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Mandade R J et al.

/ Pharmacie Globale (IJCP) 2010, 4 (07)

Available online at www.pharmacie-globale.info

ISSN 0976-8157

Research Article

PHARMACIE GLOBALE INTERNATIONAL JOURNAL OF COMPREHENSIVE PHARMACY

ANALGESIC AND ANTI-INFLAMMATORY ACTIVITIES OF Hedera helix LEAF EXTRACT


Rajesh J. Mandade1*, Avijit Choudhuri2, Vinod Mashirkar3 and Dinesh Sakarkar4
2Department

of Pharmacology, S.N. Institute of Pharmacy, Pusad, Maharashtra, India. of Pharmacology, Shree Dhanvantary College of pharmacy, Kim Surat, Gujarat, India. 3Department of Pharmacognosy, S.N. Institute of Pharmacy, Pusad, Maharashtra, India. 4Principal, S.N. Institute of Pharmacy, Pusad, Maharashtra, India.

1Department

Received: 1 October 2010; Revised: 22 October 2010; Accepted: 27 October 2010; Available online: 1 November 2010

ABSTRACT

The methanolic extract of the leaves of Hedera helix L. (Araliaceae) which contains mainly saponins and carbohydrate showed significant analgesic and anti-inflammatory activities (P<0.05) in the tail flick/ hot-plate test and egg albumen-induced rat paw oedema tests that were comparable to the test drugs (morphine 20mg/kg and indomethacin 50mg/kg respectively). These findings suggest that the methanol leaf extract of Hedera helix L. possess analgesic (which might have been peripherally mediated) and anti-inflammatory activities; and may lend credence to the ethnomedical claim of the use of the plant for the management of pain and inflammatory conditions. Keywords: Hedera helix, Analgesic activity, Anti-inflammatory activities, Egg albumin.

INTRODUCTION

Hedera helix L. (Araliaceae) is an evergreen climbing plant, growing to 2030 m high where suitable surfaces are available, and also growing as ground cover where there are no vertical surfaces. It ranges from Ireland northeast to southern Scandinavia, south to Spain, and east to Ukraine and northern Turkey and also in Asia.1 The closely related species Hedera canariensis and Hedera hibernica are also often treated as subspecies of H. helix, in traditional medicine; the leaves were used as analgesic and anti-inflammatory agents. The leaves and berries were taken orally as an expectorant to treat cough and bronchitis. The leaves can cause severe contact dermatitis in some people.2, 3 Three steroidal saponins have been isolated from the leaves of A. helix. This study was carried out to ascertain the traditional uses of the leaf of the plant as analgesic and anti-inflammatory agents.

apparatus for 24 hrs. The marc was then air-dried and extracted exhaustively with methanol for 2 days. The methanolic extract (ME) was concentrated in a rotary evaporator at reduced pressure to attain a dark brown residue. Phytochemical tests The extract (ME) was subjected to phytochemical tests to detect the various types of chemical constituents present using standard procedures.4 Pharmacological tests Animals Adult Swiss albino mice (20-25g) and rats (200-250g) of both sexes were used for the tests. The animals were obtained from the animal house of the Department of Pharmacology, S.N.Institute of Pharmacy, Pusad. They were housed in steel cages under standard conditions, fed with standard pellets and water ad libitum. Procedures for animal handling were consistent with international guidelines. Acute toxicity test The acute toxicity (LD50) of the extract (ME) was determined in mice by the method of Lorke (1983) using the oral route.5 Analgesic activity Tail-flick test: Mice (20-25g) of both sexes were fasted overnight before the study. A tail-flick analgesic meter Ugo Basile was used to measured response latencies as described by Santos et al.6 Five groups of six animals each were pretreated with 0.2ml normal saliva intraperitoneally to group one as negative control, morphine 20mg/kg subcutaneously to group two as positive control while the remaining three groups received 250, 500 and 1000mg/kg of ME orally respectively. Measurements of responses were taken at time zero, 30, Pharmacie Globale (IJCP), Vol. 01, Issue 04

MATERIALS AND METHODS


Plant material The leaves of Hedera helix was collected from botanical garden of SRM college of Phramcy, Chennai. The plant was authenticated at the Herbarium of The plant were authenticated by Prof. P. Jayaraman, Ph.D. Director plant anatomy research centre, Pharmacognosy institute, No. 411 street, sakthi nagar, west Tambram, Chennai. The roots were air-dried for several weeks and then powdered using mortar and pestle. Preparation of the extract The powdered leaves (500g) were defatted with petroleum ether (60-80C) exhaustively in a soxhlet extractor
*Corresponding Author: Rajesh J Mandade Lecturer, department of Pharmacology, S.N. Institute of Pharmacy, Pusad, (M.S.), India. Contact no: +91-8888673088, Email: raj_mandade@rediffmail.com

Mandade R J et al. / Pharmacie Globale (IJCP) 2010, 4 (07)

