“antivenom activity of cinnamomum zeylanicum extract against naja kaouthia snake venom




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“ANTIVENOM ACTIVITY OF CINNAMOMUM ZEYLANICUM EXTRACT AGAINST NAJA KAOUTHIA SNAKE VENOM "



SYNOPSIS FOR:

M. PHARM DISSERTATION


SUBMITTED TO:

RAJIV GANDHI UNIVERSITY OF HEALTH SCIENCES

BENGALURU, KARNATAKA.
SUBMITTED BY:

MITUL N PATEL

1st YEAR M.PHARM

DEPARTMENT OF PHARMACOLOGY

GUIDE

MR. MUKUND HANDRAL

Asst, Professor, Dept. Pharmacology

PES College of Pharmacy, Bangaluru.



P.E.S COLLEGE OF PHARMACY

BENGALURU – 560050

(2011-2013)

ANNEXURE II

PROFORMA FOR REGISTRATION OF SUBJECT FOR DISSERTATION



1.

Name of the candidate &

Address.



MITUL N PATEL:

#167, RAJ SADAN, 9TH CROSS, 2ND MAIN, SBM COLONY, NEAR SEETHA CIRCLE, BANGALURU-560050


PERMANENT ADDRESS:

S/o NAVINBHAI D PATEL

PAYGA STREET, AT & PO: KAMREJ,

TA: KAMREJ, DI:SURAT,

GUJARAT-394180




2.


Name of the Institution.




PES COLLEGE OF PHARMACY

HANUMATHNAGAR, 50 FEET RIAD,

BANGALURU-560050




3.

Course of the study

& subject.




MASTER OF PHARMACY (M.PHARM)

Sub:- Pharmacology




4.

Date of admission.




05-04-2011



5.

Title of the Topic:

“ANTIVENOM ACTIVITY OF CINNAMOMUM ZEYLANICUM EXTRACT AGAINST NAJA KAOUTHIA SNAKE VENOM



6.

7.

8.



Brief Resume Of Intended Work
6.1 NEED FOR THE STUDY

Snake bite is a serious medico-legal problem observed especially among the snake catchers, forest workers and agriculturists in rural India. There are over 2000 species of snakes in the world and 216 species in India, of which 52 are venomous.1 In India there are many types of venomous snakes; amongst them Indian Cobra (Naja kaouthia) is one of the largest venomous snakes. It found mainly in north-eastern hilly regions of the country.2 The Indian Cobra, family Elapidae posses a dangerous public health problems in numerous tropical and subtropical countries like India. Various estimations showed that 15,000-20,000 people die annually in India due to snake envenomation. The treatment of cobra bite patients requires a medical emergency.3

Polyvalent antivenom (prepared against the venoms of several selected species and sub-species of snakes) therapy is the most common remedy available for neutralizing the life threatening, systemically acting toxins of snake venom. However, the polyvalent antivenom therapy is less effective, nonavailability during emergency and high doses of antivenom lead to adverse reactions that may be fatal.4

Many synthetic and natural remedies are used to treat snake bite. In many countries , plant extracts have been traditionally used in the treatment of snakebite envenmations. However, in most cases, scientific comprovation of their antiophidian activity is still needed. It is believed that around 80% of the world population use plants as first therapeutic help in cases of envenomation.5


In ancient Indian system of Ayurveda and Siddha, there are many plants recommended for using a snakebite therapy. Some of these are popularly used by snake charmers of India for treating snakebite but without any scientific validation. Therefore, this type of treatment remains questionable and needs thorough scientific investigation. Some of the medicinal plants were known to have antivenom properties. According to Siddha system of medicine Cinnamomum zeylanicum is said to have antivenom property and it also contain tannins and it is already established that tannins are responsible for antivenom activity.6 Therefore, in the present investigation, an effort has been given to evaluate the neutralization capacity (antivenom activity) of Cinnamomum zeylanicum bark extracts against lethality, myotoxicity and some toxic enzymes of Naja kaouthia.
6.2 REVIEW OF LITERATURE

Cobra snake venom:

