2018 SCIENCELINE

Journal of Life Science and Biomedicine

J Life Sci Biomed, 8(2): 31-36, 2018

License: CC BY 4.0

ISSN 2251-9939

Prophylactic Administration of Ginkgo biloba Leaf

Extract (EGb 761) Inhibits Inflammation in

Carrageenan Rat Paw Edema Model

Sani Abdulrazak ^1,2^, Abdulmumin Abdulkadir Nuhu¹, and Zakka Israila Yashim¹

¹Department of Chemistry, Ahmadu Bello University, Zaria, Kaduna, Nigeria

²Department of Veterinary Physiology, Ahmadu Bello University, Zaria, Kaduna, Nigeria

Corresponding author’s Email: sazzak175@gmail.com

ABSTRACT

Original Article

PII: S225199391800006-8

Acute toxicity and anti-inflammatory effect of Ginkgo biloba leaf extract (EGb 761)

were carried out in this study. The anti-inflammatory activity was studied using the

carrageenan model whereby twenty rats were randomly divided into four groups of

five animals each. Groups one and two were administered the EGb 761 extract at 500

mg/kg and 250 mg/kg, respectively. Rats in groups three (positive control group)

and four (non-treated control group) were given piroxicam (10 mg/kg) and normal

saline (5 ml/kg), respectively. Oedema was induced by injecting 100 μl of fresh

carrageenan into the right plantar surface of the hind paw of each rat 30 minutes

after administration. The acute toxicity tests result showed that the extract is safe

at 5000mg/kg dose. Ginkgo biloba leaf extract caused a significant (P˂0.05) decrease

in the size of the paw oedema when compared to control. Of interest, EGb 761 at 250

mg/kg was as effective as, or better than piroxicam (10 mg/kg). These findings

further justify the use of Ginkgo biloba leaf extract in both medical and ethnomedical

practice and may be used in treatment of inflammation.

Rec. 08 Dec. 2017

Acc. 22 Feb. 2018

Pub. 25 Mar. 2018

Keywords

Ginkgo Biloba Leaf

Extract,

Carrageenan,

Rats,

Paw Oedema,

Inflammation

ABBREVIATION

EGb 761

GABA

IL-4

IL-6

LD₅₀

mg/kg

ml/kg

Ginkgo biloba leaf extract

Gram

Ɣ- aminobutyric acid

Interleukin

Interleukin-4

Interleukin-6

Lethal dose 50

Milligram per kilogram

Milliliter per kilogram

Nitric oxide

Prostaglandin

Standard error of mean

Microlitre

SEM

μL

To cite this paper: Abdulrazak S., Nuhu A.A., and Yashim Z.I. 2018. Prophylactic Administration of Ginkgo biloba Leaf Extract (EGb 761) Inhibits

Inflammation in Carrageenan Rat Paw Edema Model. J. Life Sci. Biomed. 8(2): 31-36; www.jlsb.science-line.com

INTRODUCTION

Inflammation is the body’s physiologic defense mechanism against infection, burn, toxic chemicals, allergens or

other noxious stimuli [1, 2]. Diseases and disorders are manifested through inflammatory responses as the body

recognizes the injury and prepares to repair the damage [3]. Endogenous mediators like prostaglandins,

histamine, serotonin, bradykinin, etc. are liberated when inflammation occurs. Prostaglandins (PG) indicate and

modulate the body’s response to inflammation. These substances can elicit pain response which in turn causes

dropped muscular activities [4].

Medicinal plants have provided biologically relevant products for centuries, and are still a source for new

medicines [5]. Ginkgo biloba is a widely used plant in treatment of asthma, bronchitis, hearing loss, tuberculosis,

cognitive dysfunction, stomach pain, skin problems, and anxiety [5, 6, 7]. Ginkgo biloba leaf extract (EGb 761)

contains flavonoids and triterpenes as the main active ingredients, and these substances possesses anti-

inflammatory activity [8]. The extracts of Ginkgo biloba is said to have promising anti-inflammatory effect.

Although it involves other mechanisms, interleukin (IL) is one of the most important in the anti-inflammatory

functions of Ginkgo biloba [9]. Haines et al. [10] showed that the synergistic interaction of Ginkgo biloba leaf

extract (EGb 761), astaxanthin and vitamin C suppress respiratory inflammation in asthmatic guinea pigs.

Bao et al. [11] reported that EGb 761 alleviate inflammatory reactions. This is done as a result of

heightened activity of Interleukin-4, an anti-inflammatory cytokine, and inhibition of Interleukin-6 (IL-6), an

inflammatory cytokine by dual activity. Using carrageenan model, Thorpe et al . [12] reported that EGb 761 has

anti-inflammatory activity. Similarly, Ou et al. [13] also reported that inflammatory processes resulting from

oxidized low density lipoproteins-induced oxidative stress in vascular endothelial cells were ameliorated by the

administration of Ginkgo biloba extract.

