B.pharm 7 Semester Project ReportFinal

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PROJECT WORK SUBMITTED TO ASSAM SCIENCE AND TECHNOLOGY UNIVERSITY, GUWAHATI, ASSAM FOR THE PARTIAL FULFILLMENT OF THE REQUIREMENT FOR THE AWARD OF DEGREE OF BACHELOR OF PHARMACY Under the guidance of Mr. Biswajit Dash (M.Pharm) Assistant Professor, Dept. of Pharmaceutical Chemistry (GIPS) SUBMITTED BY UTTAM BORAH Regd. No. 097805514 of 2014-15 Roll No. 1405511005 Sub Code- PY1327210 GIRIJANANDA CHOWDHURY INSTITUTE OF PHARMACEUTICAL SCIENCES, HATHKHOWAPARA, AZARA, GUWAHATI-781017 KAMRUP, ASSAM (INDIA)

Transcript of B.pharm 7 Semester Project ReportFinal

Page 1: B.pharm 7 Semester Project ReportFinal

PROJECT WORK SUBMITTED TO ASSAM SCIENCE AND TECHNOLOGY

UNIVERSITY, GUWAHATI, ASSAM FOR THE PARTIAL FULFILLMENT OF

THE REQUIREMENT FOR THE AWARD OF DEGREE OF

BACHELOR OF PHARMACY

Under the guidance of Mr. Biswajit Dash (M.Pharm)

Assistant Professor, Dept. of Pharmaceutical Chemistry (GIPS)

SUBMITTED BY

UTTAM BORAH

Regd. No. 097805514 of 2014-15

Roll No. 1405511005

Sub Code- PY1327210

GIRIJANANDA CHOWDHURY INSTITUTE OF

PHARMACEUTICAL SCIENCES, HATHKHOWAPARA, AZARA,

GUWAHATI-781017 KAMRUP, ASSAM (INDIA)

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PREFACE

Pharmacy is a profession which is concerned with the art and science of suitable and

convenient material for distribution and use in the treatment and prevention of disease, so it

is a fully technical profession where practical knowledge is much more important along with

theoretical knowledge. So being a Pharmacy student it is my pleasure and privilege to submit

this dissertation in its present form. The progress of scientific research has been increasing

sustainability in the last two decades particularly in the field of pharmacy. The ultimate

biochemical objective of all these are remain same, to develop better drugs for a specific

disease. Here I have tried to find out the protective effects of antimicrobial drugs.

In this literature review I have written an introduction part where the details about the

herbal drug with its advantage, disadvantage, and definition of antimicrobial activity is

summarized. In this literature review I have shown the reason why I have performed this

project work. The possible literature regarding Leucas plukenetii (Roth)Spreng are written

in literature review section. The reference of my work is given separately.

At last I expect this assignment can give some guidelines about further Phytochemical

and Pharmacological research.

UTTAM BORAH

7th SEMESTER

Bachelor of Pharmacy (GIPS)

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CERTIFICATE

This is to certify that the topic incorporated in this Project work entitled

“PRELIMINARY PHYTOCHEMICAL SCREENING & ANTIMICROBIAL

ACTIVITY OF ETHANOLIC EXTRACT OF WHOLE AERIAL PART OF THE

HERB Leucas plukenetii (Roth) Spreng” being submitted by UTTAM BORAH Roll No-

1405511005 Reg. No- 097805514 in the partial fulfilment of the requirement of the award

of degree of Bachelor in Pharmacy (B. Pharm) of Girijananda Chowdhury Institute of

Pharmaceutical Science, Guwahati, Assam is a benefited assignment which has been

carried out my direct supervision and guidance during the academic Session 2016-17.

Supervised by Verified By

Mr Biswajit Dash (Principal, GIPS)

Asstt. Professor, GIPS

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ACKNOWLEDGEMENT

“Karma-yoga is a supreme secret indeed”

- Srimadbhagwadgita

I would first like to thank to ‘LORD NARAYANA’ & “GURU MADHABDEV”

for who’s blessing only this literature review cum research work was possible.

I consider it a great privilege & honor to have had the opportunity to undergo the

Project work in Girijananda Chowdhury Institute of Pharmaceutical Science, Guwahati,

Assam. Hence, I would like to offer my heartiest thanks to Prof (Dr). Suvakanta Dash,

Principal of GIPS.

