SCREENING of AQUEOUS LEAF EXTRACT of Murraya Koenigii Linn for NEPHROPROTECTIVE ACTIVITY

SCREENING OF AQUEOUS LEAF EXTRACT OF Murraya koenigii Linn FOR NEPHROPROTECTIVE ACTIVITY IN RATS

SYNOPSIS FOR

M.PHARM DISSERTATION

SUBMITTED TO

RAJIV GANDHI UNIVERSITY OF HEALTH SCIENCES

KARNATAKA

BY

HEMANG RAMESHBHAI PATEL

DEPARTMENT OF PHARMACOLOGY

NARGUND COLLEGE OF PHARMACY

BANGALORE-560085

(2011 – 2012)

RAJIV GANDHI UNIVERSITY OF HEALTH SCIENCES

KARNATAKA, BANGALORE.

ANNEXURE-II

PROFORMA FOR REGISTRATION OF SUBJECTS FOR PG DISSERTATION

1. / Name of the candidate and address (in block letters) / HEMANG RAMESHBHAI PATEL

NO:21 , DEVAKINANDAN,

6TH MAIN, DATTATREYANAGARA,
BSK 3RD STAGE, HOSAKEREHALLI,
BANGALORE-560085
2. /

Name of the institution

/

NARGUND COLLEGE OF PHARMACY

2ND MAIN, DATTATREYANAGARA,

BSK 3RD STAGE, HOSAKEREHALLI,
BANGALORE-560085
3. /

Course of study and subject

/

MASTER OF PHARMACY IN

PHARMACOLOGY

4. /

Date of the admission

/ 16th DECEMBER 2011
5.
6. /

Title of the topic:

SCREENING OF AQUEOUS LEAF EXTRACT OF Murraya koenigii Linn FOR NEPHROPROTECTIVE ACTIVITY IN RATS
BRIEF RESUME OF THE INTENDED WORK

