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Journal of Ethnopharmacology 134 (2011) 294–297

Contents lists available at ScienceDirect

Journal of Ethnopharmacology

journa l homepage: www.e lsev ier .com/ locate / je thpharm

astric antisecretory and antiulcer activities of Cedrus deodara (Roxb.) Loud. inistar rats

vadhesh Kumara, Vandana Singhb, Amrendra Kumar Chaudharya,∗

School of Pharmaceutical Sciences, Shobhit University, NH-58, Modipuram, Meerut 250110, UP, IndiaTranslam Institute of Pharmaceutical Education and Research, Meerut 250001, India

r t i c l e i n f o

rticle history:eceived 31 August 2010eceived in revised form0 November 2010ccepted 10 December 2010vailable online 21 December 2010

eywords:

a b s t r a c t

Ethnopharmacological relevance: Cedrus deodara (Roxb.) Loud. is used in Ayurvedic medicine to treat pepticulcer.Aim of the study: To evaluate the gastric antisecretory and antiulcer activity of Cedrus deodara.Materials and methods: The volatile oil extracted by steam distillation of Cedrus deodara wood was exam-ined for its gastric antisecretory and antiulcer effect in the pylorus-ligated rat model and ethanol inducedgastric lesions in rats.Results: The volatile oil showed significant antisecretory activity as evidenced by decreased gastric fluidvolume, total acidity, free acidity and increase in the pH of the gastric fluid in pylorus-ligated rats. Our

edrus deodara

ntiulcerylorusthanolabeprazole

studies also revealed that pretreatment with Cedrus deodara significantly reduced the number of ulcer,ulcer score and ulcer index in pylorus-ligated and ethanol treated rats. The antiulcer activity of Cedrusdeodara is further supported by histopathological study which showed protection of mucosal layer fromulceration and inflammation.

ndingjustif

Conclusion: The present fiand antiulcer effects and

. Introduction

The plants Cedrus deodara (Roxb.) Loud. belonging to the familyinaceae is a species of cedar, native to the Western Himalayasn Eastern Afghanistan, Northern Pakistan, North-Central India,outh Western Tibet and Western Nepal, occurring at 1500–3200 mltitude (Shah, 2006). The chemical constituents obtained fromifferent parts of the plant include wikstromal, matairesinol,ibenzylbutyrolactol (Rao et al., 2003; Singh et al., 2007),ergapten, isopimpinellin, lignans 1,4 diaryl butane, benzofura-oid neo lignan, isohemacholone, sesquiterpenes LIII: deodarone,tlantone (Shankaranarayan et al., 1977), deodarin, deodard-one, limonenecarboxylic acid, �-himacholone, �-himacholone,edrin (6-methyldihydromyricetin), taxifolin, cedeodarin (6-ethyltaxifolin), dihydromyricetin and cedrinoside (Agrawal et al.,

980). The wood oil of Cedrus deodara has been used sincencient days in Ayurvedic medical practice for the treatment oflcer (Hussain et al., 2006; Shah, 2006). However, there is not

nough scientific data to support the claims made in the ancientiterature. Recent in vivo and in vitro studies have indicated its anti-nflammatory, analgesic (Shinde et al., 1999a), anti-hyperglycemic,nti-spasmodic, antibacterial, insecticidal (Singh and Agarwal,

∗ Corresponding author. Tel.: +91 9450883086; fax: +91 121 2575724.E-mail address: amrendrapharma@gmail.com (A.K. Chaudhary).

378-8741/$ – see front matter © 2010 Elsevier Ireland Ltd. All rights reserved.oi:10.1016/j.jep.2010.12.019

s conclude that volatile oil of Cedrus deodara wood has potent antisecretoryy the traditional usage of this herb to treat peptic ulcers.

© 2010 Elsevier Ireland Ltd. All rights reserved.

1987), anti-apoptotic, immunomodulatory (Shinde et al., 1999b),anti-cancer (Singh et al., 2007) and molluscidal (Gupta et al., 1988)activity.

Although a number of anti-ulcer drugs such as antacid, H2receptor antagonist, proton pump inhibitor, cytoprotectives, andprostaglandin analogues are available for treatment of ulcer, allthese drugs have side effects and limitations.

The need for safer and effective antisecretory and anti-ulcerdrug and the lack of enough scientific data to support the claimsmade in ancient literature prompted the present study.

2. Materials and methods

2.1. Collection and authentication of plant material

The wood of Cedrus deodara, obtained from a commercial sup-plier and authenticated by Dr. Anjula Pandey, Principal Scientist,National Botanical Plant Genetic Research Institute (NBPGR), NewDelhi, India. A voucher specimen has been deposited at the NBPGRHerbarium (NHCP/NBPGR/2009-32 dated November 19, 2009).