I am greatly indebted to Mr Biswajit Dash ; Assistant Professor, Department of

Pharmaceutical Chemistry, GIPS, for his valuable guide and suggestion.

I owe a dept of gratitude to Dr P.P. Baruah , HOD; Department of Botany, Guwahati

University for authenticate my specimen.

I convey my heartiest thanks to Dr Damiki Laloo , Assistant Professor, Department

of Pharmacognosy, GIPS for most valuable suggestions, constant encouragement, and

affectionate guidance in and Mr. Diganta Das (Librarian), GIPS for their support for

carrying out this projects.

At last, I am greatly thankful to my friends in GIPS for extending their constant

cooperation which went a long way towards the completion of this Literature review cum

research work.

UTTAM BORAH

7th SEMESTER

Bachelor of Pharmacy (GIPS)

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DEDICATED TO MY BELOVED

PARENTS….

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CONTENTS

1. AIM AND OBJECTIVES 1-2

2. INTRODUCTION 3-6

3. LITERATURE REVIEW 7-12

4. PLAN OF WORK 13-15

5. BIBLIOGRAPHY 16-19

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Page 1

CHAPTER-1

AIM & OBJECTIVES

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CHAPTER 1 AIM & OBJECTIVES 2016

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1.1. AIM:

Antimicrobial resistance is a growing problem due to overuse of antibiotics in

humans. Many of the currently used antibacterial are associated with adverse effects such as

toxicity, hypersensitivity, immune suppression, and tissue residues posing public health

hazard. As the global scenario is now changing towards the use of non- toxic and eco-friendly

products, development of modern drugs from traditional medicinal plants should be

emphasized for the control of various human and animal diseases. In the present study, the

selection of this plant for evaluation was based on its traditional usages.

So further new alternative remedies for treatment of bacterial diseases are usually

required. Although very few works have been done on the antimicrobial activity of his

medicinal plant, it needs further study for verification of its activity against disease causing

microorganisms. Thus the aim of this project work is the evaluation of the antimicrobial

potency of Leucas plukenetii (F-Laminaceae)

1.2. OBJECTIVES:

The present objective of the study is-

Extraction of active constituents from the dried leaves of Leucas plukenetii

Preliminary Phytochemical screening of the extract of leaves of Leucas plukenetii

Antimicrobial activity of the extract in comparison to suitable standard.

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Page 3

CHAPTER-2

INTRODUCTION

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CHAPTER 2 INTRODUCTION 2016

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2.1. INTRODUCTION:

The medicinal use of plants is very old. The writings indicate that therapeutic use of plants

is as old as 4000–5000 B.C. and Chinese used first the natural herbal preparations as

medicines. In India, however, earliest references of use of plants as medicine appear in

Rigveda, which is said to be written between 1600-3500 B.C.

Later the properties and therapeutic uses of medicinal plants were studied in detail and

recorded empirically by the ancient physicians in ‘Ayurveda’ which is a basic foundation of

ancient medical science in India [1]. The Hindu Physician, Susruta enlisted seven hundred

and sixty medicinal plant in Ayurveda in the 5th century AD. [2]

Antibiotic resistance is a growing problem. The widespread and indiscriminate use

of antibacterial agents resulted in development of drug resistance among many virulently

pathogenic bacteria species. Many of the currently used antibacterial are associated with

adverse effects such as toxicity, hypersensitivity, immune suppression, and tissue residues

posing public health hazard.

So, further new alternative remedies for treatment of bacterial diseases are usually

required. As the global scenario is now changing towards the use of non- toxic and eco-

friendly products, development of modern drugs from traditional medicinal plants should be

emphasized for the control of various human and animal diseases. [3]

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CHAPTER 2 INTRODUCTION 2016

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2.2. NATURAL SOURCES OF ANTOMICROBIAL AGENTS(AMA).