6.1 NEED FOR THE STUDY

Humans are exposed intentionally and unintentionally to a variety of diverse chemicals that harm the kidney.[1] As the number of drugs, natural products, industrial chemicals and environmental pollutants that cause nephrotoxicity is increasing, it has become clear that chemical molecules with very diverse structure produce nephrotoxicity.[1]
Nephrotoxicity is defined as the partial loss of function of kidney associated with nephritic syndrome, glomerulosclerosis, persistent albuminuria, declining glomerular filtration rate (GFR), elevated arterial blood pressure and fluid retention.[2]
A large number of drugs like analgesics (acetaminophene, aspirin and NSAIDs), antidepressants (doxepine), antihistamines (diphenhydramine,doxylamine), aminoglycosides(amphotericin B), β-lactams (penicillin, cephalosporines), proton pump inhibitors (pantoprazole, lansoprazole) that are in routine use and chemotherapeutic agents like carmustin, lomustin, cisplatin are known to initiate nephrotoxicy [3].
Mitochondrial damage resulting in deficiency of adenosine triphosphate (ATP), is the leading cause of renal cell injury as they are mainly depend on aerobic metabolism.[1] Drugs or their metabolites are also known to bring about nephrotoxicity by acting as compliments that trigger immune reactions resulting in T-cell infiltration into the kidney and subsequent damage leading to nephritis.[4,5]
Murraya koienigii Linn (Rutaceae) commonly known as meethi neem, is an aromatic more or less deciduous shrub or a small tree up to 6 m in height, found throughout the India up to an altitude of 1500 m and are cultivated for its aromatic leaves.[6]
In traditional system of medicine it is used as antiemetic, antidiarrhoeal, antidysentry, febrifuge, blood purifier, tonic, stomachic, flavouring agent in curries and cheutneys. The oil is used externally for bruises, eruption, in soap and perfume industries.[6]
Murraya koeinigii is having powerful antioxidant property[7], and according to Malaysian practitioners, it is beneficial in diabetic kidney damage.[8] It is also reported in Ayurvedic system of Indian medicine for the treatment of kidney pain.[9,10] In vitro studies of the plant extract was carried out for direct scavenging of NO using sodium nitroprusside. The result showed that M. koenigii might be potent and novel therapeutic agents for scavenging of NO. It may therefore be tried for possible use in vivo as a regulator of pathological conditions caused by excessive generation of NO and its oxidation product, peroxynitrite. Involvement of peroxides in the pathogenesis of drug mediated and immune mediated nephritis has been repeatedly reported in the literature.[11] The current study will be carried out to assess the usefulness of M. koeinigii as a nephroprotective agent.
6.2  REVIEW OF LITERATURE:
Scientific name: Murraya koenigii Linn
Family: Rutaceae.
Synonyms:[12]
Gujarati: Kadhilimbdo, goranimb
Kannada: Karibevu
Hindi: Curry patta, mitha neem
Marathi: Karhinimb
Tamil: Karivempu
Telugu: Karepaku
Murraya koienigii Linn (Rutaceae) commonly known as meethi neem, is an aromatic more or less deciduous shrub or a small tree up to 6 m in height, found throughout the India up to an altitude of 1500 m and are cultivated for its aromatic leaves.[6]
In traditional system of medicine it is used as antiemetic, antidiarrhoeal, antidysentry, febrifuge, blood purifier, tonic, stomachic, flavouring agent in curries and cheutneys. The oil is used externally for bruises, eruption, in soap and perfume industries.[6]
The phytoconstituents isolated so far from the leaves are alkaloids viz., mahanine[13], koenine, koenigine, koenidine[14], girinimibiol, girinimibine[15], O-methyl-murrayamin A, O-methyl mahanine, bispyrayafoline[16] and glycosides viz., scopotin, murrayanine[17], calcium, phosphorus, iron, thiamine, riboflavin, niacin, vitamin C, carotene and oxalic acid. The essential oils from leaves yielded di-α-phellandrene, D-sabinene, D-α-pinene, dipentene, D-α-terpinol and caryophyllene.[18]
It is reported to possess antioxidant, antibacterial, antifungal, larvicidal, anticarcinogenic, hypoglycemic, anti-lipid peroxidative, hypolipidemic and antihypertensive activity.[19]
Following activities have been reported:
·  Hepatoprotective activity of leaf extract of Murraya koeinigii Linn.[20]
·  Immunomodulatory activity of methanolic extract of Murraya koeinigii Linn.[21]
·  Anti-inflammatory activity of Murraya koeinigii Linn on experimental animals.[22]
·  Antioxidant activity and antitumour promoying properties of Murraya koeinigii Linn.[7]
·  Antiulcer activity of Murraya koeinigii Linn in albino rats.[23]
·  Antibacterial activity of Murraya koeinigii Linn.[24]
·  Antidiarrhoeal activity of Murraya koeinigii in rats.[25]
·  Anthelmintic activity of Murraya koeinigii Linn leaves.[26]
·  Antinociceptive activity of Murraya koeinigii Linn.[27]
·  Anticancer activity of Murraya koeinigii Linn.[28]
6.3 OBJECTIVE OF THE STUDY
The objective of this study is to evaluate the nephroprotective activity of aqueous leaf extract of Murraya koeinigii Linn in rats.
MATERIAL AND METHODS
7.1 SOURCE OF DATA :
The data will be generated by performing experiments on animals; the standard information is collected from various journals, standard text books available in library of Nargund College of Pharmacy, RGUHS-digital library and from various standard websites.
Web sites:
www.sciencedirect.com
www.pubmed.com
www.ijp-online.com
www.google.com
www.rguhs.ac.in/j-gate@helinet
7.2  METHOD OF COLLECTION OF DATA:
The data collected will be based on animal experimentation as per the parameters studied under each animal model. The sampling procedure and collection of data will be as follows.
EXPERIMENTAL MODEL
1.  NEPHROTOXIC SERUM INDUCED NEPHRITIS[29]
·  Animals: Albino Wistar Rats.
·  Weight: 150-200gm.
·  Test drug: Aqueous leave extract of Murraya koeinigii
·  Groups: 6 groups of 6 animals each.
PREPARATION OF NEPHROTOXIC SERUM
Rat kidneys are fully perfused with physiologic saline through a catheter placed in the aorta. Renal cortical tissue is removed, homogenized and diluted with physiological saline at about 20% suspension.2 ml of renal cortical homogenate is emulsified with an equal amount of Freunds adjuvant. The emulsion is injected into the rabbits. Sera is decomplemented for 30 min at 560C and absorbed with freshly harvested rat RBC. (n = 6 Rats and 6 Rabbits)
Following groups are made
Group 1 (n = 6): animals will be treated with Saline alone
Group 2 (n = 6): animals will be treated with Nephrotoxic serum alone
Group 3 (n = 6): animals will be pretreated for 2 weeks with Murraya koeinigii extract (Dose 1)
Followed by Nephrotoxic serum
Group 4 (n = 6): animals will be pretreated for 2 weeks with Murraya koeinigii extract (Dose 2)
Followed by Nephrotoxic serum
Group 5 (n = 6): animals will be pretreated with Nephrotoxic serum followed by selected dose of
Murraya koeinigii extract for 2 weeks
Parameters to be estimated: Creatinine
Blood Urea Nitrogen
Albumin
Histopathology
2.  CISPLATIN INDUCED NEPHROSIS:[30]
Rats will be administered cisplatin (7 mg/kg i.v., once) and will develope nephropathy in six to seven days.
·  Animals: Albino Wistar Rats.
·  Weight: 150-200gm.
·  Test drug: Aqueous leave extract of Murraya koeinigii
·  Groups: 6 groups of 6 animals each.
Group 1 (n = 6): animals are treated with Cisplatin alone
Group 2 (n = 6): animals are treated with Murraya koeinigii extract alone
Group 3 (n = 6): animals are pretreated for 2 weeks with Murraya koeinigii extract (Dose 1)
Followed by Cisplatin
Group 4 (n = 6): animals are pretreated for 2 weeks with Murraya koeinigii extract (Dose 2)
Followed by Cisplatin
Group 5 (n = 6): animals are treated with Cisplatin followed by Murraya koeinigii extract for 2
Weeks
Parameters to be estimated: Creatinine
Blood Urea Nitrogen
Albumin
Histopathology
7.3 STATISTICAL ANALYSIS:
Data will be analyzed using one way analysis of variance (ANOVA)
7.4 Does the study require any investigation to be conducted on patients
Or other humans or animals? If so please describe briefly
Yes, The above study requires investigation on animals. The effect of drug will be studied
on various parameters using rat as animal model.
7.5 Has ethical clearance been obtained from your institution in case of 7.4?
Application has been submitted to “Institutional animal Ethics Committee” of Nargund
College of Pharmacy.
8. LIST OF REFERENCES:
1.  Rick G. Schnellmann, Katrina J. Kelly. Pathophysiology of Nephrotoxic Acute Renal Failure.