The Cobra snake venom contains cardio toxins, cobra venom factors, a cysteine-rich toxin, cytotoxins, kaouthiagin, mocarhagin, muscarinic toxin-like proteins, neurotoxins, an oxoglutarate dehydrogenase, phospholipases, serum albumin, and a weak toxin.7 In case of Elapidae Snake bite diaphragmatic paralysis is the principal cause of death after envenomation.8 N.kaouthia envenomation causes myonecrosis, muscular degeneration, neurotoxicity, cardiotoxicity, haemolytic and damage to biological membranes.9 The venom produced a progressive decrease in the amplitude of miniature end-plate potentials in the rat hemidiaphragm.10



Cinnamomum zeylanicum:

GC-MS analysis of essential oils of Cinnamomum zeylanicum were reported to identified as benzaldehyde, (E)-cinnamaldehyde and linalool having insecticidal and ovicidal activity.11 The fruit and other part of Cinnamomum zeylanicum reported to contain phenolic antioxidants which counteract the damaging effects of free radicals and protect against mutagenesis.12 The cinnamaldehyde isolated from cinnamon bark extract showed significant decrease in glycosylated haemoglobin, serum total cholesterol, triglyceride levels and at same time increase in plasma insulin, hepatic glycogen and high density lipoprotein-cholesterol.13Aldehydic components of Cinnamon bark namely cinnamaldehyde and 2-methoxycinnamaldehyde suppressers RANKL (receptor activator of nuclear factor-B ligand) induced osteoclastogenesis through NFATc1 (nuclear factor of activated T cell 1) down regulation.14 GC-MS and bioassay revealed that major compound in the Cinnamon oil were (E)cinnamaldehyde , benzaldehye, and (E)cinnamyl acetate and find an application as an antibacterial agent and anticancer agents.15 Extract of Cinnamomum zeylanicum bark reduces palladium ions to nano-scale palladium particles.16



Some of the scientific reports on plant based preparations or agent having antivenom property are given here under:

The aqueous extract of root of Mimosa pudica in vitro neutralize the toxic enzymes and in vivo antagonises the lethality and myotoxicity of Naja kaouthia venom.9 The plant Vitex negundo and Embellica officinalis extracts significantly antagonized the Vipera russellii and Naja kaouthia venom induced lethal activity both in in vitro and in vivo studies.17 Lupeol acetate isolated from the methanolic root extract of Indian medicinal plant Hemidesmus indicus family: Asclepiadaceae which could neutralize venom induced action of Daboia russellii and Naja kaouthia on experimental animals.18 The root extract of Pluchea indica family: Asteraceae neutralize the effect of venom of Viper and cobra like lethality, hemorrhagic, defibrinogenating, and edema-producing action , phospholipase activity, neurotoxicity and cardio toxicity.19







   PLANT PROFILE

Cinnamomum zeylanicum also called as Cinnamon bark is the dried inner bark of the coppiced trees, belonging to the family Lauraceae.

Description: Cinnamomum zeylanicum is considered to be native of Sri Lanka and Malabar coast of India. However, most of the world requirement are met by Sri Lanka and hence true cinnamon is also known as Sri Lanka cinnamon. The altitude at which it grows favourable is 800 to 1000 meters , Sheltered situation with an annual rainfall of 200-250 cm is ideal for cultivation of cinnamon. The bark of Cinnamomum zeylanicum found in form of quills and it is around 1 m in length and 1 cm in diameter and about 0.5mm thick. The outer surface of bark is marked by wavy longitudinal striations with small holes of scars left by the branches. The inner surface also shows the longitudinal striations. Bark is free of cork.

Part used : Bark

Chemical constitutution : Cinnamomum contains about 0.5 to 1% of volatile oil, 1.2% tannins, mucilage, calcium oxalate, starch and a sweet substance known as mannitol. cinnamon oil contains 60-70% of cinnamaldehyde, 5-10% eugenol, benzaldehyde, cuminaldehyde and other terpenes like phellandrene, pinene, cymene, caryophylline, etc.

Uses: Traditionally cinnamon bark is used as carminative, stomachic, mild astringent, flavouring agent, stimulant, aromatic and antiseptic. Commercially, it is used as a spice, condiment, in the preparation of candy, dentifrices and perfumes.