2.2. Extraction of volatile oil

The plant material cut into small pieces, was subjected to hydrodistillation using Clevenger apparatus. The oil was separated fromaqueous layer, dried over anhydrous sodium sulphate and stored

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n an amber colored glass bottle in cool place (Shinde et al., 1999a;anda et al., 2008).

.3. Animals

Wistar rats of either sex (160–240 g) and Swiss albino mice25–30 g) were obtained from Indian Veterinary Research Institute,areilly, UP, India. The animals were housed in polypropylene cagender standard conditions (25 ± 2 ◦C, 12 h light and dark cycle) withree access to standard pellet feed (Ashirwad Industries, Mohali,unjab) and water ad libitum. All the experimental procedures androtocols involving animals were reviewed by the Institutional Ani-al Ethical Committee (Registration Number: 1279/ac/09/CPCSEA)

nd were in accordance with the guidelines of CPCSEA.

.4. Phytochemical testing

The volatile oil was subjected to preliminary phytochemicalcreening for the presence of terpenoid, phenolics, alcohol, alde-yde and ketone. The terpenoid was evaluated by using SalkowskiEdeoga et al., 2005) and Liebermann–Burchard’s test (Parekh andhanda, 2007). The total phenolic content in Cedrus deodara wasetermined by using bromine water and Liebermann test. Alco-ol content of Cedrus deodara was determined by Lucas test. Theldehyde was evaluated by Benedict’s solution, Fehling’s solutionnd Tollens test. The presence of ketone in Cedrus deodara wasetermined by using 2,4-dinitrophenylhydrazine test.

.5. Acute toxicity study

The Cedrus deodara was administered orally in dose of 50, 100,00, 300, 400 and 500 mg/kg to groups of mice (n = 6) and percent-ge mortality was noted 24 h later.

.6. Gastric secretion in pylorus-ligated rats

Gastric secretion content, pH, total acidity, free acidity, numberf ulcer, ulcer score and ulcer index were measured according tohe method of Shay et al. (1945). One hour after oral administrationf volatile oil of Cedrus deodara (50 and 100 mg/kg) or Rabeprazole20 mg/kg) or vehicle (10 ml/kg mixture of 3% (w/v) acacia and tra-acanth in distilled water), the animals were subjected to pylorusigation under thiopental sodium anaesthesia. The animals wereacrificed with an overdose of thiopental sodium after 4 h of pyloricigation. The abdomen was opened, cardiac end of the stomach

as dissected out and the contents were drained into a glass tube.he volume of the gastric juice was measured and centrifuged at000 rpm for 10 min. From the supernatant, aliquots (1 ml of each)ere taken for the determination of pH, total and free acidity. Total

cidity and free acidity were determined using titrimetry (Treasend Evans, 1992). The inner surface of free stomach was examinedor gastric lesions. The number of ulcers was counted. Ulcer scor-ng was done according to the method by Vogel and Vogel (1997)s given below.

The scores were: 0 = no ulcer, 1 = superficial ulcer, 2 = deep ulcer,= perforation.

Ulcer index was measured by using the following formula (Vogelnd Vogel, 1997)

I = UN + US + UP × 10−1

I is the ulcer index; UN is the average number of ulcers per animal;S is the average number of severity score; and UP is the percentagef animals with ulcers.

macology 134 (2011) 294–297 295

Percentage inhibition of ulceration was calculated as follows:

% Inhibition of ulceration = (Ulcer index control − Ulcer index test) × 100Ulcer index control

2.7. Gastric lesions induced by ethanol

Lesions were induced according to the method of Vogel andVogel (1997). Rats, fasted for 24 h but with free access to water wereused. One hour after the treatments (Cedrus deodara: 100 mg/kg orRabeprazole: 20 mg/kg or vehicle: 10 ml/kg mixture of 3% (w/v)acacia and tragacanth in distilled water), 90% ethanol (5 ml/kg)was administered orally. The animals were sacrificed under etheranaesthesia, 1 h after ethanol treatment. The stomach of each ani-mal was excised and opened along the greater curvature. Thenumbers of ulcer, ulcer score, ulcer index and % inhibition of ulcerwere determined.

2.8. Histopathological evaluation

The stomach samples from the pylorus ligated and ethanoltreated groups were preserved in 10% buffered formalin andprocessed for routine paraffin block preparation. Using a rotarymicrotome, sections of thickness of about 5 �m were cut andstained with haematoxylin and eosin. These were examined underthe microscope for histopathological changes such as degeneration,hemorrhage, edematous appearance, erosion and necrosis.

