IJAMS I International Journal of Ayurveda & Medical Sciences ISSN: 2455-6246

ABSTRACT

Experimental Evaluation of Local Effect of Glycyrrhiza glabra Linn. on Bleeding time in Wistar Strain Albino Rats

Richa Khandelwal, Sumit Nathani, Gaurav Sharma

Department of Dravya Guna, National Institute of Ayurveda, Jaipur, Rajasthan, India

INTRODUCTION

Glycyrrhiza glabra Linn. (G. glabra) which is known by the

name of Yashtimadhu or Madhuka in Ayurvedic classics, is an

abundantly used drug in different ailments. Madhuka is mentioned

in 11 Mahakashaya (group of 10 drugs) in Charaka Samhita, viz.

Jivaniya (~vitalisers), Sandhaniya (~union promoting), Varnya

(~Complexion promoting), Kanthya (~beneficial for throat),

Kandughna (~anti-pruritics), Snehopaga (~sub-oleatives),

Vamanopaga (~adjuvants of emesis), Asthapanopaga (~adjuvants of

asthapana type enema), Mutravirajaniya (~correcting the color of

urine), Angamardaprashamana (~cure the malaise) and Shonita-

sthapana (~haemostatics). [1]

It is one of the drugs of Madhura

skandha [2]

(~group of drugs having sweet taste). Among 36 groups

of drugs (gana) of Sushruta Samhita, Madhuka is included in 8

groups. [3]

In Ashtanga Hridaya, Madhuka has been included in ten

out of thirty three Gana. [4]

All this indicates that it is a broad-

spectrum drug which is used very extensively in Ayurvedic

pharmacology.

As per Ayurvedic classics, Yashtimadhu is used in various

bleeding conditions, both internally and externally. Oil which is

prepared with Madhuka and milk is used in nasal bleeding

conditions. [5]

Madhuka decoction, which is prepared with some

other drugs, is used for parisheka (~hot fomentation), [6]

and for

avagahana (~sitz bath), [7]

in excessive bleeding in Arsha (~piles).

Sushruta has recommended use of Madhuka as avacurnana dravya

(~sprinkled powder) in Atisruta rakta (~haemorrhagic condition). [8]

Sushruta used Madhuka as a very good wound cleanser and healer.

In Ashtanga Hridaya, milk prepared with Madhuka is indicated in

Raktashthivi kasa (~bleeding with cough). [9]

All the brihattrayi

granthas (~three big treatise of Ayurveda) indicate use of

Yashtimadhu in various bleeding disorders like Raktapitta

(~bleeding disorder) and Raktatisara (~diarrhoea with bleeding). As

per Chakradatta, Madhuka and Chandana (Santalum album Linn.)

pounded with milk should be taken in hematemesis. [10]

He also used

Madhuka in Atisruta rakta (~haemorrhagic condition). [11]

Use of

this plant in various bleeding conditions is seen in other classics of

Ayurveda also.

Also, many products and formulations are available in

market as hemostatic agents and the common content in most of

them is G. glabra, as it is claimed as an agent for cessation of

bleeding on external injuries. [12]

However, its main derivative

glycyrrhizin shows antithrombotic activity. [13]

Thus Madhuka is very commonly used in various bleeding

conditions externally as well as internally.

Corresponding author: Dr. Richa Khandelwal,

P.G. Scholar, Department of Dravya Guna, National Institute of

Ayurveda, Amer Road, Jaipur, Rajasthan,India.

Email: khandelwalricha98@gmail.com

ORIGINAL RESEARCH ARTICLE (EXPERIMENTAL)

Background: Glycyrrhiza glabra Linn. (G. glabra) is the botanical source of Yashtimadhu, which has been recommended for internal as well as

external use by Ayurveda classics in various bleeding conditions. Aims and Objectives: This study was carried out to evaluate the effect of local

application of G.glabra on the bleeding time (BT) in wistar strain albino rats. Material and Method: 24 adult wistar strain albino rats were

divided into four groups having six rats in each. Group A was treated as Control Group. Group B (Test group 1) was treated with Glycyrrhiza

root powder locally. Group C (Test group 2) was treated with Glycyrrhiza root decoction locally. Group D was treated as standard group.