Vernacular name:

Bengali - Dalchin, Gujarati - Ceylon taj, Hindi - Dalchini, Malylam - Dalchini, Hindi - Dalchini, Marathi- Dalchini, , Gujarati - Ceylon taj, Sanskrit - Darushila, Tamil - Lavangapatti.20



6.3 Objective of the study

  1. Evaluation of neutralization of lethality against N.kaouthia venom using mice.

  2. Evaluation of neutralization of myotoxicity of N.kaouthia venom in mice.

  3. Evaluation of neutralization activity of phospholipase A2 of N.kaouthia venom by extract

  4. Evaluation of anti-inflammatory activity of extracts against N.kaouthia snake venom using mice


MATERIALS & METHODS

7.1 Source of data

The plant material will be procured from authenticated suppliers. Whole experiment is planned to generate data from laboratories studies. Experiment will be performed as described in the standard bibliography, may be obtained from standard journals and text books available within the college or from other pharmacy colleges or from libraries of National Institutes or through internets from industry.



www.sciencedirect.com

www.pubmed.com

www.google.co.in

7.2 Methods of collection of data (including sampling procedure if any)

The data will be based on laboratory animal experimentation.



1.Animals

Swiss albino mice (25–33g) of either sex will be collected from central animal facility, PES COLLEGE OF PHARMACY, Bangaluru, India. Animals will be maintained at a temperature of 25±1 ◦C under 12-h light: 12-h dark cycle. The animals will be fed on autoclaved standard mice food pellets and water ad libitum.


2.Venom sample

Lyophilized Naja kaouthia venom will be procured from the authenticated supplier from Hindustan park, Kolkata or Irula snake catchers Industrial Cooperative society, Chennai.



3.Chemicals

All chemicals of analytical grade will be procured from licensed vendor.



4. Collection of plant material

The bark of Cinnamomum zeylanicum will be procured from Authenticated supplier from Bengaluru and it will be authenticated by Taxonomist.



5.Preparation of Exracts and mode of administration

Dried powder of bark material successively extracted with water and alcohol (70%) in a soxhlet apparatus for 72 hours. The residue obtained after extraction is dried. The residue mass obtained is evaporated to dryness under reduced pressure. A % yield of extracts will be determined.

Extracts will be administered by oral route, N.kaouhtia venom and standard(polyvalent antivenom) will be administered by i.p. and i.m. route.
6.Preliminarary phytochemical investigation

Preliminary phytochemical investigation of extracts will be done as described by Practical Pharmacognosy- Techniques and Experiments.21



7.Acute oral toxicity study of Cinnamomum zeylanicum.

Acute oral toxicity studies for the extract of Cinnamomum zeylanicum will be performed according to The Organization of Economic Co-operation and Development (OECD) guidelines NO: 423 and low and high dose will be selected for treatment.22
7.3 EVALUATION OF ANTIVENOM ACTIVITY

1.Determination of Median Lethal Dose of venom (LD50 or MLD)

MLD or LD50 of Naja Kaouthia venom was already established in our laboratory as 0.6 μg/g of mice with a mean survival time of 1.05 hrs by i.p route and 0.3 μg/g by i.v route with a mean survival time of 0.48 hrs.23



2.Neutralization of lethality

In-vivo method

36 Swiss Albino mice weighing (18 – 20 gm) will be devided into 6 groups each containing 6 mice. Group-I will receive phosphate buffer by i.p route , Group-II will receive venom by i.p route and polyvalent antivenom by i.m route , Group-III & IV will receive venom i.p route followed by low and high dose of aqueous extract by oral route Group-V&VI will receive venom i.p route followed by low and high dose of alcoholic extract by oral route.

The animals will be administered either 1.5,2,3 MLD (LD50) of venom (i.p). Venom will be dissolved in 0.1ml of phosphate-buffered saline solution, pH 7.2.Immediately after envenomation mice will receive 250µl of polyvalent antivenom in the left thigh (i.m.)/ required dose of bark extract of Cinnamomum zeylanicum/ Phosphate buffer for comparative purpose. Animals will be observed for 48hrs and time for death will be recorded.24


Group I

Control (Phosphate buffer) i.p

Group II

Venom (i.p)+ Polyvalent Antivenom(i.m)

Group III

Venom(i.p)+ Aq. Extract of C.zeylanicum low dose (p.o)

Group IV

Venom(i.p)+ Aq. Extract of C.zeylanicum high dose (p.o)

Group V

Venom(i.p)+ Alc. Extract of C.zeylanicum low dose (p.o)

Group VI

Venom(i.p)+ Alc. Extract of C.zeylanicum high dose (p.o)



3.Neutralization of myotoxicity of venom in mice

36 Swiss Albino mice weighing (18 – 20 gm) will be divided into 6 groups each containing 6 mice. Group-I will receive phosphate buffer by i.m route, Group-II will receive venom(25μg) by i.m route, Group-III & IV will receive venom(25μg) i.m route followed by low and high dose of aqueous extract by oral route Group-V&VI will receive venom(25μg) i.m route followed by low and high dose of alcoholic extract by oral route.