60, 90, and 120 minutes after treatment of the animals with drugs and extract by application of pressure from the analgesiometer onto their tail (1cm from the tip of the tail). Hot-plate test: Mice (20-25g) of both sexes were fasted overnight before the study. Hot-pate was used to measure response latencies according to the methods of Turner,7 and Guzman et al.8 with minor modifications. In this study, the hot-plate was maintained at 55 1OC and the animals were individually placed on the heated surface. The time in seconds between placement and shaking, paw licking and jumping off the plate was recorded as response latency. Five groups of six animals each were pre-treated with 0.2ml normal saline intraperitoneally to group one as negative control, morphine 20mg/kg intraperitoneally to group two as positive control while the remaining three groups received 250, 500 and 1000mg/kg of ME orally respectively.9 Measurements were taken at zero, 30, 60, 90 and 120 minutes after the treatment of animals.

inhibition was calculated according to the formula of Barbera et al.11

Where = mean paw volume at start of experiment = mean paw volume at time 30, 60, 90 and 120 minutes (A) = % Oedema increase Statistical analysis Results were expressed as mean standard error of mean (SEM). Students t-test was used to analyze the data. P<0.05 was considered to be statistical significant.

RESULTS AND DISCUSSION

Anti-inflammatory activity Anti-inflammatory activity was carried out using the rat paw oedema test according to the method of Winter et al.10 And Guzman et al., (2001)8 with minor modifications. Rats (200-250g) of both sexes were divided into five groups of six animals each. 0.2ml of fresh undiluted egg albumen was injected into the right hind paw of the rats after pretreatment with 0.2ml of normal saline intraperitoneally to group one as negative control, indomethacin 50mg/kg intraperitoneally to group two as positive control while the remaining three groups received 250, 500 and 1000mg/kg of ME orally respectively. Paw volumes were measured using Ugo Basile 7140 phethysometer immediately after injection of egg albumen and at 30, 60, 90 and 120 minutes after treatment. Percentage oedema Table 1. Effect of ME on response latencies in the tail-flick test mean response latencies SEM doses
Treatment groups Control Morphine ME ME ME Treatment groups Control Morphine ME ME ME Doses mg/kg 0 Min. -----20 250 500 1000 Doses mg/kg 0 -----20 250 500 1000 4.98 1.22 7.03 1.58 5.85 1.16 5.55 1.31 6.33 1.21 3..99 0.83 5.24 1.88 4.64 0.76 5..37 0.70 4.42 0.74 Mean response latencies SEM 30 Min. 60 Min. 5.49 0.78 17.434.25* 6.90 1.72 9.97 2.60* 11.250.08* 5.36 1.21 15.323.42* 10.872.32* 14.864.56* 15.335.28* 90 Min. 6.12 1.18 13.553.42* 14.203.33* 13.033.54* 14.833.61*

The extraction process yielded 16.33% of the methaolic extract. Phytochemical tests showed the presence of carbohydrates and saponins mainly with small quantity of flavonoids and tannins. The methanolic extract of the leaf of Hedera helix did not cause any mortality upto a dose of 5000 mg/kg orally and was thus considered to be none toxic.12 The mathanolic extract of the leaf of A. helix showed significant dose-dependent analgesic activity in the tail-flick and hot-plates tests (Tables 1 and 2). Analgesic activity of the extract increases with time while that of the test drug (morphine) decreases with time. The extract has shown central and peripheral analgesic activities since the results showed significant (p<0.05) analgesic activity of the extract at all the doses used in the tests. Agents such as histamine, serotonin and bradykinin are natural pain substances liberated by injured tissues which stimulate the nociceptive mechanism to elicit pain.13

120 Min. 6.42 1.16 10.953.20* 10.483.54* 11.982.32* 13.253.43*

Table 2. Effect of ME on Response Times in the Hot-Plate Test


Mean response latencies SEM 30 60 5.47 0.75 36.364.34* 9.20 2.59* 8.63 1.7* 19.5 2.74* 5.63 2.07 29.5 3.86* 14.17 2.3* 17.4 4.24* 24.1 4.05* 90 6.0 2.44 18.172.22* 22.508.00* 28.538.33* 30.5 3.83* 120 6.10 2.10 11.040.64* 12.003.84* 17.407.35* 21.335.35*

Analgesic agents are known to diminish the perception of that stimulate the nociceptive mechanism. The results of pain by depressing the nociceptive mechanism. The the anti-inflammatory effects of the methanolic extract of extract may possibly have acted as an analgesic agent by the leaf of A. helix on egg albumen-induced oedema in rats, antagonizing the activities of the natural pain substances hind paws are presented in Table 3. Table 3: Effects of ME on Egg Albumen Induced Rat Paw Oedema Test
Treatment groups Control Indomethacin ME ME Doses mg/kg 0 -----20 250 500 0.66 0.30 0.99 0.21 0.77 0.14 0.83 0.11 Mean Paw Volume SEM (Percentage Oedema Inhibition) 30 1.22 0.28 1.29 0.31 (25.23)* 1.30 0.26 (12.44) 1.32 0.22 (17.22) 1.31 0.4 (25.85)* 60 1.51 0.30 1.50 0.37 (27.50)* 1.58 0.21 (22.40)* 1.46 0.18 (24.70)* 1.43 0.46 (35.68)* 90 2.00 0.34 1.66 0.36 (34.65)* 1.98 0.11 (25.44)* 1.55 0.17 (44.50)* 1.69 0.62 (38.20)* 120 2.10 1.96 2.10 1.89 1.99