Result: The BT of rats treated with Glycyrrhiza powder and Glycyrrhiza decotion externally was not significantly affected, when compared with

control. Conclusion: G. glabra does not have any significant activity on bleeding time in wistar strain albino rats after local application in the

form of powder or decoction.

Key words: Albino Wistar Rats, Bleeding time, Glycyrrhiza glabra Linn., Local effect

Khandelwal, et al.: Local Effect of Glycyrrhiza Glabra Linn. on Bleeding Time in Albino Wistar Rats

International Journal of Ayurveda & Medical Sciences I July-September 2017 I Vol 2 I Issue 3

60

Aim of this study

This study was carried out to evaluate the effect of G. glabra on

the bleeding time (BT) by tail cut method in wistar strain albino rats,

after local application of G. glabra root powder and decoction.

MATERIALS AND METHODS

Experimental study was conducted in Institute of Biomedical

and Industrial Research, Vidyadhar Nagar, Jaipur (CPCSEA

Registration No: 1737/PO/Rc/S/14/CPCSEA), after approval from

the Institutional Ethics Committee (IAEC Approval No:

IBIR/IAEC/2015/I/4).

Test Sample

Glycyrrhiza glabra Linn. dried roots was purchased from Jaipur

crude drug market and authentication was done from Raw Material

Herbarium and Museum, Delhi (RHMD) of National Institute of

Science Communication And Information Resources (NISCAIR)

with Ref. no. NISCAIR/RHMD/Consult/2016/2977-04.

Powder was made by the grinder. For making decoction, 10 grams

of Glycyrrhiza root powder was boiled with 40 ml of water and

reduced to around 25 ml and filtered. This decoction was used for

the experiment.

Experimental Animals

24 wistar strain albino rats of either sex weighing 150 20 g

were used for this experiment.

Housing and feeding conditions

The temperature in the experimental animal room was

maintained at 22C (3C) and relative humidity was maintained at

50-60% during room cleaning. Lighting was artificial, with a

sequence of 12 hours light and 12 hours dark. For feeding,

conventional laboratory diets of Gulmohar brand animal feed

manufactured by Lipton India Limited was used with an unlimited

supply of drinking water.

Group Design

24 adult wistar strain albino rats were divided into three

groups having six rats in each. These groups received different

treatment in following manner:

Group A (Control Group): No any application applied on six albino

wistar rats.

Group B (Test Group 1): Six wistar strain albino rats had been

applied locally Glycyrrhiza root powder.

Group C (Test Group 2): Six wistar strain albino rats had been

applied locally Glycyrrhiza root decoction.

Group D (Standard Group): Six wistar strain albino rats had been

applied locally Feracrylum solution.

Experimental procedure

The adult wistar strain albino rats (150 20 g) were

randomly divided into four groups of six rats per group. The rats

were slightly anesthetized. Bleeding time was measured by cutting

off the rats tail tip (5 mm), and no any application (in control

group), after applying test drug (in test group 1 and test group 2) and

standard drug (in standard group) and blood was blotted on a filter

paper at 15 sec interval until no blood was observed on the filter

paper. The period between the tail amputation and the cessation of

bleeding was taken as the bleeding time in minutes.

Statistical analysis

The results were expressed as mean SEM. Comparison

between the treatment groups and control were performed by

analysis of variance (ANOVA) followed by Dunnets multiple

comparison test. In all tests the criterion for statistical significance

was P < 0.05.

RESULTS

The effect of G. glabra on the bleeding time in wistar strain

albino rats were summarized in [Table 1] and [Table 2]. The BT of

rats treated Glycyrrhiza root powder and Glycyrrhiza decotion

externally was not significantly affected compared with control.

Table 1 : Bleeding time ( Mean SEM) in Group A,B,C,D

Group A Group B Group C Group D P value and

Significance Mean SEM Mean SEM Mean SEM Mean SEM

221.8321.916 265.3332.809 262.6745.944 8845.100 0.0105 (S)

Fig 1: Mean Bleeding time in Group A,B,C,D

Table 2: Multiple Comparisons in Between Groups

Dunnett's multiple

comparisons test

Mean

Diff.

95.00% CI of

diff.