Sl

No

Plant Scientific Name Major components

1 Allspice Pimenta dioica Eugenol, methyl ether cineol

2 Basil Ocimum asilicum d-linalool, methyl chavicol,

geraniol

3 Black Pepper Pipper nigrum Monoterpenes, Sesquiterpenes

4 Caraway

Seed

Carum carvi Carvone, limonene

5 Celery Seed Apium graveolens d-limonene

6 Cinamon Cinamonum

zeylanicum

Cinnamic aldehyde, l-linalool,

p-cymene, eugenol

7 Clove Syzygium aromaticum Eugenol, cariofilene

8 Coriander Coriandum sativum d-linalool, d-pinene,

9 Cumin Cuminum cyminum Cumin aldehyde, p-cymene

10 Fennel Foeniculum vulgare Anethole

11 Garlic Allium sativum Diallyl disulphide, diethyl

sulphide, allicin

12 Lemongrass Cymbopogon citratus Citral, geraniol

13 Marjoram Origanum majorana Linalool, cineol, methyl

chavicol, eugenol, terpininecol

14 Mustard Brassica hirta Allyl-isothiocynanate

15 Onion Allium cepa d-n-propyl disulphide, methyl-n-

propyl disulphide

16 Oregano Origanum vulgare Thymol, carvacrol, p-cymene

17 Parsley Pertroselinum

crispum

Pinene, fenol-eter-apiol

18 Rosemary Rosmarinus officinalis Borneol, cineol, borneol,

thymol, eugenol

19 Sage Salvia officinalis Thujone, cineol, borneol,

thymol, eugenol

20 Tarragon Artemissia

dracunculus

Methyl chavicol, anethode

21 Thyme Thymus Vulgaris Thymol, carvacol, geraniol, p-

cymene

22 Vanilla Vanilla planifolia Vanillin, Vanillic, p-

hydroxybenzoic, p-cumaric

acids

Fig-1: Sources of Natural AMA [4]

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2.3.PLANT PROFILE

2.3.1. BIOLOGICAL SOURCE: It consisted of the dried whole herb of Leucas

Plukenetii belonging to the family Lamiaceace.

2.3.2 PLANT DESCRIPTION-

Habit: An annual herb with square stem.

Leaves : Linear-oblong or oblong lanceolate.

Flowers : Sessile, ciliate with long hairs.

Fruit: Oblong, inner face sharply angular

.Flowering and Fruiting Time : July-October

Plant Form : Herb [5]

2.3.3 VERNICULAR NAME- Doron Bon [6]

2.3.4 SYNONYM- Leucas aspera [7]

Fig-2. Leucas plukeneti

2.3.5 ETHNOMEDICINAL USES OF LEUCAS PLUKENETII-

The leaves and flowers are used by the Kavirajes to treat tooth infections and mucus.

[8]

It is applied on inflamed parts to relieve pain and inflammation. [9]

Leave paste made with lime juice applied externally in the treatment of Headache.

Flowers were macerated and extract put dropwise into opposite side of nostril to

reduce migraine.

Leaves with pepper and garlic chewed and spilt into the nostril with force in case of

SNAKE BITE.

Whole plant dugout early in the morning and made into paste with water to treat

Wounds and worms. [10]

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Page 7

CHAPTER-3

LITERATURE

REVIEW

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CHAPTER 3 LITEARATURE REVIEW 2016

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3.1. PHYTO-CHEMICAL EVALUATION:

3.1.1. Sadhu et al. (2003) isolated eight lignans namely nectandrin B , (-)-chicanine ,

meso-dihydroguaiaretic acid, macelignan , myristargenol B , erythro-2-(4-allyl-2,6-

dimethoxyphenoxy)-1-(4-hydroxy-3-methoxy phenyl) propan-1-ol, Machilin C, (7R,8R)-

and (7S,8S)-licarin from the methanol extract of the whole plant.

3.1.2. Manivannana and Sukumar (2007) reported free flavonoid ‘baicalein’ was reported in

the ethereal fraction of hydro methanolic extract of flower.

3.1.3. Gerige et al. 2007 & Mangathayaru et al. (2006) found to contain high amount of a-

farnesene, α-thujene and menthol.

3.1.4. A new type of diterpenes, leucasperones A and B ; leucasperols A and B, have been

reported by Sadhu et al. (2006).

3.1.5. Linoleic acid were found which was contingent upon crop variation by Chen et al.,

(1979).

3.1.6. Rajyalakshmi et al., (2001) found Leucas aspera and Leucas cephalotes contain

significant amounts of total carotenoid and β-carotene. [11]

3.2. PHARMACOLOGICAL ACTIVITY:

3.2.1 Antimicrobial activity: Mangathayaru et al., (2005) reported significant antimicrobial

activity was reported for the alkaloidal fraction and the total methanol extract the

flowers [12].