2.9. Statistical analysis

The results were expressed as mean ± standard deviation ofmean (SD). Analysis of variance (ANOVA) was applied to compareand analyse the data followed by Dunnett’s t-test.

3. Results

3.1. Yield of plant extract

The 1185 g wood powder of Cedrus deodara was taken for extrac-tion of volatile oil by water distillation method using Clevengerapparatus. Total 5.80 ml volatile oil was obtained. The yield ofvolatile oil of Cedrus deodara was found to be 0.47% (v/w).

3.2. Phytochemical testing of Cedrus deodara

Phytochemical testing showed that the volatile oil of Cedrusdeodara contains terpenoids, phenols, alcohol and ketone.

3.3. Acute toxicity

The mice treated with oral administration of 50 and 100 mg/kgof Cedrus deodara were normal. The animals which received 200,300, 400 mg/kg showed signs of depression. The animals treatedwith 500 mg/kg of Cedrus deodara showed 50% mortality. In thestudy, volatile oil of Cedrus deodara was administered in 50 and100 mg/kg dosage, which was determined as the most effectivedosage in a previous report (Shinde et al., 1999b).

3.4. Gastric secretion in pylorus-ligated rats

Gastric secretion measurements of pylorus-ligated rats showedthat Cedrus deodara significantly decreased the gastric content,pH, total and free acidity at doses of 50 mg/kg and 100 mg/kg.Rabeprazole (20 mg/kg), the reference compound used also showedsignificant reduction of all these secretory parameters (Table 1). The

296 A. Kumar et al. / Journal of Ethnopharmacology 134 (2011) 294–297

Table 1Effect of volatile oil of Cedrus deodara on gastric content, pH, total acidity and free acidity in pylorus ligation induced ulceration in rats.

Treatment Dose (kg−1) Gastric content (ml) pH of gastric content Total acidity (meq./l) Free acidity (meq./l)

Control 10 ml 2.32 ± 0.12 2.95 ± 0.23 4683.25 ± 211.40 3307.83 ± 328.15Cedrus deodara 50 mg 2.13 ± 0.06a 3.19 ± 0.15 4346.07 ± 192.42b 2844.07 ± 163.71b

Cedrus deodara 100 mg 1.92 ± 0.05b 5.62 ± 0.20c 2312.42 ± 165.33c 1714.96 ± 164.5c

Rabeprazole 20 mg 2.05 ± 0.12c 6.18 ± 0.27c 2180.90 ± 83.93c 1676.57 ± 253.82c

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is not involved in the formation of such lesions (Miller and Henagan,1984). Ethanol rapidly penetrates the gastric mucosa apparentlycausing cell and plasma membrane damage leading to increasedintra cellular membrane permeability to sodium and water. The

alues are represented as mean ± SD. Statistical analysis was done by one-way ANOa P < 0.05 as compared to control.b P < 0.01 as compared to control.c P < 0.001 as compared to control.

igher doses of Cedrus deodara (100 mg/kg) showed similar level ofction as that of reference compound.

.5. Gastric lesions in pylorus-ligated rats

Gastric lesion measurements of pylorus-ligated rats showedhat Cedrus deodara significantly decreased the number of ulcer,lcer score and ulcer index at the doses of 50 mg/kg and 100 mg/kg.abeprazole (20 mg/kg), the reference compound used also showedignificant reduction of all these secretory parameters. The ulcernhibition was found to be 9.69 and 41.49% at a dose of 50 and00 mg/kg, respectively, while standard drug showed 67.74% ulcer

nhibition in comparison to control group (Table 2). The higheroses (100 mg/kg) showed similar level of action as that of refer-nce compound.

.6. Gastric lesions induced by ethanol

Cedrus deodara (100 mg/kg) treatment showed significant pro-ection from ulcerative lesions caused by ethanol, when comparedo Rabeprazole (20 mg/kg) and control. In the ethanol-inducedlceration study, pretreatment with Cedrus deodara extract pro-uced a significant decrease in number of ulcer, ulcer score andlcer index at 100 mg/kg dose levels (Fig. 1A–C). The ulcer inhibitionas found to be 50.17% at a dose of 100 mg/kg while standard drug

howed 69.95% ulcer inhibition in comparison to control group.

.7. Histopathological evaluation

Pylorus ligation caused histopathological lesions includingegeneration, hemorrhage, and edematous appearance of the gas-ric tissue. Pretreatment with Cedrus deodara (100 mg/kg) andabeprazole (20 mg/kg) offered significant protection against alluch damage to the mucosa.