Significant

? Summary

Adjusted P

Value

Group A vs. Group B 43.5 179.1 to 92.15 No Ns 0.7553

Group A vs. Group C 40.83 176.5 to 94.81 No Ns 0.7867

Group A vs. Group D 133.8 1.815 to 269.5 Yes * 0.0436

Group A (Negative Control) without application of any

medicine, showed mean bleeding time 221.83 sec, Group B (Test

Group 1) with application of Glycyrrhiza root powder, showed mean

bleeding time 265.33 sec, Group C (Test Group 2) with application

of Glycyrrhiza decoction show mean bleeding time 262.67 sec,

Group D (Positive Control) with application of Feracrylum Solution,

showed mean bleeding time 88 sec.[Table 1], also shown by[Fig. 1].

0.00

50.00

100.00

150.00

200.00

250.00

300.00

Group A Group B Group C Group D

Bleeding Time in Sec

Bleeding Time in Sec

Khandelwal, et al.: Local Effect of Glycyrrhiza Glabra Linn. on Bleeding Time in Albino Wistar Rats

International Journal of Ayurveda & Medical Sciences I July-September 2017 I Vol 2 I Issue 3

61

Statistically comparison between Negative Control, Test

Group and Positive Control found P value 0.0105 and it is

significant. Then multiple comparision was done. Statistically

comparison between Group A vs Group B has been shown

insignificant results (P value = 0.7553), Group A vs Group C has

been shown insignificant results (P value = 0.7867) and Group A vs

Group D has been shown significant results (P value = 0.0436)

DISCUSSION

In this experiment, blood coagulation effect after local

application of test drug G. glabra root (in two different form

powder and decoction), was evaluated by rat tail cut method in four

different group (Negative Control, two Test Groups and Positive

Control) and bleeding time was recorded. It is suggestive of no

significant effect of test drug on bleeding time after local application

in powder and decoction both forms.

Blood coagulation is a host defense mechanism that assists in

maintaining the integrity of the closed, high-pressure mammalian

circulatory system after blood vessel injury. In the abnormal

conditions, it is also involved in the thrombosis, atherosclerosis,

inflammation and metastasis by the activation of enzymes in the

coagulation cascade and the platelets. [14]

Bleeding time is a measure of the primary hemostasis where-

in platelets plugs are formed by collagen-induced aggregation of

platelets and thrombin-induced formation of fibrin. [15]

In this

experiment bleeding time was evaluated.

Bleeding time is affected by many factors including

vasoconstrictive effect of blood vessels, the formation of hemostatic

plug and platelet activity. In general, fatty acids, palm oil and aspirin

have been reported to increase BT in animals and humans, whereas

saturated fatty acids and cholesterol decrease BT. [16]

The present

work shows that Glycyrrhiza have no any significant effect on

bleeding time in powder and decoction both forms after external

application when compared with control.

In this study, the effect of test drug on bleeding time was

evaluated on the basis of some classical references like in Sushrut

Samhita, this drug has been used (with some other drugs) to stop the

hemorrhage or excessive bleeding (Atisrut rakta chikitsa), but no

significant result was found on bleeding time in experimental model.

This indicates that G. glabra do not have any activity on bleeding

time in wistar strain albino rats by rat tail cut method.

In Ayurveda classics this drug has been used for blood

clotting internally as well as externally. In Sushruta Samhita,

Sutrasthana, this is used for Atisrut rakta chikitsa (excessive

bleeding treatment) in the form of powder applying externally.

To evaluate the coagulation effect of G. glabra bleeding time

was evaluated in this experiment but bleeding time is not a single

parameter to evaluate the coagulation. Some other factor like

clotting time, prothrombin time, activated plasma thromboplastim

time, INR ratio are also related with the coagulation. May be these

parameters are affected by G. glabra rather than bleeding time

which was chosen in this experiment. Hence the concept of classical

application of G. glabra which has mentioned by Acharya Sushruta

in Atisrut rakta chikitsa may be considered through other such

surrounding area of parameters.

CONCLUSION

On the basis of this experiment, we concluded that G. glabra

does not have any significant activity on bleeding time in wistar

strain albino rats after local application in the form of powder and

decoction both. For further evaluation some other factor like clotting

time, prothrombin time, activated plasma thromboplastim time, INR

ratio are also will be included, because these are important factor for

coagulation.

