Atta 1998- Plantas Antiinflamatorias

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Journal of Ethnopharmacology 60 (1998) 117124

Anti-nociceptive and anti-inflammatory effects of someJordanian medicinal plant extracts

A.H. Atta a, A. Alkofahi b,*

a Department of Veterinary Basic Sciences, Faculty of Veterinary Medicine, Jordan Uniersity of Science and Technology,

P.O. Box 3030, Irbid, Jordanb Department of Medicinal Chemistry and Pharmacognosy, Faculty of Pharmacy, Jordan Uniersity of Science and Technology,

P.O Box 3030, Irbid, Jordan

Received 15 October 1996; received in revised form 14 July 1997; accepted 4 November 1997

Abstract

The anti-nociceptive effect of ethanolic extract of 11 traditionally used Jordanian plants was studied by using the

acetic acid-induced writhing and hot-plate test in mice. The anti-inflammatory effect of these plants was determined

by xylene-induced ear oedema in mice and cotton pellet granuloma test in rats. Mentha piperita, Cinnamomum

zeylanicum, Apium graeolens,Eucalyptus camaldulentis, andRuta graeolens possess an anti-nociceptive effect against

both acetic acid-induced writhing and hot plate-induced thermal stimulation. M. piperita, Jasminum officinale,

Commiphora molmol, andBeta ulgarispossess an anti-inflammatory effect against acute (xylene-induced ear oedema)

and chronic (cotton-pellet granuloma) inflammation. The anti-nociceptive and anti-inflammatory effects were dosedependent. These data affirm the traditional use of some of these plants for painful and inflammatory conditions.

1998 Elsevier Science Ireland Ltd. All rights reserved.

Keywords: Anti-nociceptive activity; Anti-inflammatory activity

1. Introduction

There are more than 49 plant families having

more than 120 plant species used in Jordanian

traditional medicine, especially among people who

have little or no access to medical assistance(Karim and Quraan, 1986; Al-Khalil, 1995). The

anti-inflammatory and/or the analgesic effects of

Apium graeolens, Beta ulgaris, Eucalyptus

camaldulentis, Jasminum officinale, Mentha piper-

ita, Ruta graeolens, Lactuca satia, Cinnamomum

zeylanicum, and Commiphora molmol have been

described in folk medicine (Al-Jarwani and Khal-ifeh, 1936; Jabour, 1983). Moreover, A. graeo-

lens, M. piperita, and C. molmol have been

traditionally used as anti-spasmodics, anti-* Corresponding author.

0378-8741/98/$19.00 1998 Elsevier Science Ireland Ltd. All rights reserved.

PII S 0 3 7 8 - 8 7 4 1 ( 9 7 ) 0 0 1 3 7 - 2


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A.H. Atta, A . Alkofahi/Journal of Ethnopharmacology 60 (1998) 117124118

Table 1

List of plants used in the screening for anti-nociceptive and anti-inflammatory activity

Part used Voucher specimen number Yield g/kgScientific name

Dried juice 89 860A. era L. (Liliaceae)

3462A. graeolens L. (Umbelliferae) Seed

9655B. ulgaris L. (Chenopodiaceae) Root

Bark 11C. zeylanicum L. (Lauraceae) 50

114 168Oleo-gum resinC. molmol Engl. (Burseraceae)

Leaf 46E. camaldulentis Dehn. (Myrtaceae) 118

132 80J. officinale L. (Oleaceae) Flower

Seed 58L. satia L. (Compositeae) 104

Leaf 33M. piperita L. (Labiatae) 112

52 38O. syriaca (L.) Rafin (Labiatae) Leaf

Leaf 94R. graeolens L. (Rutaceae) 130

rheumatics, and to relieve teeth and ear pains,

skin diseases and headache (Kotb, 1985). In addi-

tion, these plants have been used for gastrointesti-

nal, respiratory, and reproductive disorders.Moreover, some of them are used as germicides

and vermifuges. The aim of this work is to eluci-

date the anti-nociceptive and anti-inflammatory

effects of 11 Jordanian plants (Table 1) of known

traditional use in relieving pain and inflammation.

2. Materials and methods

2.1. Plant materials

Extracts were prepared from 11 plants com-

monly used in Jordan as analgesics and/or anti-

inflammatory. Most of these plants grow locally.