. Discussion

Cedrus deodara is known to possess various therapeutic proper-ies and has been one of the most noteworthy plant mentionedn various medicinal system. In addition to other therapeuticotentials, Cedrus deodara is also reported to possess some anti-lcerogenic activity. In our studies, we went a step ahead and havetudied gastric antisecretory and antiulcer activities of Cedrus deo-ara in two different models including pyloric ligation and ethanolnduced ulcer model.

The causes of gastric ulcer in pyloric ligation are believed to beue to increase in gastric hydrochloric acid secretion and/or stasisf acid, leading to auto digestion of the gastric mucosa and break-

own of the gastric mucosal barrier. These factors are associatedith the development of upper gastrointestinal damage including

esions, ulcers and life threatening perforation and hemorrhage.n the present study, Cedrus deodara wood oil administration sig-ificantly decreased the gastric content, pH, total acidity and free

llowed by Dunnett’s t-test.

acidity induced by pylorus ligature. This may possibly be due to theantisecretory property of Cedrus deodara wood oil.

The volatile oil of Cedrus deodara significantly reduced num-ber of ulcer, ulcer score, and ulcer index at a dose of 100 mg/kg.The ulcer inhibition was found to be 41.49% at a dose of 100 mg/kgwhile standard drug showed 67.74% ulcer inhibition in comparisonto control group. It could be suggested that Cedrus deodara woodoil can suppress gastric damage induced by aggressive factors. Theantiulcer activity of Cedrus deodara wood oil is further supported byhistopathological study which showed protection of mucosal layerfrom ulceration and inflammation.

Ethanol-induced gastric lesions are due to superficial damage tomucosal cells caused by the direct action of ethanol and gastric acid

Fig. 1. Effect of volatile oil of Cedrus deodara at 100 mg/kg, and reference compoundRabeprazole at 20 mg/kg on number of ethanol induced gastric lesions in rats. (A)Number of ulcer; (B) ulcer score; (C) ulcer index. Statistical analysis was done byone-way ANOVA followed by Dunnett’s t-test. *P < 0.05, **P < 0.01 and ***P < 0.001were considered significantly different as compared to control (n = 6 in each group).

A. Kumar et al. / Journal of Ethnopharmacology 134 (2011) 294–297 297

Table 2Effect of volatile oil of Cedrus deodara on gastric lesion in pylorus ligation induced ulceration in rats.

Treatment Dose (kg−1) Number of ulcers Ulcer score Ulcer index Ulcer inhibition (%)

Control 10 ml 5.66 ± 0.7 2.33 ± 0.4 18.08 ± 1.2 –Cedrus deodara 50 mg 4.66 ± 0.5 1.66 ± 0.5 16.33 ± 1.03a 9.69Cedrus deodara 100 mg 1.50 ± 1.0c 0.75 ± 0.5c 10.58 ± 1.57b 41.49Rabeprazole 20 mg 0.33 ± 0.2c 0.33 ± 0.2c 5.83 ± 0.85c 67.74

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assive intracellular accumulation of calcium represents a majortep in the pathogenesis of gastric mucosal injury. Cedrus deodaraas found to offer significant protection against gastric ulceration

aused by 90% ethanol. In the present study Cedrus deodara wasound to be effective in decreasing the number of ulcer, ulcer scorend ulcer index at a dose of 100 mg/kg. The ulcer inhibition wasound to be 50.17% at a dose of 100 mg/kg while standard drugabeprazole 20 mg/kg, showed 69.95% ulcer inhibition in compar-

son to control group.The phytochemical tests of volatile oil of Cedrus deodara wood

howed the presence of various phytoconstituents viz. terpenoids,henols, alcohol and ketone. It is known that triterpenoid com-ound have antisecretory and cytoprotective activity (Mahato anducharita, 1997) which partially explain the mechanism of actionf volatile oil of Cedrus deodara. Further studies are warranted tosolate the antiulcer compound and to elucidate their exact mech-nism of action.

. Conclusion

The results of this study confirm the use of the volatile oil ofedrus deodara in the traditional management of peptic ulcer dis-ase. The mode of action of the volatile oil may pave the way for thestablishment of a new gastric antisecretory and antiulcer therapyegimen that will not require the use of antacids and antisecretorygents. However, further studies to identify the active moieties andlucidations of the mechanism of action are recommended.

cknowledgements

Authors would like to thank Prof. (Dr.) Ranjit Singh, Director,chool of Pharmaceutical Sciences, Shobhit University and otheraculty members for their technical assistance.

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