Satyal et al., (2013) reveals that the essential oil of the plant showed no activity

against E. coli, P. aeruginosa, and C. albicans (MIC ≥ 1250μg/mL). The oil did exhibit good

activity against S. aureus (MIC = 625 μg/mL), B. cereus (MIC = 313μg/mL), and A. niger

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(MIC = 313 μg/mL), most likely attributable to the sesquiterpenes present in the oil. Both

(E)-caryophyllene and α-humulene have shown antibacterial activity against B. cereus and

S. aureus, and α-humulene was antifungal to A. niger [13]

Ilango et al., (2008) evaluated and found that Ethyl acetate extract (EAE) of whole

plant exhibited moderate to significant and concentration dependent antibacterial activity

against all the tested microorganisms at the concentrations of 50, 100, 200, 300 and 400

µg/disc and comparable to the various antibiotics used for individual microorganism. This

study also reveals that EAE was found to be highly active against Staphylococcus

epidermidis and Klebsiella pneumoniae [14]

Chew et al. (2012) evaluated the antimicrobial activity of crude extracts of root,

flower, leaf having notable antibacterial activity. The root extract showed the highest mean

zone of inhibition ranging from 9.0–11.0 mm at a concentration of 100 mg/mL.[15]

Antony et al. (2013) describes green synthesis of silver nanoparticles (AgNPs)

utilizing the plant. Antimicrobial activity of the AgNPs was tested against Aeromonas

hydrophila. Catla catla, the model organism used for the experiment was divided into six

groups with 15 animals in each group. In vivo analysis of biochemical parameters and

histological architecture provided evidence for the antibacterial effect of AgNPs in the fish

model.[16]

3.2.2 Central nervous system activity: Rahman et al. (2007) found that ethanolic extract of

root showed significant peripheral antinociceptive activity at a dose of 400 mg/kg . [17]

3.2.3 Antioxidant activity: Rahman et al. (2007) reported significant activity was found in

the ethanolic extract of root (IC50 = 7.5 µg/ml) .[17]

Chew et al. (2012) evaluated antioxidant activity methanol extract of root and it

possessed antioxidant activity near the range of vitamin E and thus could be a potential rich

source of natural antioxidant.[15]

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3.2.4 Hepatoprotective activity: According to Thenmozhi et al (2013) the Hydroalcoholic

leaf extract has found to possess hepatoprotective property by suppressing the abnormal

elevation of liver enzymes induced by lead acetate . [18]

Mangathayaru et al.,(2005) showed The cold methanolic extract of the whole plant

of was found to exhibit significant hepato protection in CCl4 induced liver damage.[19]

Banu et al. (2012) found that d-galactosamin administration induced hepatotoxicity

in rats which was manifested by increased levels of alanine aminotransferase, aspartate

aminotransferase, alkaline phosphatase, total cholesterol, triglycerides, total bilirubin and

oxidative stress. Pre-treatment with LA extract significantly protected the liver in d-

galactosamin administered rats. LA extract significantly elevated antioxidant enzymes like

superoxide dismutase, catalase, glutathione peroxidase and decreased lipid peroxidation

levels in liver. The total phenolic and flavonoid content in LA aqueous extract was found to

be 28.33 ± 0.19 gallic acid equivalents mg/g of extract and 3.96 ± 0.57 rutin equivalent mg/g

of extract, respectively. LA extract (200 and 400 mg/Kg) treatment with CCl4 decreased the

hexobarbitone-induced sleeping time in mice by 56.67 and 71.30%, respectively, which

indicated the protective effect of LA on hepatic MDMEs. Histological studies showed that

LA at 400 mg/kg attenuated the hepatocellular necrosis in d-galactosamin intoxicated

rats.[20]

3.2.5 Anti-inflammatory activity: Reddy et al. (1986) reported that whole plant extract

have anti-inflammatory activity and caused degranulation of mast cells [17].

Reddy et al. (1986) reported anti-inflammatory activity of the yellow colored

chromatographic fraction of extract was observed in the chronic and acute models of

inflammation. It was observed that, the activity was due to the inhibition of histamine and

serotonin [21].