Samples were obtained either as dried plants from

herbal stores or collected from the wild. The

taxonomic identity of the plants was confirmed by

Professor A. El-Oqlah, Department of Biological

Sciences, Yarmouk University, Irbid, Jordan. A

voucher specimen of each species studied has been

deposited at the Department of Medicinal Chem-

istry and Pharmacognosy, Faculty of Pharmacy,

Jordan University of Science and Technology,

Irbid, Jordan.

2.2. Preparation of plant extract

Each dehydrated plant was ground to a fine

texture and 100 g of the dried plant were repeat-

edly extracted with 80% ethanol. The ethanol

extracts were concentrated under vaccum and

weighed and the residue was used in the experi-ments. The dried plant extracts were freshly dis-

solved or suspended in distilled water just before

administration.

2.3. Acetic acid writhing in mice

The writhing test was performed as described

by Koster et al. (1959). Groups of ten Swiss mice

(five males and five females) were fasted over

night prior to dosing, but had free access to

water. After 30 min of receiving an oral dose of

the extract, each mouse was given intraperitonealy0.7% aquous solution of acetic acid (10 ml/kg b.

wt.) and then placed in an observation box. The

number of writhes were counted for 20 min after

acetic acid injection. The number of writhes in

each treated group was compared to that of a

control saline-treated group.

2.4. Hot-plate test in mice

The method described by Eddy and Leimback

(1953) was applied. Groups of ten mice each were

used. Mice were placed in a 2-l glass beakerplaced on a hot plate maintained at 55C. Latency

to exhibit the nociceptive response such as licking

paws or jumping was determined before and 15,


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A.H. Atta, A . Alkofahi/Journal of Ethnopharmacology 60 (1998) 117124 119

Table 2

Effect of ethanolic extracts on acetic acid-induced writhes in mice

200 mg/kg 400 mg/kgTreatment groups

Mean+S.D. Protection (%) Mean+S.D. Protection (%)

40.2+10.049.6+12.2Normal saline (10)

38 22.4+3.1***M. piperita (10) 30.2+9.5** 44

21 35.8+

6.0O. syriaca (10) 39.4+

5.3 11

18.8+3.8***48 5325.0+4.3***C. zeylanicum (8)

21 27.5+6.0**E. camaldulentis (8)a 39.2+10* 32

23.0+6.6*** 43A. graeolens (10) 29.6+10.2*** 40

48 20.6+4.6***J. officinale (9) 26.0+5.2*** 49

0 40.4+9.2 063.6+9.4B. ulgaris (10)

24 30.3+4.2**C. molmol (10) 37.5+11.5* 25

24 28.4+6.1**A. era (10) 37.6+10.3* 29

42.0+8.40 049.4+7.5L. satia (10)

32 27.0+4.1***R. graeolens (10)a 3333.8+7.0**

( ) Number of animals.a A dose of 100 and 200 mg/Kg b. wt.

*P0.05, **P0.01, ***P0.001 compared to control.

30, 45, 60, and 75 min after intraperitoneal ad-

ministration of the extract. A cut-off time of 60 s

was selected to avoid tissue damage.

2.5. Xylene-induced ear oedema in mice

Male Swiss mice were divided into groups of

ten mice each. After 30 min of the i.p. injection of

the extract, xylene (0.03 ml) was applied to the

anterior and posterior surfaces of the right ear.

Mice were sacrificed 2 h after xylene application

and both ears were removed. Circular sections of

both treated and untreated ears were taken using

a 7 mm diameter cork borer and weighed. The

difference in weight between left untreated ear

sections and right treated ear section was calcu-

lated (Tang et al., 1984).

2.6. Cotton pellet granuloma in rats

Sterilized cotton pellets of 20 mg weight each,

were impregnated with 0.4 ml of ampicillin

aquous solution. Pellets were implanted subcuta-neously in the groin region of Wistar rats, one on

each side. The drug was given orally by stomach

tube once daily for 6 days starting with the day of

implantation. On day 7, the rats were killed and

the pellets were removed, dried at 70C for 24 h.

The weight of granuloma and adrenal gland was

recorded and compared to that of saline treated

control (Schiatti et al., 1986). All plant extracts

were given in doses of 200 and 400 mg/kg body

weight except E. camaldulentis and R. graeolens

which were given in doses of 100 and 200 mg/kg

based on pilot experiments and appearance of

some toxic manifestations after larger doses.

