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Fitoterapia 78 (2007) 365 369 www.elsevier.

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Analgesic and antipyretic effects of Capparis zeylanica leaves


B.V. Ghule a,, G. Murugananthan b , P.G. Yeole a
a

Institute of Pharmaceutical Education and Research, Borgaon (Meghe), Wardha 442 001, MS, India b J. S. S. College of Pharmacy, SS Nagar, Mysore 570 015, Karnatak, India Received 13 February 2005; accepted 4 February 2007 Available online 11 April 2007

Abstract The ethanol and water extracts of Capparis zeylanica leaves showed dose-dependent and significant (P b 0.05) increases in pain threshold in tail-immersion test. Moreover, both the extracts (100200 mg/kg) exhibited a dose-dependent inhibition of writhing and also showed a significant (P b 0.001) inhibition of both phases of the formalin pain test. The water extract (200 mg/kg) significantly (P b 0.01) reversed yeast-induced fever. Preliminary phytochemical screening of the extracts showed the presence of alkaloids, flavonoids, saponins glycosides, terpenoids, tannins, proteins and carbohydrates. 2007 Elsevier B.V. All rights reserved.
Keywords: Capparis zeylanica; Analgesic activity; Antipyretic activity

1. Introduction Capparis zeylanica is commonly known as Indian caper; a climbing shrub found throughout India and has been used as a Rasayana drug in the traditional Ayurvedic system of medicine. In Northern India, the leaves are widely used as counter-irritant, febrifuge and as a cataplasm in swellings, boils and piles [1,2]. We found no relevant literature substantiating the uses indicated. Phytochemical screening of the plant has shown the presence of fatty acids [3] and flavonoids [4] in the leaves since no data are till now reporting on the claimed activities. The purpose of the present study was to evaluate the analgesic effect of the ethanolic and water extracts using different acute and chronic models of pain in mice and rats and also to evaluate their antipyretic effects in brewer's yeast-induced pyrexia in rats. 2. Experimental 2.1. Plant Fresh leaves of the C. zeylanica L. (Capparidaceae), collected at the flowering stage in December from Nagpur District, Maharashtra State, India were authenticated by the Authority of Botany Department, Nagpur University, Nagpur, India. A voucher specimen (no. CSV/B-1 (12), 2004) is deposited in the departmental herbarium.
Corresponding author. Tel.: +91 7152 240284; fax: +91 7152 241684. E-mail address: dr_yeole@rediffmail.com (B.V. Ghule). 0367-326X/$ - see front matter 2007 Elsevier B.V. All rights reserved. doi:10.1016/j.fitote.2007.02.003

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2.2. Extraction Air-dried, powdered leaves were Soxhlet extracted with petroleum ether and EtOH and the marc was macerated giving a water extract. The extracts evaporated in vacuo gave petroleum ether, EtOH and water extracts (yields 6.8 %, 12.3 % and 15.5 % w/w, respectively). Moreover, the extracts were subjected to preliminary phytochemical screening for the detection of various plant constituents [5]. 2.3. Animals Swiss mice (1820 g) and Wistar rats (150200 g) of either sex kept at the Laboratory Animal Center of the Institute of Pharmaceutical Education and Research, Wardha, India were used. The animals were housed under standard environmental conditions and had free access to standard pellet diet (Goldmohar brand, Lipton India Ltd.) and water ad libitum. 2.4. Acute toxicity studies The method described by Lorke et al. [6] was employed in the determination of the LD50. Mice were separated into two sets of five groups of animals, each consisting of five male and female mice (N = 5). They were fasted overnight and then were administered with the EtOH and water extracts both orally and intraperitoneally at the following doses; 1, 10, 100, 1000, and 2000 mg/kg. Animals were observed for 24 h after treatment and the final LD50 value was calculated as the square root of the product of the lowest lethal dose and highest non-lethal dose. 2.5. Mouse writhing assay The method of Koster et al. [7] was used. The EtOH and water extracts (100200 mg/kg, oral) or acetylsalicylic acid (100 mg/kg, s.c.) was administered to mice 60 and 30 min, respectively, before intraperitoneal injection of acetic acid (0.6%, v/v in saline, 10 ml/kg). 10% v/v propyleneglycol was used as the control. The number of writhes was counted for 15 min. 2.6. Tail-immersion test Mice were divided into six groups each containing five animals. The lower 5 cm portion of the tail was immersed in a beaker of water maintained at 55 0.5 C [8]. The time in s for tail withdrawal from the water was taken as the reaction time, with a cut-off time of immersion set at 10 s. The reaction time was measured 1 h before and 1 h after oral administration of EtOH and water extracts (100200 mg/kg) or 10% v/v propylene glycol (10 ml/kg). Morphine (10 mg/kg) was administered subcutaneously, 30 min before the test. 2.7. Formalin test The method used was similar to that described previously by Shibata et al. [9]. Twenty microliters of 1% v/v formalin was injected subcutaneously into the right hind paw of mice. The time in s spent in licking and biting
Table 1 Effects of the C. zeylanica leaves EtOH and water extracts on acetic acid-induced writhing in mice Material Control EtOH extract Water extract Acetylsalicylate Dose (mg/kg) 100 200 100 200 100 No. of writhes z( 15 min) 37.0 0.70 32.6 0.54 27.6 0.89b 27.6 0.54b 25.2 0.83b 11.8 0.83a Inhibition (%) 11.90 25.41 25.41 31.90 68.11