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Similar observations were made by Saundane et al., (2000) and concluded that, on

preliminary screening of ethanol and dis­tilled water extracts exhibited significant anti-

inflammatory activity, whereas only ethanol (95%) extract produced long term analgesia in

the experimental animals. [22]

Srinivas et al. (2000) reported that the dried leaves of for the alcoholic and aqueous

extracts of the plant possessed significant anti-inflammatory activity against carrageenan-

induced paw oedema and cotton pellet induced granuloma. [23]

Goudgaon et al. (2003) concluded that the alkaloid fraction of the crude ethanolic

extract is accountable for the antiinflammatory activity. [24]

3.2.6 Cytotoxicity: Krishnaraju et al. (2005) proves that the hydro alcoholic extract of whole

plant exhibited cytotoxicity (LC50 = 1,900 µg/ml)[16] and this activity was more in the root

extract (LC50 = 52.8µg/ml) [25].

Chew AL et al. (2012) performed brine shrimp lethality bioassay , it was evident

that the methanol root extract did not show significant toxicity. The LC50 value for 12 h and

24 h observation was 2.890 mg/mL and 1.417 mg/mL, respectively. [15]

3.2.7 Larvicidal Activity: Maheswaram et al. (2008) The findings of the present

investigation revealed that L. aspera has good larvicidal activity of the plant activity against

Culex quinquefasciatus Say. and Aedes aegypti [26]

3.2.8 Antihyperglycemic Activity: Mannan et al. (2010) performed an experiment upon

antihyperglycemic activity of plant extract at mice. The methanol extract of leaves, when

administered to glucose-challenged mice demonstrated dose-dependent and significant

antihyperglycemic activity The stem extract also demonstrated significant and dose-

dependent antihyperglycemic activity in glucose tolerance tests in mice. [27]

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However, dose for the antihyperglycemic activity demonstrated by stem extract was

lower than that of leaf extract. At the highest dose of 400 mg extract per kg body weight,

serum glucose levels fell by 28.39% versus 34.01% obtained with the same dose of leaf

extract. To summarize, antihyperglycemic activity at a statistically significant level was

observed with leaf extract, when the extract was administered at doses of 200 mg per kg body

weight to glucose-loaded mice. [27]

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CHAPTER-4

PLAN OF WORK

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CHAPTER 4 PLAN OF WORK 2016

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Step-I

• Collection of Plant Material.

• Washing of plan material.

• Authentication of Plant Material by GU.

Step - II• Drying and Grinding of plant material.

Step-III

• Extraction of Plant Material (Maceration Process).

• Preliminary phytochemical screening of extract.

Step-IV• Determination of Antimicrobial Activity.

4. PLAN OF WORK

Fig 3-Plan of work

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CHAPTER 4 PLAN OF WORK 2016

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4.1. Collection of Plant Material: The fresh whole aerial parts of the plant Leucas

plukentii was collected from various parts of the District Lakhimpur, Assam.

4.2. Washing of Plant Material: The collected plant material was washed using water

and then it is sundried to remove moisture.

4.3. Authentication of Plant Material: Herbarium sheet was prepared using the cleaned

plant material. Then it was authenticating at Department of Botany, Gauhati University.

4.4. Drying and grinding of plant material: The cleaned plant material will be dried at

600c using Tray dryer and then it will be grinded to form powder.

4.5. Extraction of Plant Material: The sieved powder drug will be extracted by

maceration process using ethanol as solvent.

4.6. Preliminary phytochemical screening of extract: The Preliminary phytochemical

screening of extract will be done by tested the extract for glycosides, alkaloids etc.

4.7. Determination of Antimicrobial Activity: The potency of extract will be

determined by agar disc diffusion method against a standard.

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CHAPTER-5

BIBLIOGRAPHY

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CHAPTER 5 BIBLIOGRAPHY 2016

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BIBLIOGRAPHY

1. Prakash P and Gupta N. Therapeutic uses of Ocimum sanctum linn (tulsi) with a note

on eugenol and its pharmacological actions; a short review. Indian Journal of

Physiology and Pharmacology 2005; 49: 125–131.

2. Guthrie D.A. History of Medicine. London, Thomas Nelson and Sons Ltd : 1945;

p-33.

3. Das A. A study of antibacterial activity of ethanolic extracts and aqueous extracts of

Leucas longifolia (doron) leaves against Eschreiashia coli. International Research

Journal of Pharmacy 2012; 3: 130-132.