ME 1000 0.89 0.21 * = Statistical significant (P<0.05) compared to control

The extract at a dose of 500mg/kg showed the highest percentage inhibition of oedema (44-55%) at 90 mins

which was comparable to that of indomethacin 50mg/kg (34.65%) at the same time. Saponins isolated from about Pharmacie Globale (IJCP), Vol. 01, Issue 04

Mandade R J et al. / Pharmacie Globale (IJCP) 2010, 4 (07)

50 plants have been shown to possess anti-inflammatory activities against several experimental models of inflammations in mice and rats.14 The initial phase of oedema which develops in rats paw after the injection of oedemagenic agents which is known to last for 90 minutes is due to the release of histamine and serotonin. The last phase of the inflammation which last for 5 hrs is due to the release of prostaglandins (DiRosa et al. 1971).15 Egg albumen is an oedemagenic agent which produces inflammatory reactions through the release of serotonin and histamine. The resulting oedema has been shown to be inhibited by antihistaminic agents.16 The activity of the extract was observed to be more pronounced in the first

phase of the rat paw oedema (90 mins) thus suggesting that the extract may possess antihistaminic activity.

CONCLUSION

Based on the results of the present study it can be concluded that the methanolic extracts of the leaf of Hedera helix has potential dose-dependent analgesic and anti-inflammatory activities. Hence this study has confirmed the use of the plant in traditional medicine as a pain reliever in the treatment of headache, backache, as an aid in child birth and in the treatment of inflammatory diseases such as haemorrhoids, rheumatism and chronic gout. 10. Winter C A; Non Steroid Anti-inflammatory Agents. In: Progress in Drug Research, Vol 10. (ed.Jucker, E.) 1966; Pp 139-203. Birkhauser Verlag Basel and Stuttgart. 11. Berbera R J, Trovato A J and Requsa S; Analgesic and Anti- inflammatory Activity in Acute and Chronic Conditions of Trema guineense (Schum. et Thonn) Ficalhu and Trema micantha Blume Extracts in Rodents. Phytother. Res. 1992; 6. 144-146. 12. Dubois H P and Geilin g. E M H; Textbook of Toxicity. 1959; Pp 302-303 Oxford University Press, London, U.K. 13. Collier H O J; Analgesic. In: Evaluation of Drug Activities: Pharmacometric. (Ed. Laurence, D.R. and Bacharach, A.L.) Vol. 1. 1964; Pp 183-203. Academic Press, London and New York. 14. Lacaille-Dubois, M A and Wagner H; A Review of the Biological and Pharmacological Activities of Saponins. Phytomedicine 1996; 2 (4): 363-386 15. DiRosa M, Givoud J P and Wiloughby; Studies of the Mediators of Acute Inflammatory Responses Induced in Rats in Different Sites by Carragenan and Turpentine. J. Pathol. 1971; 104: 15-29. 16. Winter C A, Risky E A and Nuss G W; Carragenaninduced Edema in Hind Paw of the Rat as an Assay for Anti-inflammatory Drugs. Proc. Soc. Exp. Biol. Med. 1962; 111:544-547.

REFERENCES

1. Metcalfe D J; Biological Flora of the British Isles no. 268 Hedera helix L. Journal of Ecology 2005; 93: 632648. 2. Jhnke, H and Bjarnason, B; Contact dermatitis allergy to common ivy (Hedera helix L.). Ugeskr. Laeger 1994; 156 (25): 3778-3779. 3. Boyle, J and Harman R M H; Contact dermatitis to Hedera helix (Common Ivy). Contact Dermatitis 2006; 12 (2): 111112. 4. Evans W C Trease and Evans; Pharmacognosy. 14th edition. 1996; pp 293-321. W.B Sounders Company Ltd. U.K. 5. Lorke, D; A New Approach to Practical Acute Toxicity Testing. Arch. Toxicol. 1983; 54: 275-287. 6. Santos A R S, Filho V C, Yunes R A and Calixto S B; Further Studies on Antinoceptive Action of the Hydroalcoholic Extracts from Plants of the Genus phyllanthos. Pharm. Pharmacol. 1995; 47:66-71. 7. Turner R A; Screening Methods in Pharmacology. Vol. 1. 1975; Pp 105-108. Academic press, New York, U.S.A. 8. Guzman S, Gato A and Callega J M; Anti-inflammatory, Analgesic and Free Radial Scavenging Activities of the Marine Microalgea Chlorella stigmaphora and Phaeodactylum tricormutrum. Phytother. Res. 2001; 15: 224-230 9. Toma W, Graciosa J S; Hiruma-Lima Evaluation of the analgesic and antiedematogenic activities of Quassia amara bark extract. J Ethnopharmacol. 2003; 85: 19-23.

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