Mice will be injected intramuscularly (i.m) in the right gastrocnemius with 2.5µg of venom in 50 μl of PBS and control animals will receive Phosphate buffer, test animals will receive aqueous and alcoholic extracts of C. zeylanicum. Animals will be bled on 1, 3, 6, 12 and 24 h later through the ophthalmic plexus under light ether anaesthesia. Sera will be collected after centrifugation and immediately assayed for creatine kinase activity by using biochemical kit(Span Diagnostics Ltd, India). One unit corresponds to the amount of enzyme that hydrolyses 1 μm of creatine per min at 250C. Enzyme activity is expressed in U/l. 25

Groups:

Group I

Control (Phosphate buffer) (i.m)

Group II

Venom(i.m)

Group III

Venom(i.m) + Aqueous Extract of C.zeylanicum (Low dose) (p.o)

Group IV

Venom(i.m) + Aqueous Extract of C.zeylanicum (High dose) (p.o)

Group V

Venom(i.m) + Alcoholic Extract of C.zeylanicum (Low dose) (p.o)

Group VI

Venom(i.m) + Alcoholic Extract of C.zeylanicum (High dose) (p.o)



4.Neutralization of Phospholipase A2 activity

Phospholipase activity (PLA2) will be measured by using an indirect haemolytic assay. Increasing concentration of venom in 15µl of saline will be added to 3mm wells in agarose gels (0.8% in phosphate-buffered saline, pH 8.1) containing 1.2% sheep erythrocytes, 1.2% egg yolk and 10mM CaCl2. Plates will be incubated at 370C overnight (12-16 h) and the diameters of the haemolytic haloes will be measured. Control disc contains only 15 µl of saline and standard disc contains only 15 µl of polyvalent antivenom. One unit corresponds to a concentration of venom which produce a haemolytic halo of 11 mm diameter. Activity was expressed as U/μg.26
5.Anti-inflammatory activity of extracts against inflammatory reaction induced by N.kaouthia venom

36 Swiss Albino mice weighing (18 – 20 gm) will be divided into 6 groups each containing 6 mice. Group-I will receive N.kaouthia venom (2.5μg) 27 by intraplantar route , Group-II will receive venom (2.5μg) by intraplantar route and dexamethasone (1mg/kg) as anti-inflammatory control, Group-III & IV will receive venom(2.5μg) intraplantar route followed by low and high dose of aqueous extract by intraplantar or oral route Group-V&VI will receive venom(2.5μg) intraplantar route followed by low and high dose of alcoholic extract by intraplantar or oral route.

Fifty microliters of sterile saline containing 2.5μg of N.kaouthia venom for venom control group will be injected in the subplantar region of the right hind paw. For anti-inflammatory control control group 2.5μg of N.kaouthia venom followed by dexamethasone (1mg/kg) in the subplantar region of the right hind paw. Group III,IV,V and VI mice will receive 2.5μg of N.kaouthia venom followed by extracts of C.zeylanicum in the subplantar region of the right hind paw or by oral route. The left hind paw received an equal volume of sterile saline alone and served as control. The volumes of both hind paws were measured by plethysmometry before and 15, 30 minutes, 1, 2, 4 and 6 hours after venom or myotoxins administration. The edema was expressed as the percentage increase in the volume of the treated (right) paw relative to that of control (left) paw at each time interval.28


Group I

Venom (intraplanter) 2.5μg

Group II

Venom (intraplanter) 2.5μg + dexamethasone (1mg/kg) (intraplanter)

Group III

Venom (intraplanter) 2.5μg + Aqueous Extract of C.zeylanicum (Low dose) (intraplantar)(p.o)

Group IV

Venom (intraplanter) 2.5μg + Aqueous Extract of C.zeylanicum (High dose) (intraplantar)(p.o)

Group V

Venom (intraplanter) 2.5μg + Alcoholic Extract of C.zeylanicum (low dose) (intraplantar) (p.o)

Group VI

Venom (intraplanter) 2.5μg + Alcoholic Extract of C.zeylanicum (High dose) (intraplantar) (p.o)



Statistical analysis

The data obtained from experimentation will be subjected to one way Analysis Of Variance (ANOVA) with suitable post-hoc test.