2.7. Statistical analysis

The data were presented as meanS.D. Sig-

nificance between control and treated groups was

tested by ANOVA.

3. Results

Ethanolic extract ofC. zeylanicum, J. officinale,M. piperita, A. graeolens, and R. graeolens sig-

nificantly (P0.01 and P0.001) reduced thenumber of acetic acid-induced writhes in mice

with a protection percent ranging from 32 to 48

and from 33 to 53 after administration of 200 and


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Table 3

Effect of ethanolic extracts on time to response of mice to heat stimulation in the hot-plate test

Treatment Doses mg/kg Time to response (s)

groups

45 min 60 min0 min 15 min 30 min 75 min

Normal saline

7.48+

2.4 7.34+

1.9 8.42+

3.8(20) 8.34+

3.25.10 +

1.1 5.66+

1.5

M. piperita

7.72+1.1 8.42+0.4(6) 200 7.9+2.6 4.7+0.2 5.76+1.4 8.28+0.5

10.63+ 3.7** 15.52+6.1***(10) 400 6.1+2.3 7.15+2.1 7.10 +0.5 8.04+2.3

O. syriaca

8.1+0.4 8.3+0.6 7.86+0.4(7) 200 8.2+0.56.2+1.8 6.4+0.8

13.2 +6.24**10.88+1.1*(10) 5.93+0.9400 8.96+2.2*5.6+1.5 5.19+1.4

C. zeylanicum

8.7+0.4 7.0+0.9(5) 200 6.56+0.7 6.0+1.4 8.82+0.9 7.58+0.4

11.55+3.0*** 10.59+0.9*(10) 10.7+1.9*400 9.82+3.6**5.22+1.6 7.9+1.7***

E. camaldulen-

tis

6.4+

1.86.16+

1.5 6.9+

1.8(5) 6.1+

1.08100 7.0+

2.8 5.8+

1.211.51+ 4.4*** 13.4+5.3**(10) 200 5.18+0.5 14.3+3.8*** 17.4+6.9*** 12.8+5.7*

A. graeolens

9.5+1.98.0+2.5 10.1+2.9(5) 7.9+1.8200 5.01+1.0 6.34+1.5**

12.1+3.1**11.2+2.1*(10) 400 5.87+2.0 9.58+4.3*** 9.4+2.2* 10.6+2.3***

J. officinale

6.62+0.9 7.7+0.6(5) 200 7.02+2.5 4.92+1.5 6.9+2.9 6.98+1.2

7.8+1.9 8.08 +2.01(10) 400 4.81+2.4 6.72+2.1 6.28 +3.3 7.88 +3.9

B. ulgaris

46+ 1.2 6.12+0.6 6.9+1.3 7.42+0.9(5) 200 6.3+2.7 4.84+1.4 7

8.81+4.47.27+3.9(10) 7.71+3.2400 6.9+3.14.09+1.8 6.02 +1.3

C. molmol

6.9+0.9 8.16+0.6(5) 200 5.64+0.9 6.44+3.4 7.9+3.0 8.68+0.2

8.01+

2.66.91+

2.66.01+

0.7(10) 400 6.15+

1.56.0+

1.8 5.13+

1.3

A. era

7.5+0.87.6+0.67.3+0.5(5) 6.6+1.2200 6.4+0.6 5.6+0.9

6.64+2.1 6.33+2.6(10) 400 5.03+1.6 6.18+1.4 8.24+3.9 8.8+2.2

L. satia

6.78+2.47.3+1.0 5.8+2.7(5) 6.2+0.3200 6.38+0.4 6.44+0.6

7.7+2.66.89+1.4(10) 400 4.52+1.2 6.7+1.9 7.4+2.0 8.85+1.8

R. graeolens

6.99+2.6 8.9+1.36.1+2.5(5) 6.2+2.6100 6.95+2.3 7.9+2.7*

8.89+0.8* 7.68+1.4 6.49+0.6(10) 8.08+2.4200 4.07+1.8 7.38+2.3**

( ) Number of animals.

*P0.05, **P0.01, ***P0.001 compared to control. Mean+S.D.

400 mg/kg, respectively (Table 2). Ethanolic ex-

tract of E. camaldulentis, C. molmol, and Aloe

era also decreased the number of acetic acid-in-

duced writhes but to a less extent (P0.05 and

P0.01 for the low and high dose, respectively).