2.  Ogi M, Kojima S, Kuramochi M. Effect of postural change on urine volume and urinary sodium excretion in diabetic nephropathy. Am J Kidney Dis 1998;31:41-8.
3.  Cynthia A. Naughton, PharmD, BCPS, North Dakota State University College of Pharmacy, Nursing, and Allied Sciences, Fargo, North Dakota. Drug induced nephrotoxicity. Am Fam Physician.2008Sep15;78(6):743-750.
4.  Anders HJ, Vielhauer V, Schlondorff D: Chemokines and chemokine receptors are involved in the resolution or progression of renal disease. Kidney Int 2003; 63: 401–415.
5.  Vielhauer V, Anders HJ, Schlondorff D: Chemokines and chemokine receptors as therapeutic targets in lupus nephritis. Semin Nephrol,2007; 27: 81–97.
6.  Ajay S, Rahul S, Sumit G, Paras M, Mishra A, Gaurav A, Himalayan Institute of Pharmacy, Kala Amb (HP), Venkateshwara College of Pharmacy, Meerut (U.P.), Faculty of Pharmacy, Amity University, Lucknow (U.P.). AND College of Pharmacy, Babnan, Gonda (U.P). Faculty of Pharmacy, RPETGI, Karnal (HR). Comprehensive review: Murraya koenigii Linn. Asian Journal of Pharmacy and Life Science. ISSN 2231 – 4423
7.  Yih Yih Kok, Lim Yang Mooi, Kartini Ahmad, Mohd Aspollah Sukari,
Nashriyah Mat, Mawardi Rahmani and Abdul Manaf Ali. Anti-Tumour Promoting Activity and Antioxidant Properties of Girinimbine Isolated from Murraya koenigii. Molecules 2012, 17, 4651-4660; doi:10.3390/molecules17044651
8.  Hassan Yankuzo, Qamar Uddin Ahmed, Rahajoe Imam Santosa, Seikh Farid Uddin Akter, Norlewati A. Talib. Beneficial effect of the leaves of Murraya koeinigii (Linn) on diabetes induced renal damage invivo. Journal of Ethnopharmacology,Volume 135, Issue 1,26 April 2011,Pages 88-94
9.  Saligrama Ramachandra Rao encyclopaedia of Indian Medicine- Materia Medica- Herbal drugs. Volume 4, Popular Prakashan, New Delhi, 2005 pg. 116.
10.  Dr. H.K. Bakhru, Indian Spices & Condiments as Natural Healers, Jaico Publishing House, Mumbai. 2007 pg. 67-68.
11.  Baliga MS, Jagetia GC, Rao SK, Babu K. Evaluation of nitric oxide scavenging activity of certain spices in vitro: A preliminary study. Nahrung. 2003; 47(4): 261-4.
12.  Kripi Vohra and Vivek Kumar Gupta. Murraya koenigii (Linn.) Spreng (Rutaceae): A precious Gift from the Nature. IJPRR 2011, 3(1), 18-25.
13.  Narasimhan NS, Paradkar MV, Kelkar SL. Alkaloids of Murraya koenigi, Structures of mahanine, koenine, koenigine and koenidine. Indian J Chem 1970; 8:473-476.275
14.  Narasimhan NS, Paradkar MV, Chitguppi VP, Kelkar SL. Alkaloids of Murraya koenigi, Structures of mahanimbine, koenimbine, mahanine, koenine, koenigine and koenidine. Indian J Chem 1975; 13: 993-995.
15.  Adebajo AC, Avoola OF, Iwalewa EO, Akindahunsi AA, Omisore NO, Cadewunmi CO et al. Anti-trichomonal, biochemical and toxicological activities of methanolic extract and some carbazole alkaloids isolated from the leaves of Murraya koenigii growing in Nigeria. Phytomedicine 2006; 13(4): 246-54.
16.  Tachibana Y, Kikuzaki H, Lajis NH, Nakatani N. Comparison of antioxidative properties of carbazole alkaloids from Murraya koenigii leaves. J Agric Food Chem 2003; 51(22): 6461-7.
17.  Adebajo AC, Reisch J: Minor furocoumarins of Murraya koenigii. Fitoterpia 2000; 71(3): 334-7.
18.  Gopalan C, Rama Shastri BV, Balasubramanian SC. Nutritive value of Indian Foods. New Delhi: ICMR, 1984: 66,117.
19.  Iyer D, Uma DP. Phyto-pharmacology of Murraya koenigii. Pharmacognosy Reviews 2008; 2: 180-184.
20.  Sadhana Sathaye, Yogita Bagul, Sanjay Gupta, Harpreet Kaur, Roopali Redkar. Hepatoprotective effects of aqueous leaf extract and crude isolates of M.koenigiiagainstin vitroethanol-induced hepatotoxicity model. Experimental and Toxicologic Pathology,Volume 63, Issue 6,September 2011,Pages 587-591.
21.  Abhishek S. Shah, Alok S. Wakade, Archana R. Jouvekar. Immunomodulatory activity of methanolic extract of Murraya koeinigii leaves. Indian Journal of Experimental Biology. Vol. 46, July 2008, pp. 505-509.
22.  Vaibhav M. Darvekar, Vijay R. Patil and Amol B. Choudhari. Anti-inflammatory activity of Murraya koenigii Spreng on experimental animals. J. Nat. Prod. Plant Resour., 2011, 1 (1): 65-69
23.  Dinesh Kumar Patidar. Anti-ulcer activity of aqueous extract of murraya koenigii in
Albino rats. International Journal of Pharma and Bio Sciences. Jan-Mar 2011 Vol. 2/ Issue- 1

