 mg once in a day. As per earlier studies both 20mg and 40mg human dose was proved effective in reduction of serum lipids in rats and there was no significant difference between them. Hence the dose selected was 20mg, conversion of human dose to rat dose based on formula.

Rat dose = HED × Conversion factor for rat that is 6.17 As, dose of Atorvostatin for human is 20 mg. Average weight of an individual is 60kg So, according to formula, 20/ 60 = 1/3 mg is per kg bodyweight for human Therefore, for rat 1/3 x 6.17 = 2.05 mg is per kg bodyweight For 210 gm rat, 2.05 mg x 0.210 = 0.43 mg 20 mg of standard was dissolved in 25 ml of distilled water.

0.5ml of Atorvostatin was administered in morning hours to the standard group.

Preparation of Kashaya

Fehling method was applied for removal of bark, then it was cut into small pieces and was allowed to dry in shade. When bark of both the drugs got dried then they are pounded well and make to coarse powder form by passing through the sieve no. 2000/355, According to W.H.O. guidelines. Then powders of both the bark were stored in air tight container in dark.

Test Drug:

To prepare freshly Twaka Kashaya of drug Chirbilva (Holoptelea integrifolia Planch.) as mentioned in classics (Sarangadhara) i.e., 1 Pala (48gms) of powdered drug was taken with 16 parts of water. It was reduced to 1/8th part. The mentioned methods were adopted to prepare Kashaya of bark of drugs Chirbilva (Holoptelea integrifolia Planch.) Roxb.). Fresh Kashaya was prepared under Mandagni (low flame) as per told in Sharangdhara every day to give to rats. The dose given was 2ml as obtained by dose conversion formula. The drug was administered in morning hours to both trial groups.


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Vinod K et al. An experimental evaluation of bark of Chirbilva

Experimental study design and protocol

Sample: 18 Albino rats of either sex will be randomly selected.

Inclusion: Criteria Healthy, active rats each weighing in between 150-200 g.

Exclusion Criteria: Pregnant, diseased rats weighing below 150 g or above 200 g and rats under trial for other experiments will be excluded.

Grouping

GroupsNo. of Albino ratsDrugsFormsDose / 200 g Body WeightPurpose
Control Group6Distilled water2mlTo serve as prophylactic control
Standard Group6Atorvastatin 20mgLiquid0.4mlAs positive Control
Trial Drug I6Stem bark of ChirabilvaKashaya2mlTo bring about Medohara effect

Initial body weights of all the animals were taken and the animals were fed on the hyperlipidemic diet except control (Group-1) where they were kept on pellet feed and water. Rats of groups 2 and 3 were given with hypercholesterolemic diet. In Group-2 standard drug Atorvastatin 20mg diluted in 25ml of water and 0.5ml of saturated solution was administered (in morning hrs) whereas Groups-3 received Chirabilva Twak Kshaya respectively for 30days. The hypercholesterolemic diet included hydrogenated vegetable oil (Vanaspati Ghee and Cholesterol extra pure powder) made into suspension form. The suspension was administered in the dose of 0.5 ml/210 g bodyweight of rats daily for 30 days (at evening hours) to all the rats. On the 30th day final weight of all the rats will be taken and on the 31st day after overnight fasting, rats were anaesthetized by giving mild dose of Ketamine subcutaneously. Then, the blood was collected by Retro-orbital puncture and was sent to laboratory for biocicalestimations[8-10]

Duration: 31days

Statistical Test: Anova followed by Post hoc. Scheffe test was done.

Observation 

  • Subjective Criteria: Weight (B.T. & A.T.) are taken
  • Objective Criteria: HDL, LDL, VLDL, Total Cholestrol, Triglycerides

Collection of Blood

According to guidelines for Collection of Blood from Laboratory Animals following procedure is opted for collection of blood: Under general anesthesia rats were grasped so that its back rested on palm with its head toward thumb. The thumb was placed just lateral to animal's trachea so that jugular vein on same side as eye from which occluded blood was collected. The fur on animals head was drawn into palm, this caused animal’s eye to proptose (bulge) slightly. A 50 μL sterile micro hematocrit tube was directed into medial canthus (junction of eyelids closest to animal's nose) of eye rotating slightly as tube was directed to a point directly behind globe. Sufficient pressure was applied to cut through fibrous layer that surrounded sinus. Blood flowed through tube once sinus had been penetrated. After blood collection, tube was removed and eyelids were closed and a dry cotton pad was applied over eye with gentle pressure to prevent retro-orbital hemorrhage. Blood was not collected from same eye more than 2 times. An antibiotic ophthalmic ointment was applied following bleeding.

Mode of Drug Administration: Oral route was selected for administration of drug to respective group of animals by using syringe with an attached gastric tube.