The ethanolic extract of O. syriaca, B. ulgaris,


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Table 4

Effect of ethanolic extracts on xylene-induced ear swelling in mice

200 mg/kg 400 mg/kg

Weight (mg) Inhibition (%) Weight (mg) Inhibition (%)

3.4+1.44.3+1.9Normal saline

1.7+0.8* 50M. piperita 2.2+1.2* 49

47 2.1+

0.8O. syriaca 2.3+

1.6 38

3.3+1.10 34.9+1.2C. zeylanicum

3.0+1.0 12E. camaldulentisa 3.9+2.7 9

63.2+1.0163.6+0.9A. graeolens

0 1.6+0.3*J. officinale 5.0+6 4.2 53

0 1.7+0.8*B. ulgaris 5.2+6.2 50

501.7+0.5*352.8 +2.1C. molmol

5.0+0.7 0 1.0+0.5** 71A. era

0 2.8+0.8L. satia 5.3+0.23 18

3.4+2.00 05.7+3.0R. graeolensa

a A dose of 100 and 200 mg/kg b. wt.

*P0.05, **P0.01. MeanS.D., n=5.

andL.satia in an oral dose of 200 or 400 mg/kg,showed no significant effect on the number of

acetic acid-induced writhes.

Ethanoilic extract ofC. zeylanicum, E. camald-

ulentis, and A. graeolens produced a highly sig-

nificant (P0.001) increase in the latency to

response of mice to hot plate thermal stimulation.

This effect starts 15 min and persisted for at least

75 min after administration of the plant extract

(Table 3). Mild or no effect was observed by the

small dose. Mild anti-nociceptive effect has also

been observed after administration ofM. piperita,

O. syriaca, and R. graeolens. However, theireffect was either temporary as in R. graeolensor

delayed in onset (45 min) as in M. piperita and O.

syriaca. On the other hand the ethanolic extracts

of J. officinale, B. ulgaris, C. molmol, A. era,

and L. satia showed no anti-nociceptive effect.

Ethanolic extract of A. era in a dose of 400

mg/kg significantly (P0.01) reduced the weight

of xylene-induced ear oedema in mice with a

calculated inhibition of 71%, while the smaller

dose produced no significant effect (Table 4). M.

piperite (both doses), J. officinale, B. ulgaris, and

C.molmolalso significantly (P0.05) reduced thesize of ear oedema with an inhibition percent

ranging from 50 to 53 when used at the high dose

only. O. syriaca, C. zeylanicum, E. camaldulentis,

A.graeolens, L. satia, andR. graeolensshowedno significant effect on xylene-induced oedema in

mice ears.

Ethanolic extract of A. graeolens, B. ulgaris,O.syriaca,C. zeylanicum, J. officinale,C. molmol,

and L. satia in a dose of 400 mg/kg and R.graeolens in a dose of 200 mg/kg significantly

reduced the weight of cotton granuloma in rats

(Table 5). Non of these extracts except A. graeo-lens (P0.05) was effective by the smaller dose.

The most effective extracts was that ofA. graeo-lens and B. ulgaris (P0.001). E. camaldulentis

and A. era have no effect at any of the useddoses. None of the tested plant extracts in the

used doses affect the weight of adrenal glands in

rats.

4. Discussion and conclusion

In the present study, two animal models for

investigation of the anti-nociceptive and anti-infl-

ammatory effects of the selected plants were used.

The hot plate thermal stimulation and the acetic

acid induced writhes in mice were selected toinvestigate the central and peripheral anti-noci-

ceptive effects. Xylene-induced ear swelling in

mice and the cotton pellet granuloma in rats were


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Table 5

Effect of ethanolic extracts on the weight of granuloma and adrenal glands in rats

Doses (mg/kg) Weight of granuloma (mg)Treatment Weight of adrenal (mg)

Normal control

0.107+0.030 0.046+0.030(8)

0.015+0.011(5) 0.059+0.017

M. piperita0.086+0.017 0.010+0.003(5) 200

400 0.045+0.009**(6) 0.066+0.009

O. syriaca

200 0.076+0.038 0.006+0.004(6)