Total No. of animals required

No. of mice required for Acute oral toxicity test - 12 mice

No. of mice in each group - 6 mice

Total no. of animals used =120 mice


7.4 DOES THE STUDY REQUIRE ANY INVESTIGATION OR INTERVENTIONS

TO BE CONDUCTED ON PATIENTS OR OTHER HUMANS /ANIMALS? IF SO

PLEASE DESCRIBED BRIEFLY.

Yes, the above study requires investigation to be done on the Swiss Albino mice for the determination of antivenom activity.


7.5 HAS ANIMAL ETHICAL COMMITTEE CLEARANCE BEEN OBTAINED

FROM YOUR INSTITUTION IN CASE?

YES, The study has been referred to the ethical committee of the institution and clearance has been obtained.



LIST OF REFERENCE:

1. Brunda G, Sashidhar RB, Sarin RK. Use of egg yolk antibody (IgY) as an immunoanalytical tool in the detection of Indian cobra (Naja naja naja) venom in biological samples of forensic origin. Toxicon. 2006;48:183-94


2. Gomes A, Pallabi D, Dasgupta SC. Occurrence of a unique protein toxin from the Indian King Cobra (Ophiophagus hannah) venom. Toxicon. 2001;39:363-70.
3. Mukherjee AK, Maity CR. Biochemical composition, lethality and pathophysiology of venom from two cobras Naja naja and N. kaouthia. Comp Biochem and Physiol Part B 2002;131:125-32.
4. Shashidharamurthy R, Kemparaju K. Region-specific neutralization of Indian cobra (Naja naja) venom by polyclonal antibody raised against the eastern regional venom: A comparative study of the venoms from three different geographical distributions. Int Immunopharmacol 2007;7:61–9.
5. Clayton ZO, Victor AM, Silvana M, Carolina DS, Ana HJ, Miriam VL et al. Anticoagulant and antifibrinogenolytic properties of the aqueous extract from Bauhinia forficata against snake venoms. J Ethnopharmacol 2005;98:213–6.
6. Gomes A, Das R, Sarkhel S, Mishra R, Mukherjee S, Bhattacharya S, et al. Herbs and herbal constituents active against snake bite. Indian J Exp Biol. 2010;48(9):865-78.
7. Kasem K, Wanpen C, Yuwaporn S, Pongsri T, Praman T. Proteome and immunome of the venom of the Thai cobra, Naja kaouthia. Toxicon 2007;49: 1026–41.
8. Regis G, Jean-Pierre R, Herve R. Anticholinesterases and experimental envenomation by Naja. Comp. Biochem. Physiol 1994;109C:265-8.

9. Monimala M, Mukherjee AK. Neutralisation of lethality, myotoxicity and toxic enzymes of Naja kaouthia venom by Mimosa pudica root extracts. J Ethnopharmacol. 2001;75:55-60.

10. Marielga R, Francine G, Maria A, Cruz-Hofling., Marcos D. Neurotoxic and myotoxic actions of Naja naja kaouthia venomon skeletal muscle in vitro. Toxicon. 2003;41:657-65.
11. Yang. Y-C, Lee. H-S, Lee. SH, Clark. JM, Ahn. Y-J. Ovicidal and adulticidal activities of Cinnamomum zeylanicum bark essential oil compounds and related compounds against Pediculus humanus capitis (Anoplura: Pediculicidae). Int J Parasitol. 2005;35:1595-600.
12. Jayaprakasha GK, Negi PS, Jena BS, Jagan Mohan Rao L. Antioxidant and antimutagenic activities of Cinnamomum zeylanicum fruit extracts. J Food Compos Anal. 2007;20:330–6.
13. Subash Babu P, Prabuseenivasan S, Ignacimuthu S. Cinnamaldehyde-A potential antidiabetic agent. Phytomedicine. 2007;14:15-22.
14. Tsuji-Naito K. Aldehydic components of Cinnamon bark extract suppresses RANKL-induced osteoclastogenesis through NFATc1 downregulation. Bioorg Med Chem. 2008;16:9176-83.
15. Mehmet U, Emel E, Gulhan VU, Hulya SZ, Nilufer V. Composition, antimicrobial activity and in vitro cytotoxicity of essential oil from Cinnamomum zeylanicum Blume (Lauraceae). Food Chem Toxicol. 2010;48:3274-80.
16. Sathishkumar M, Sneha K, Seob Kwak I, Mao J, Tripathy SJ, Yun Y-S. Phyto-crystallization of palladium through reduction process using Cinnamom zeylanicum bark extract. J Hazard Mater. 2009;171:400-4.
17. Alam MI, Gomes A. Snake venom neutralization by Indian medicinal plants (Vitex negundo and Emblica officinalis) root extracts. J Ethnopharmacol 2003;86:75-80.
18. Ipshita C, Chakravarty AK, Gomes A. Daboia russellii and Naja kaouthia venom neutralization by lupeol acetate isolated from the root extract of Indian sarsaparilla Hemidesmus indicus R.Br. J Ethnopharmacol. 2006;106:38-43.

