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Antidiarrhoeal activity of the methanol extract of the barks of Xylocarpus moluccensis in castor oil- and magnesium sulphate-induced diarrhoea models in mice

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Antidiarrhoeal activity of the methanol extract of the barks of Xylocarpus moluccensis in castor oil- and magnesium sulphate-induced diarrhoea models in mice
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  ISSN 2229  –  6859 IJPI’s Journal of Pharmacology and Toxicology  Visit www.ijpijournals.com Antidiarrhoeal Activity of Methanol Extract of Leaves of Calotropis Procera R. Br Patil S.H.   , Adkar P.P., Shelke T.T., Oswal R.J., Borase S.P.,   Department of Pharmacology & Department of Pharmaceutical Chemistry, JSPM‟s Charak College of Pharmacy and Reasearch, University of Pune-412207, Maharashtra, Corresponding Author:   Adkar Prafulla P.   Email address : prafi.phd@gmail.com   . ABSTRACT The leaves of Calotropis procera R.Br   was undertaken for evaluation of antidiarrhoeal activity in Albino mice (male) weighing between 40-50gm was used in this study. The selected and authenticated plant extract was prepared by maceration with using a methanol as a solvent for 7 days. From the toxicity study; it was observed that plant extract is non toxic and caused no death up to the dose of 500 mg/kg. It is safe and used for further experiment. The methanol extract of Calotropis procera showed significant activity against castor oil induced diarrhoea to compared standard drug Loperamide in albino mice. All the values obtained in the studies were expressed as mean ± SEM. by one way of analysis of variance for determining the significant difference. On the basis of these findings, it can be assumed that Calotropis procera R.Br could be a potential source for novel „lead‟ discovery for an tidiarrhoeal drug development. Further study is necessary to evaluate the active principle responsible for the antidiarrhoeal activity and clear mechanism of action involved. Keywords:   Calotropis procera  R.Br; Castor oil; Antidiarrhoeal activity; Loperamide.  Vol 1:3 (2011)   IJPI’S Journal of Pharmacology and Toxicology  Patil .S  et al   Page 26 1. INTRODUCTION Diarrhoea is the abnormal increase of liquid in stool and increase in the frequency of passing stool (defecation). (1) To most persons, diarrhoea means an increased frequency or softer consistency of bowel movements; however, the medical definition is more exact than this. Diarrhoea best correlates with an increase in stool weight; stool weights above 300 g per day generally indicates diarrhoea. This is mainly due to excess water, which normally makes up 60-85% of fecal matter. In this way, true diarrhoea is distinguished from diseases that cause only an increase in the number of bowel movements (hyperdefecation), or incontinence (involuntary loss of bowel contents). (2-8)  The plant Calotropis procera R.Br is found in most parts of the world in dry, sandy and alkaline soils and warm climate. (9)  The plant Calotropis procera R.Br commonly known as small crown plant is used commonly for tooth ache, dysentery, and as antidote for snake bite. The main constituent in the plant are proteins, tannins and resin. The latex of the plant is used as an antidysenteric, antirheumatic, a diaphoretic, an expectorant and for the treatment of bronchial asthma and skin conditions.  (10-11) In the present study antidiarrhoeal activity of methanol extract of Calotropis procera R.Br. has been explored to gain its possible mechanism. 2. MATERIAL AND METHODS 2.1 Plant Material Collection & Extraction The leaves of Calotropis procera R.Br was collected from the surrounding area of Village Wagholi, Pune, Maharashtra in November 2009 .The leaves were dried under sunlight with occasional shifting and then powdered with a mechanical grinder and The obtained powder was subjected to maceration process for 7days by using methanol as solvent; after that which is filtered and simple distillation is done to get the final extract. The extracted drug is used for evaluation of antidiarrhoeal activity. (9)   2.2 Animal Albino mice (male) weighing between 40-50gm was used in this study. The cages of animals were placed in a room temperature with controlled cycles of 12h light and 12h of darkness the relative humidity was maintained at 44-45%. All animal were fed with standard pellet diet & water ad libitum. Animal experiments were conducted according to the guidelines of institutional animal ethical committee. The animal bed in the cages was renewing thrice a week to ensure hygienic condition and maximum comfort of animals. 