Values are expressed as mean S.D. aP b 0.01, bP b 0.05 significantly different from control. Student's t-test. N = 6.

B.V. Ghule et al. / Fitoterapia 78 (2007) 365369 Table 2 Effects of C. zeylanica leaves EtOH and water extracts on tail-immersion test in mice Material Control EtOH extract Water extract Morphine Dose (mg/kg) 100 200 100 200 10 Reaction time (s) 2.6 0.54 3.4 0.54 4.8 0.83b 3.8 0.83a 5.2 0.44b 7.0 0.70b

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Increase (%) 30.76 45.83 46.15 100.00 169.00

Values are mean S.D. aP b 0.01, bP b 0.001; significantly different from control; Paired t-test. N = 6.

responses of the injected paw was taken as an indicator of pain response. Responses were measured for 5 min after formalin injection (first phase) and 1530 min after formalin injection (second phase). The EtOH and water extracts (100200 mg/kg, oral) and acetylsalicylic acid (100 mg/kg, s.c.) were administered 60 and 30 min, respectively, before formalin injection. Control animals received 10% v/v propylene glycol (10 ml/kg). 2.8. Antipyretic activity Antipyretic activity of drug was measured by slightly modifying the method described by Adams et al. [10]. Rats were fasted overnight with water ad libitum before the experiments. Pyrexia was induced by subcutaneously injecting 20% w/v brewer's yeast suspension (10 ml/kg) into the animal's dorsum region. Nineteen h after the injection, the rectal temperature of each rat was measured using a thermometer. Only rats that showed an increase in temperature of at least 0.7 C were employed for the experiments. The EtOH and water extracts (100200 mg/kg) or 10% v/v propylene glycol solution (10 ml/kg) was administered orally and the temperature was measured at 0, 1, 2 and 3 h after drug administration. 2.9. Statistical analysis All data were expressed as mean S.D. and analyzed statistically by using Student's t-test and paired t-test. A difference was considered significant at P b 0.05. 3. Results and discussion 3.1. Acute toxicity studies The LD50 in mice for the EtOH and water extracts was estimated to be 1624.5 42.6, 1428 62.8 mg/kg, p.o. and 324.6 12.4, 310.4 11.8 mg/kg i.p., respectively. During observation the animals exhibited decreased mobility, respiratory distress (gasping) with eventual immobility but without convulsions or loss of righting reflex prior to death.