19. Gomes A, Archita S, Ipshita C, Chakravarty AK. Viper and cobra venom neutralization by b-sitosterol and stigmasterol isolated from the root extract of Pluchea indica Less.(Asteraceae). Phytomedicine. 2007;14:637-43.


20. KOKATE CK, PUROHIT AP, GOKHALE SB. Pharmacognosy. 22 ed: Nirali Prakashan; 2003.
21. Khandelwals KR. Practical Pharmacognosy- Techniques and Experiments. Pune: Nirali Prakashan; 1996.
22. 423 Acute Oral toxicity - Acute Toxic Class Method (Updated Guideline,adopted 17th December 2001)

Guidelinehttp://www.oecd-ilibrary.org/environment/test-no-423-acute-oral-toxicity-acute-toxic-class-method_9789264071001-en (cited on 9-12-2011).

23. Makarand G, Mukund H. Unpublished work :Antivenom activity of Amaranthus spinosus root extracts against Naja kaouthia snake venom Dissertation work Rajiv Gandhi University of Health Science, Bangaluru; May 2010:53

24. Fernando C, Gilbert DL, Adriana S, Jose MG. Intramuscular administration of antivenoms in experimental envenomation by Bothrops asper: comparison between Fab and IgG. Toxicon 2003;41:237–44.

25. Moraes FV, Sousa-e-Silva MCC, Barbaro KC, Leitao MA, Furtado MFD. Biological and immunochemical characterization of Micrurus altirostris venom and serum neutralization of its toxic activities. Toxicon 2003;41:71–9.

26. Kyoko UC, David T, Velarde, Eladio FS. Pharmacological characterization of the venoms used in the production of Bothropic antivenom in Brazil. Toxicon 2002;40:501-9.

27. Yingprasertchai S, Bunyasrisawat S, Ratanabanangkoon K. Hyaluronidase inhibitors (sodium cromoglycate and sodium auro-thiomalate) reduce the local tissue damage and prolong the survival time of mice injected with Naja kaouthia and Calloselasma rhodostoma venoms. Toxicon. 2003;42:635-46.
28. Pereira I, Barbosa A, Salvador M, Soares A, Ribeiro W, Cogo J, et al. Anti-inflammatory activity of Blutaparon portulacoides ethanolic extract against the inflammatory reaction induced by Bothrops jararacussu venom and isolated myotoxins BthTX-I and II. J Venom Anim Toxins incl Trop Dis. 2009;15(3):527-45










9.

Signature of the candidate

(Patel Mitul N)





10

Remarks of the Guide:


Recommended and forwarded





11

Name and Designation of:

11.1 Guide



Mr. Mukund Handral

Asst. Professor

Dept. Pharmacology

PES college of pharmacy

Bangaluru-560050




11.2 Signature









11.3 Co-Guide

11.4 Signature



NOT APPLICABLE


11.5 Head of the Department




Mr. R.SRINATH.

M. Pharm.

HOD& ASST PROFESSOR

DEPT. OF PHARMACOLOGY

P.E.S. COLLEGE OF PHARMACY




11.6 Signature of HOD







12

12.1 Principal





Prof. Dr. S. Mohan

Principal & Director

PES college of pharmacy

Bangaluru-560050




12.2 Remark of the principal




Recommended and forwarded




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