2.3 Toxicity study An acute toxicity study relating to the determination of LD50 value was performed using different doses of the extracts. From the toxicity study; it was observed that plant extract is non toxic and caused no death up to the dose of 500 mg/kg. It is safe and used for further experiment. 2.4 Castor Oil Induced Diarrhea The animals were screened initially by giving 0.5ml of castor oil and only those showing diarrhea were selected for final experiment. The animals were divided in to control, positive and test groups, with six mice in each group. The control group received vehicle (1% tween 80 in water) at a dose of 10 ml/kg orally. The positive control group received Loperamide at the dose of 3 mg/kg orally. The test group received methanol extract at the doses of 250 and 500 mg/kg orally. Each animal was placed in an individual cage, the floor of which was lined by blotting paper. The floor lining was changed every hour. Diarrhoea was induced by oral administration of castor oil; 30 min after the above treatments. During an observation period of 4h total number of feces and the number of diarrheic feces excreted  Vol 1:3 (2011)   IJPI’S Journal of Pharmacology and Toxicology  Patil .S  et al   Page 27 by animals were recorded. The numerical score based on stool consistency was assigned as follows; normal stool=1, semisolid stool=2, and watery stool=3. (12-20) 2.5 Statistical Analysis The data obtained in the studies were subjected to one way of analysis of variance (ANOVA) for determining the significant difference. The inter group significance was analyzed using Dunnet‟s - t test. A „P‟ value<0.05 we re considered to be significant. All the values were expressed as mean ± SEM. 3. RESULTS In the mice with castor oil-induced diarrhea, the methanol extract of leaves of Calotropis procera R.Br. at doses of 250 and 500 mg/kg, reduced the total number of feces as well as diarrheic feces as shown in the Table 1.  In the present study antidiarrhoeal activity of selected plant Calotropis procera R.Br methanol extract on Swiss albino mice at dose 500mg/kg was carried out. Thus Histogram 1 shows total number of feces and Histogram 2  show weight of feces. Both the graph shows diarrhoea between controls, standard and extract (test) drug. 4. DISCUSSION Diarrhoea is a very common ailment and national problem in many tropical countries and the cause of 4-5million deaths throughout the world annually. Apart from modern medical therapy, the use of herbal drugs in the treatment of diarrheal diseases is a common practice in many countries of Asia including India. In the present study methanol is used as the solvent system to prepare crude methanol extract. To avoid any solvent effect on the experimental animal, the solvent was evaporated completely to dryness to yield a non sticky solid mass. The methanol extract of Calotropis procera R.Br shows significant reduction in the total number of wet feces in a dose dependent manner. Castor oil induced diarrhea is a secretary diarrhea since ricinolic acid, the active ingredient of castor oil, induces diarrhea by a hypersecretory response. Since the methanol extract Calotropis procera  R.Br successfully inhibited castor oil diarrhea, it can be assumed that the antidiarrhoeal action was mediated by an antisecretary mechanism. This was also evident from the reduction of total number of wet feces in test group in the experiment. Table 1:- Antidiarrhoeal Activity of Methanol Extract of Leaves of Calotropis Procera R. BR. In albino Mice  Sr.   Treatment   Dose (mg/kg)   No of feces   Total number of feces in hr.   Weight of feces in gm.   2hr   4hr   2hr   4hr   1   Control (1% tween 80) in water.   