Table 3 Effects of the C. zeylanica leaves EtOH and water extracts on formalin-induced pain in mice Material Control EtOH extract Water extract Morphine Dose (mg/kg) 100 200 100 200 10 05 min 89.0 4.18 84.4 1.09 70.0 3.80 69.4 3.57a 65.2 3.34b 57.8 2.95b Inhibition (%) 0.04 21.35 22.03 26.75 35.06 1530 min 83.4 3.20 48.2 3.19b 38.6 2.60b 31.4 2.60b 24.2 2.16b 12.2 1.30b Inhibition (%) 42.21 53.72 62.36 70.99 85.38

Values are mean S.D. aP b 0.01, bP b 0.001; Significantly different from control; Paired t-test. N = 6.

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Table 4 Effects of the C. zeylanica leaves EtOH and water extracts on brewer's yeast-induced pyrexia in rats Material Dose (mg/ kg) 100 200 100 200 100 Average rectal temperature (C) 0h 31.05 0.44 32.25 0.27 32.25 0.40 30.75 0.75 30.31 0.28 30.50 0.08b 1h 36.58 0.58 35.91 0.37 37.08 0.58 36.16 0.81 35.16 0.81a 33.26 0.32b 2h 38.20 0.34 38.06 0.72 39.91 0.37 37.60 0.24 36.83 0.75b 34.65 0.48b 3h 39.50 0.91 39.75 0.52 40.05 0.59 38.50 0.44 37.16 0.68b 33.41 0.25b

Control EtOH extract Water extract Acetylsalicylic

Values are mean S.D. aP b 0.05, bP b 0.01; Significantly different from control; Paired t-test. (N = 6).

3.2. Analgesic effects In the mouse writhing assay, EtOH and water extracts caused a significant and dose-dependent inhibition of the control writhes (Table 1). The inhibition produced by the highest dose (200 mg/kg) of the extracts was significantly (P b 0.01) lower than that by acetylsalicylic acid (100 mg/kg). The effect of EtOH and water extracts on tail-immersion tests are shown in Table 2. These extracts showed a dose-dependent inhibition of pain with the water extract being more active than the EtOH one. There was a significant, dose-dependent inhibition of both phases of the formalin-induced pain response in mice, with a more potent effect on the second than the first phase (Table 3). 3.3. Antipyretic effects The EtOH extract of C. zeylanica did not show significant (P b 0.05) effect on pyrexia induced by yeast while the water extract (200 mg/kg) significantly (P b 0.01) reversed yeast-induced fever (Table 4). 4. Discussion Several tests (acute and chronic) were employed in evaluating the analgesic effect of the EtOH and water extracts of C. zeylanica. It is necessary to apply tests which differ with respect to stimulus quality, intensity and duration, to obtain as complete a picture as possible of the analgesic properties of a substance using behavioral nociceptive tests [11]. The results obtained indicate that the extracts possess a moderate dose-dependent analgesic effect on the various pain models used. A potent inhibitory effect was exerted by both the extracts on the mouse writhing assay (a test useful for evaluating mild analgesic non-steroidal anti-inflammatory agents). This suggests that the analgesic effect of the extract may be peripherally mediated. The extracts also had a significant effect in the tail-immersion tests. Centrally acting analgesic drugs elevate pain threshold of animals towards heat and pressure. The effect of the extract on this pain models indicates that it might be centrally acting. The extracts inhibited both phases of the formalin-induced pain with a more potent effect on the second than the first phase. The formalin pain test is very useful for evaluating the mechanism of pain and analgesia. Drugs which act mainly centrally, such as narcotic analgesics, inhibits both phases of pain in this model while peripherally acting drugs, such as acetylsalicylic acid or indomethacin, only inhibit the late phase [12]. The EtOH extract of C. zeylanica did not show any significant effect on brewer's yeast-induced fever in rat; however water extract (200 mg/kg) significantly reversed yeast-induced pyrexia. The results obtained in this study indicate that the extracts possess mild analgesic properties. Even if further studies are needed this seems to provide a rationale for the use of this plant in pain and inflammatory disorders. Acknowledgements We owe our thanks to the authority of Botany Department, Nagpur University, Nagpur for the authentication of plant specimen. The facilities provided by the Institute of Pharmaceutical Education and Research, Wardha during this course of study are gratefully acknowledged.

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