0.036+0.002(6) 0.054+0.006**400

C. zeylanicum

200 0.057+0.012(6) 0.013+0.007

(6) 0.037+0.0050.075+0.007*400

E. camaldulentis

0.009+0.0040.056+0.021(5) 100

0.060+0.001 0.032+0.020(5) 200

A. graeolens

200 0.037+

0.012*(5) 0.020+

0.0160.046+0.0130.061+0.001***(9) 400

Jasminum officinale

0.017+0.0060.076+0.017(5) 200

400 0.069+0.014*(6) 0.087+0.010

Beta ulgaris

0.009+0.0030.054+0.001(6) 200

400 0.059+0.011***(8) 0.027+0.006

Commiphora molmol

0.060+0.011 0.012+0.003(5) 200

0.071+0.016* 0.024+0.008(5) 400

Aloe era

200 0.071+0.032(5) 0.010+0.003

400 0.083+0.005 0.015+0.001(5)

L. satia

200 0.073+0.024(5) 0.001+0.003

400 0.060+0.014*(5) 0.037+0.002

R. graeolens

0.016+0.0010.080+0.034(6) 100

200 0.047+0.001*(5) 0.029+0.004

( )Number of animals.

*P0.05, **P0.01, ***P0.001. MeanS.D.

selected to represent models of acute (exudative

phase) and chronic (the poliferative phase) inflam-mation respectively.

The present results show that M. piperita, C.

zeylanicum, E. camaldulentis, A. graeolens, and

R. graeolens induced a dose-dependent analgesic

protective effect against both thermal stimuli andthe writhing syndrome indicating central and pe-

ripheral effects. The increased latency to response

of mice to thermal stimuli only after administra-


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tion of the ethanolic extract of O. syriaca indi-

cates central analgesic effect only. On the other

hand, J. officinale, A. era, and C. molmol in-

duced an anti-nociceptive effect against acetic

acid-induced writhing only indicating the lack of

central anti-nociceptive effect of these three ex-

tracts.

The present results demonstrate that ethanolicextracts of M. piperita (dose dependent), J.officinalis, B. ulgaris, and C. molmol(large dose

only) possess a dose-dependent anti-inflamma-

tory effect against both acute (exudative) and

chronic (proliferative) inflammation. The activity

of ethanolic extract of A. era against acute infl-

ammation (xylene-induced ear oedema) only in-

dicates possible anti-phlogestic but not

anti-proliferative effect. O. syriaca, C. zey-lanicum, A. graeolens, L. satia, and R. graeo-lens showed an anti-inflammatory effect only

against chronic inflammation induced by cottonpellet granuloma indicating anti-proliferative ef-

fect. E. camaldulentis ethanolic extract in the

used dosage and route of administration showed

no anti-inflammatory effect.

All the tested extracts, except E. camaldulentis,

have variable degrees of both anti-nociceptive

and anti-inflammatory effect. Such an association

is well-known for various non-stroidal autin-infl-

ammatory (NSAI) compounds especially salicy-

late by-products (Reuse, 1978; Beuoist and

Misse, 1979; Famaey, 1983; Gyires et al., 1985).M. piperita, C. zeylanicum, A. graeolens, E.camaldulentis, and R. graeolens have both anti-

nociceptive and anti-inflammatory effects. These

five plant extracts contain common active princi-

ples; volatile oils, resins, and flavenoids (Kotb,

1985). Volatile oils, resins and flavenoids isolated

from other plant extracts have been proved to

posses analgesic and/or anti-inflammatory effect

(Duke, 1992). Therefore, it could be suggested

that the anti-nociceptive and anti-inflammatory

effects of the tested plant extracts may be due to

their content of volatile oils, flavenoids and

resins. From these results, it can be affirmed that

the traditional indication of some of these plantextracts for inflammation and pain associated

with sprains, bruises, wounds, spasmodic colics,

and rheumatic arthritis (Al-Khalil, 1995; Bajpai

and Sant, 1995; Sudarsanam et al., 1995). Fur-

ther investigations are necessary to elucidate the

exact mechanism of the anti-nociceptive and

anti-inflammatory activity of these extracts.

Acknowledgements

The authors would like to thank Jihan Abu-

Zaher for her technical assistance and the Dean-

ship of the Scientific Research in the University

of Science and Technology for financial support

(Grant No. 44/95).

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Atta 1998- Plantas Antiinflamatorias