The anti-inflammatory agents of plant origin have been the major focus of most research globally. Thus,

evaluation of anti-inflammatory effects of Ginkgo biloba leaf extract is of great importance in the effective

treatment and prophylaxis of several disease conditions in both humans and animals.

MATERIAL AND METHODS

Experimental animals and Ethical approval

Albino rats weighing an average of 180 g were acclimatized for 2 weeks prior to the experiment, fed

standard diet and water was provided ad-libitum. All animal experimentation was done in accordance with

Ahmadu Bello University Animal Use and Care Guidelines. Ethical clearance with approval number

ABUCAUC/2016/015 was obtained from Committee on Animal Use and Care, Directorate of Academic Planning

and Monitoring, Ahmadu Bello University, Zaria before the commencement of the study.

Experimental design

Acute toxicity study

The method of Lorke [14] with modification was used to determine the median lethal dose (LD₅₀) of the

extracts in rats. This modification involves the introduction of uniform number of rats per group and the use of

18 albino rats instead of 12 for the study. In this study, 18 albino rats were randomly allocated into 6 groups of 3

rats each. The animals were starved of food ad libitum and water for 12 hours to avoid formation of complexes

with food substances. Groups 1, 2, 3, 4, 5 and 6 were treated with the extract orally at 10, 100, 1000, 1600, 2900

and 5000 mg/kg body weight respectively. Rats were observed for 48 hours for any sign of toxicity or mortality.

Anti-inflammatory study

The method as described by Suleiman et al. [15] with modification was employed. Twenty rats were

randomly divided into four groups of five animals each. Groups one and two received the extract at 500 mg/kg

and 250 mg/kg, respectively. Rats in groups three (positive control group) and four (non-treated control group)

were given piroxicam (10 mg/kg) and normal saline (5 ml/kg), respectively. All treatments were administered by

oral route. Oedema was induced by injecting 100 μL of fresh carrageenan into the right plantar surface of the

hind paw of each rat 30 minutes after administration. The paw diameter was measured at 0, 30 minutes, 1,

2,3,4,5, and 6 hours after administration.

To cite this paper: Abdulrazak S., Nuhu A.A., and Yashim Z.I. 2018. Prophylactic Administration of Ginkgo biloba Leaf Extract (EGb 761) Inhibits

Inflammation in Carrageenan Rat Paw Edema Model. J. Life Sci. Biomed. 8(2): 31-36; www.jlsb.science-line.com

Statistical Analysis

Data were expressed as mean ± standard error of mean (S.E.M) and then analysed by one-way analysis of

variance (ANOVA) followed by Tukey’s post-hoc test. The analyses were done using Graphpad Prism version 5.

Values of P<0.05 were considered significant.

RESULTS

Acute Toxicity Study

Table 1 shows the results of acute toxicity study of Ginkgo biloba leaf extract (EGb 761). The extract

administered at doses of 10, 100, and 1000, 1600, 2900, and 5000 mg/kg respectively did not produce any sign of

toxicity or mortality. Also, Ginkgo biloba leaf extract (EGb 761) did not alter the behavior of the animals during

the period of the study. Therefore, Ginkgo biloba leaf extract is considered relatively safe.

Anti-inflammatory study

Sub-plantar injections of carrageenan induced inflammation as evident in the increased paw diameter of

the untreated control rats. Oedema was visible within the first 5-10 minutes of administration of carrageenan,

the peak of swelling occurred approximately 2-3 hours following injection of carrageenan. Ginkgo biloba leaf

extract produced a significant (P<0.05) decrease in the size of the paw oedema as shown in Table 2. The activity

of Ginkgo biloba leaf extract was highest at 250 mg/kg after 3 hours and was comparable to Piroxicam (standard

anti-inflammatory agent; 10 mg/kg).

Table 1. Acute toxicity study of Ginkgo biloba leaf extract (EGb 761)

Groups

Group 1

Group 2

Group 3

Group 4

Group 5

Group 6

Dose/Day

10 mg/kg

100 mg/kg

1000 mg/kg

1600 mg/kg

2900 mg/kg

5000 mg/kg

Mortality (x/N)

0/3

*Group 1 (10 mg/kg Extract); Group 2 (100 mg/kg Extract); Group 3 (1000 mg/kg Extract); Group 4 (1600 mg/kg Extract); Group 5 (2900

mg/kg Extract); Group 6 (5000 mg/kg Extract).

Table 2. Effect of Ginkgo biloba leaf extract on carrageenan induced acute inflammation measured as paw size in

mm (mean ± SEM)

0 hr

3 hrs

4 hrs

5 hrs

6 hrs

Items

0.5 hr

1 hr

2 hrs

Group A

Group B

3.32±0.18^a

4.89±0.23

4.64±0.40

5.71±0.25

5.14±0.32

5.99±0.20

5.97±0.38

5.94±0.30

5.60±0.56

5.23±0.39

4.48±0.17

3.72±0.19

3.55±0.17^a

3.00±0.14

4.60±0.35^a

3.92±0.25^a

Group C

Group D

2.46±0.18^b

2.55±0.13^b

4.36±0.22

5.02±0.11

4.72±0.18

5.65±0.25

5.52±0.09

6.67±0.48

5.13±0.16^a

6.83±0.49^b

4.91±0.16

4.02±0.20^a

5.30±0.30^b

3.47±0.13^a

4.34±0.21^b

6.16±0.30^b

*ANOVA: Indicates that Comparism for all groups is statistically significant (P˂0.05) within the same column. Tukey's test: Means having

different superscript (^a,b) letters are significantly different (P˂0.05). Group A (500 mg/kg Extract); Group B (250 mg/kg Extract); Group C

(Piroxicam (10 mg/kg); Group D (Normal saline (5 ml/kg).