Criteria for assessment of results: Blood cholesterol levels were checked before and after treatment. Blood was drawn from the rats by retro-orbital method. These values were subjected to statistical analysis to evaluate the hypercholesterolemic activity.

Results

Animal Experiment

jaims_3703_01.JPG
Animals (Rats) in Cage


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Vinod K et al. An experimental evaluation of bark of Chirbilva

jaims_3703_02.JPG
Feeding to Rat

jaims_3703_03.JPG
Collection of Blood by Retro-orbital Puncture

Bodyweight Comparison:

GroupBodyweight (Before Treatment)Mean + SDBodyweight (After Treatment)Mean + SD
Control210.00 + 14.49236.66 + 9.83
Standard216.66 + 10.80235.83 + 11.58
Trial I211.66 + 14.71246.66 + 9.83

Mean of Change in Bodyweight

GroupsMean + SD
Control26.66 + 6.83
Standard18.33 + 4.08
Trial I35.00 + 6.32

Mean and Standard Deviation of Total Cholesterol (mg/dl) Before and After Treatment

GroupsBefore TreatmentAfter Treatment
Control53.60 + 4.4855.13 + 4.57
Standard53.81 + 2.6941.36 + 2.66
Trial I52.85 + 3.3343.90 + 2.65

GroupsBefore TreatmentAfter Treatment
Control68.03 + 4.7569.56 + 4.59
Standard74.88 + 7.7651.35 + 6.79
Trial I71.66 + 8.8552.27 + 8.58

Mean and Standard Deviation of Triglycerides (mg/dl) Before and After Treatment

Mean and Standard Deviation of HDL (mg/dl) Before and After Treatment

GroupsBefore TreatmentAfter Treatment
Control21.13 + 1.8520.26 + 1.88
Standard23.51 + 2.5124.23 + 2.57
Trial I23.52 + 2.1124.09 + 2.17

Mean and Standard Deviation of LDL (mg/dl) Before and After Treatment

GroupsBefore TreatmentAfter Treatment
Control18.86 + 3.7220.98 + 4.04
Standard15.31 + 1.556.69 + 1.59
Trial I15.00 + 0.759.18 + 1.74

Mean and Standard Deviation of VLDL (mg/dl) Before and After Treatment

GroupsBefore TreatmentAfter Treatment
Control13.60 + 0.9513.88 + 0.89
Standard14.97 + 1.5510.27 + 1.35
Trial I12.87 + 1.8310.45 + 1.71

Discussion

When bodyweight before and after treatment was compared, it was seen that increase in weight was maximum with Trial I being 35 gm while it was 19.17 gm, 34gm and 26.66 gm respectively. When total cholesterol was attained, it was increased by 1.53 gm/dl for control group while for standard it got reduced, 12.45 gm/dl. For Trial drugs, quantity was reduced by 8.95 gm/dl. Total Triglycerides were observed increasing in control group slightly by 1.53 mg/dl while it was reduced by 23.53 mg/dl, 19.39 mg/dl respectively for standard, Trial drug I HDL was noted decreased by 0.87 mg/dl while it was marked increased by 0.72mg/dl and 0.57 mg/dl in sequence for standard, Trial drugs I.

Conclusion

Mean of bodyweight changes before treatment and after treatment was 26.66, 18.33 and 35 respectively for control, standard and trial I groups. Mean of changes in total cholesterol before and after treatment in mg/dl for control, standard and trial I were respectively 1.53 (increased), 12.45 (reduced) and 8.95 (reduced). Mean of changes in triglycerides before and after treatment in mg/dl for control, standard and trial I were respectively 1.53 (increased), 23.53 (reduced) and 19.39 (reduced).


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Mean of changes in HDL before and after treatment in mg/dl for control, Standard and trial I were respectively 0.87 (reduced), 0.72 (increased) and 0.57 (increased). Mean of changes in LDL before and after treatment in mg/dl for control, Standard and trialI were respectively 2.12 (increased), 8.62 (reduced) and 5.82 (reduced). Mean of changes in VLDL before and after treatment in mg/dl for control, Standard and trial I were respectively 0.28 (increased), 4.70 (reduced) and 2.42 (reduced). Holoptelea bark was having better in action in case of HDL, LDL and VLDL. Statistical analysis of total cholesterol by means of ANOVA revealed Bark as significant after treatment.

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10. Madhavi Jagtap et al. (2011) ‘A comparative evaluation of cardioprotective activity of two Makandi (Coleus forskohlii ) formulations against isoproterenol induced myocardial infarction in hyperlipidaemic rats’. Indian Journal of Natural Products and Resources. Volume 2(3), September 2011,pp.335-344.


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