REFERENCES

1. Yadavji Trikamji acharya (Ed.), Charaka Samhita of Agnivesha

elaborated by Charaka and Dridhbala with the Ayurvedadipika

commentary by Chakrapanidatt, Chaukhambha Surbharati

Prakashana, Varanasi, 2014 Edition, Sutra Sthana 4th

chapter,

verse no. 9-18, pg. no. 32-34.

2. Ibidem (1). Charaka Samhita, Vimana Sthana 4th chapter, verse

no. 139, pg. no. 284.

3. Jadavji Trikamji (Ed.), Narayan Ram Acharya Kavyatirth (Ed.),

Sushruta Samhita of Sushruta with the Nibandhsangrha

commentary of Dalhanacharya and the Nyayachandrika Panjika

of Gayadasacharya on Nidana Sthana, Chaukhambha Surbharti

Prakashan, Varanasi, 2017 Edition, Sutra Sthana, Chapter 38th

,

pg. no. 164-170.

4. Pt. Hari Sdashiva Shastri Paradkar (Ed.), Astangahridaya of

Vagbhatta with the commentaries Sarvangsundara of Arundatta

and Ayurveda Rasayana of Hemadri, Chaukhambha Surbharti

Prakashan, Varanasi, 2016 Edition, Sutra Sthana, Chapter 15th

,

pg. no. 230-240.

5. Ibidem (1). Charaka Samhita, Chikitsa Sthana 4th Chapter, verse

no. 101, pg. no. 434.

6. Ibidem (1). Charaka-Samhita, Chikitsa Sthana 14th Chapter, verse

no. 214, pg. no. 510.

7. Ibidem (1). Charaka-Samhita, Chikitsa Sthana 14th Chapter, verse

no. 215, pg. no. 510.

8. Ibidem (3), Sushruta Samhita, Sutra Sthana, Chapter 14th, verse

no. 36, pg. no. 66.

9. Ibidem (4), Astanghridaya Chikitsa Sthana, Chapter 3rd, verse no.

85, pg. no. 593.

10. Tripathi Indradev, Chakradatta of Shri Chakrapanidatta with the

Vaidyaprabha Hindi Commentary, Chaukhambha Sanskrit

Bhawan, Varanasi, Edition 2014, Chardi Chikitsa Adhyaya,

verse no. 26, pg. no. 117.

Khandelwal, et al.: Local Effect of Glycyrrhiza Glabra Linn. on Bleeding Time in Albino Wistar Rats

International Journal of Ayurveda & Medical Sciences I July-September 2017 I Vol 2 I Issue 3

62

11. Ibidem (10), Chakradatta, Siravyadhavidhi adhyaya, verse no.

16, pg. no. 435.

12. Haznedaroglu Berat Z. et al. Ultrastructural and Morphological

Analyses of the in-vitro and in-vivo Hemostatic Effects of

Ankaferd Blood Stopper. Clinical and Applied

Thrombosis/Hemostasis, 2009;16(4):446-453

13. Mendes-silva Wallace et al. Antithrombotic effect of

Glycyrrhizin-a plant-derived Thrombin Inhibitor. Thrombosis

Research, 2003; 112 (1-2):93-98

14. Yu Lin G.O.U, Alice Lok Sze H.O., Dewi Kenneth Rowlands,

Yiu Wa CHUNG, and Hsiao Chang Chan. Effects of Bak Foong

Pill on Blood Coagulation and Platelet Aggregation. Biol.

Pharm.Bull ., 2003; 26(2), 241246

15. GKA Adepoju, T Adeyemi. Evaluation of the Effect of Lime

Fruit Juice on the Anticoagulant Effect of Warfarin. J Young

Pharm. 2010;2(3): 269-272

16. Hadi Abdul-Hadi A. , Elderbi Mahmoud A., Mohamed Abdel-

Wahab H. Effect of Gum Arabic on Coagulation System of

Albino Rats. Int.J. Pharm.Tech. Res.2010,2 (3): 1762-1766.

How to cite this article: Khandelwal R, Nathani S, Sharma G.

Experimental Evaluation of Local Effect of Glycyrrhiza glabra

Linn. on Bleeding Time in Wistar Strain Albino Rats. Int J

Ayurveda & Med Sc 2017; 2(3): 59-62.

Source of Support: Nil. Conflict of Interest: None declared.