DISCUSSION

Acute Toxicity Study

Toxicological study is first assayed to determine the safety of drugs and plant products for human and

animal use [15]. The calculated LD₅₀of Ginkgo biloba leaf extract (EGb 761) was greater than 5000 mg/kg. This

value falls within the practically non-toxic range [14]. Doses up to 5000 mg/kg, orally administered, did not alter

the behavior of the animals during the period of the study, thus, the extract was considered relatively safe.

This finding was consistent with the outcome of a similar study carried out by Salvador [16], who

reported that the LD₅₀of standardized Ginkgo biloba extract administered orally to mice was 7,730 mg/kg. He

To cite this paper: Abdulrazak S., Nuhu A.A., and Yashim Z.I. 2018. Prophylactic Administration of Ginkgo biloba Leaf Extract (EGb 761) Inhibits

Inflammation in Carrageenan Rat Paw Edema Model. J. Life Sci. Biomed. 8(2): 31-36; www.jlsb.science-line.com

also reported no organ damage or impairment of hepatic or renal function when Ginkgo biloba extract was

administered orally over 27 weeks to rats and mice at doses ranging from 100 to 1,600 mg/kg.

Anti-inflammatory study

Results from this study suggest Ginkgo biloba leaf extract possessed anti-inflammatory effect. This may

be as a result of inhibition of inflammatory mediators, such as nitric oxide (NO), prostaglandins, and

proinflammatory cytokines into the paw tissue, because evidence shows that Ginkgo biloba and its constituents

suppress induction of these mediators [17].

Of interest, EGb 761 at 250 mg/kg was as effective as, or better than piroxicam (10 mg/kg). However,

administration of higher dose (500 mg/kg) of the extract did not produce such or higher anti-inflammatory

effect. This may not be unconnected to the reports of Ivic et al. [18] and Kiewert et al. [19] that EGb 761 contains

triterpenes; ginkgolides and bilobalide, and these active components at higher doses are known antagonists at

both glycine and Ɣ- aminobutyric acid (GABA) in the body, which are neurotransmitters that are known to

inhibit the activities of neurons that activate the release of inflammatory agents and regulate inflammation in

the body.

Our finding is consistent with the work of Abdel Salam et al. [20] and Han [21], who reported that oral

administration of Ginkgo biloba extract significantly reduced carrageenan induced paw oedema. Other studies

have shown that treatment with Ginkgo biloba extract (30–120mg/kg; orally) reduced inflammation and acute

colonic damage induced by acetic acid [22]. Similar studies on the anti-inflammatory properties of flavonoids,

quercetin and kaempferol have also demonstrated reduced carrageenan-induced hind paw oedema in mice [23].

However, our result disagrees with the findings of Biddlestone et al. [24], who reported that Ginkgo biloba had

no effect on paw oedema regardless of dose or duration of administration.

CONCLUSION

This study shows that Ginkgo biloba leaf extract (EGb 761) is practically non-toxic and is considered

relatively safe. Also, the extract possessed prophylactic anti-inflammatory effect and was as effective as, or

better than Piroxicam, a standard anti-inflammatory drug.

DECLARATIONS

Acknowledgement

We appreciate Abdulwahab Hashimu Yau and Yusuf Abdulraheem Oniwapele of the Department of

Veterinary Pharmacology and Toxicology, Ahmadu Bello University, Zaria for their technical support.

Authors’ Contributions

AAN designed the study. SA, AAN, and ZIY carried out the experimental research, collected the data,

analysed and interpreted the results. The first draft of manuscript was prepared by SA and reviewed by the rest

of the authors and the final version of the manuscript was read and accepted by all the authors.

Ethics Committee Approval

This experimental research was approved by the committee on animal use and care, directorate of

academic planning and monitoring, Ahmadu Bello University, Zaria. Ethical clearance with approval number

ABUCAUC/2016/015 was obtained for this experiment.

Consent to Publish

Not applicable

Competing Interests

The authors declare that there is no conflict of interest.

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To cite this paper: Abdulrazak S., Nuhu A.A., and Yashim Z.I. 2018. Prophylactic Administration of Ginkgo biloba Leaf Extract (EGb 761) Inhibits

Inflammation in Carrageenan Rat Paw Edema Model. J. Life Sci. Biomed. 8(2): 31-36; www.jlsb.science-line.com