24.  Bhaskar K. Gupta, Mukul Tailang, Alkesh K. Lokhande, Eman A. Subhey, Narayan P. Gavatia, Arnab Sarkar. Antimicrobial activity of ethanolic extracts of Murraya Koenigii by disc diffusion and broth dilution method. Journal of Pharmacy Research,(2011).Vol 4, No 4.

25.  Praveen Sharma, Gali Vidyasagar, Anil Bhandari, Sunder Singh, Upendra Bhadoriya, Santosh Ghule, Nitin Dubey. A pharmacological evaluation of antidiarrhoeal activity of leaves extract of Murraya koenigii in experimentally induced diarrhoea in rats. Asian Pacific Journal of Tropical Disease (2012)230-233.
26.  Uma Shankar Sharma, Umesh Kumar Sharma, Abhishek Singh, Niranjan Sutar, and Puspak Jyoti Singh. InVITRO ANTHELMINTIC ACTIVITY OF Murraya Koeinigii Linn LEAVE EXTRACT. International Journal of Pharma and Bio Sciences. Sep.2010 Vol.1/Issue-3/Jul-.
27.  Rupali Arun Patil,Padmaja Mukund Langade,Pramod Babarao Dighade,Yogesh Ashok Hiray. Antinociceptive activity of acute and chronic administration ofMurraya koenigiiL. leaves in experimental animal models. 2012 /Volume: 44/Issue: 1/Page: 15-19.
28.  P. Muthumani, S.Venkatraman , K.V.Ramseshu, R.Meera , P.Devi , B. Kameswari, B.Eswarapriya. Pharmacological studies of anticancer, anti inflammatory activities of Murraya koenigii (Linn) Spreng in experimental animals. P.Muthumani et al /J. Pharm. Sci. & Res. Vol.1(3), 2009, 137-141.
29.  Vogel HG, Wolfgang HV. Drug discovery and evaluation- Pharmacological assays. 2nd Edition. Springer Verlag Berlin Heisdelberg Publications; 1997.
30.  Pitchai Balakumar, Vishal Arvind Chakkarwar, Vijay Kumar, Akash Jain, Jayarami Reddy, Manjeet Singh. Experimental models for nephropathy. Journal of Renin-Angiotensin-Aldosterone System 2008 9: 189.
09. / SIGNATURE OF THE CANDIDATE /
HEMANG RAMESHBHAI PATEL
10. / REMARKS OF THE GUIDE / RECOMMANDED
11. / NAME AND DESIGNATION
11.1 NAME OF THE GUIDE / Prof D S PURANIK
ASST. PROFESSOR
DEPT. OF PHARMACOLOGY,
NARGUND COLLEGE OF PHARMACY, BANGALORE-560 085.
11.2 SIGNATURE
11.3 CO-GUIDE (IF ANY) / NAGAMANI KOPPARAPU
ASST. PROFESSOR
DEPT. OF PHARMACOLOGY,
NARGUND COLLEGE OF PHARMACY, BANGALORE-560 085.
11.4  SIGNATURE / .
11.5  HEAD OF THE DEPARTMENT
/ DR.H.J.HRISHIKESHAVAN
PROFESSOR & H.O.D,
DEPT. OF PHARMACOLOGY,
NARGUND COLLEGE OF PHARMACY, BANGALORE-560 085.
11.6 SIGNATURE
12. / 12.1 REMARKS OF THE PRINCIPAL
/ RECOMMANDED FOR REGISTRATION
12.1 SIGNATURE
12.2 PRINCIPAL / Prof D S PURANIK
PRINCIPLE,
NARGUND COLLEGE OF PHARMACY,
BANGALORE-560085

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