0.01ml/g   22   8.80±0.663   13.60±0.245   0.76±0.17   0.8980±0.032   2   Loperamide   3.0   2   0.600±0.400   0.200±0.201   0.23±0.20   0.3100±0.011   3   Methanol extract   250.0   6   2.610±0.245 *   2.00±0.316 **   0.43±0.20 *   0.5900±0.014 **   4   500.0   3   1.600±0.510 **   0.60±0.400 **   0.31±0.46 **   0.4460±0.00 **   Values are expressed as mean ± S.E.M. n = 6, ** P< 0.01, *P< 0.5 compared with vehicle Control one way ANOVA followed by Dunnet‟s comparison test.    Vol 1:3 (2011)   IJPI’S Journal of Pharmacology and Toxicology  Patil .S  et al   Page 28 Histogram-1   Histogram-2 8   .813.60.60.22.6121.70.602468101214 Total no. of feces 1 2 3 4 Treatments Histogram ofAntidirrhoeal Activity :- Total no. of feces Time in 2 & 4 hr In Mice. Series1 Series2 0.760.90.230.310.430.590.310.4400.10.20.30.40.50.60.70.80.9 Weight of feces in Time 2hr,4hr, in (gm) 1234 Treatments Antidirrhoeal Activity of Calotropis procera Linn in Mice :-Weight of feces weight of feces in 2hr.in gm.Weight of feces in 4hr,in gm.  Vol 1:3 (2011)   IJPI’S Journal of Pharmacology and Toxicology  Patil .S  et al   Page 29 5. CONCLUSION The extract contains many biologically active constituents which are known to affect gastrointestinal function. From this current study, it can be concluded that methanol extract of leaves of Calotropis procera  R.Br possesses marked antidiarrhoeal activity in dose dependent manner. The present study establishes the effectiveness and pharmacological rationale for use of Calotropis procera  R.Br as an antidiarrhoeal drug. The drug may be further explored for its phytochemical profile to identify the active constituents responsible for the antidiarrhoeal activity.   On the basis of these findings, it can be assumed that Calotropis procera  R.Br could be a potential source for novel „lead‟ discovery for antidiarrhoeal drug development and number of preclinical trial s. Further study is necessary to evaluate the active principle responsible for the antidiarrhoeal activity and clear mechanism of action involved. 6, ACKNOWLEDGEMENT The Authors greatly thankful and expresses deep sense of gratitude to JSPM‟S Group of Institute and University of Pune, Pune, India, and also last but not least to Botanical Survey of India, Pune. for authenticating the plant. 7. REFERENCES 1.   Adkar P.P., Kaswa K. S., Oswal R.J., Shelke T.T. Bhaskar V.H., “The Pharmacological evaluation of Punica granatum   Linn. For antidiarrhoeal activity”. Poster presented in the  International Conference on   “Resent developments in Pharmacological research”,  Pravaranagar, Ahemadnagar, Maharashtra, India. On 3 rd  -5 th  March, 2010. 2.   Fine, Kenneth D. "Diarrhoea." In Sleisenger & Fordtran's Gastrointestinal & Liver Disease.  Edited By Mark Feldman, Et Al. Philadelphia: W. B. Saunders Company, (1997), 129-130. 3.   Wilson ME, "Diarrhoea in Nontravelers: Risk and Etiology". Clinical. Infectious Disease  41th Supplement, (2005), 541-546. 4.   Ghishan, Fayez K. "Chronic Diarrhoea." In  Nelson Textbook Of Pediatrics , Edited By Richard E. Behrman Et Al. Philadelphia: Saunders, 1 7th  Edition, (2003), 1276  –  1280. 5.   Bodhankar .S.L & Vyawhare.N.S,  A Textbook of Pathophysiology, Nirali Publication, Pune. 3 rd  Edition,(2007), 7.9-7.11. 6.   Sack, R. Bradley. "The Diarrhoea of Travelers." In Cecil Textbook Of Medicine , Edited By Lee Goldman Et Al. Philadelphia: Saunders, 22 nd  Ed., (2003), 1864  –  1845. 7.   Lenhard, Raymond E, Robert T. Osteen, and Ted Gansler. Clinical Oncology . American Cancer Society, (2000), 245-247. 8.   Barar FSK, “  Essential of Pharmacotherapeutics”  S. Chand & Company Ltd, New Delhi, 4 th  edition, (2007), 540. 9.   Jarald E., Edwin S., Pandian S., Damodaran S., Jain P., Patel J., Antimicrobial activity of leaves of  Lantana camara  Linn & Calotropis procera  R. Br. Indian drugs 45(9) September (2009). P-736. 10.   Lawrence S., And Kurt J. Isselbacher. "Diarrhoea." In Harrison's Principles of Internal Medicine. Edited By Anthony S. Fauci, Et Al. New York: Mcgraw Hill, (1998), 443-445. 11.   Ramarao A.V. & Gujar M.K.: “  Drugs from Plant Resources ” An overview, Pharmatimes. P. 19 -27. 12.   Nirmala R Nargis akhtar. Isolation and structural studies on the constituent Procera, Plumeria rubra ,  Amberboa ramose  & Calotropis procera  Thesis (1992), 138-163.
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