4. Michael P, Sofos J, and Branen A . Antimicrobials in Food, Tailor & Francis group;

Third Edition : 2005; p- 429-507.

5. Http://www.efloraofgandhinagar.in/herb/leucas-aspera [Accessed 21 October 2016]

6. Http://www.flowersofindia.net/catalog/slides/Common%20Leucas.html [Accessed

02 November 2016]

7. http://www.theplantlist.org/tpl1.1/record/kew-111857 [Accessed 02 November

2016]

8. Mohammed R, Das AK, Ariful Md., Haque M, Rownak J, Khan M, Rahman T, and

Chowdhury MH. An Ethnomedicinal Survey of Dhamrai Sub-district in Dhaka

District, Bangladesh. American-Eurasian Journal of Sustainable Agriculture 2009; 3:

881-888.

9. Rahmatullah Q, Bhatti R, and Rabia A. Ethnomedicinal uses of herbs from northern

part of nara desert,Pakistan. Pakistan Journal of Botany 2010; 42(2): 839-851.

10. Parinitha M, Harish GU, Vivek NC, Mahesh T, and Shivanna MB. Ethnobotanical

wealth of Bhadra Wild life sanctuary of Karnataka. Indian Journal of Traditional

knowledge 2004; 3(1): 37-50.

11. Singh SK, and Chowhan HS. A review of plants of genus Leucas. Journal of

Pharmacognosy and Phytotherapy 2011; 3: 13-23.

12. Mangathayaru K, Lakshmikant J, Sundar NS, Swapna R, Grace XF, and Vasantha J.

Antimicrobial activity of Leucas aspera flowers. Fitoterapia 2005; 76: 752-754.

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13. Satyal P, Paudel P, Poudel A, and Setzer WN. Microbiological activities of volatile

constituents of Leucas aspera (Willd). Nepal Journal of Natural Pharmaceuticals

2012 ; 3 : 118-119.

14. Ilangos k. Ramya s and Gopinath G. Antibacterial activity of Leucas aspera spreng.

International Journal of Chemical Sciences 2008; 6 :526-530.

15. Chew AL , Jessica JJA and Sasidharan S. Antioxidant and antibacterial activity of

different parts of Leucas aspera. Asian Pacific Journal of Tropical Biomedicine,

2012; 2: 176-180.

16. Antonya JJ, Nivedheethaa M, Sivaa D, Pradeephaa G, Kokilavania P, Kalaiselvia S,

Sankarganesha A, Balasundaramb A, Masilamanib V, and Achiramana S.

Antimicrobial activity of Leucas aspera engineered silver nanoparticles against

Aeromonas hydrophila in infected Catla catla. Colloids and Surfaces B:

Biointerfaces 2013; 109: 20-24.

17. Rahman MS, Sadhu SK, and Hasan CM. Preliminary antinociceptive, antioxidant and

cytotoxic activities of Leucas aspera root. Fitoterapia 2007; 78: 552-555.

18. Thenmozhi M, Dhanalakshmi M, Devi KM, Sushila K, and Thenmozhi S. Evaluation

of hepatoprotective activity of Leucas aspera hydroalcoholic leaf extract during

exposure to lead acetate in male albino wistar rats. Asian Journal of Pharmaceutical

and Clinical Research 2013; 6 : 78-81.

19. Mangathayaru K, Fatima GX, Bhavani M, Meignanam E, Karna SLR and Kumar

DP. Effect of Leucas aspera on hepatotoxicity in rats. Indian Journal of

Pharmacology 2005 ; 37 : 329-330.

20. http://www.tandfonline.com/doi/abs/10.3109/13880209.2012.685130

21. Redy MK, Viswanathan S, Sambantham PT, Ramachandran S, and Kameswaran L.

Effects of Leucas aspera on experimental inflammation and mast cell degranulation.

Ancient Science of Life 1986; 5: 168-171.

22. Saha K, Mukherjee PK, Mandal SC, Saha BP, and Pal M. Antiinflammatory

evaluation of Leucas lavandulaefolia Rees Extract. Natural Product Sciences 1997;

2: 119-122.

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23. Srinivas K, Rao MEB, and Rao SS. Anti-inflammatory activity of Heliotropium

indicum Linn. And Leucas aspera Spreng. in albino Rats. Indian Journal of

Pharmacology 2000; 32: 37-38.

24. Goudgaon NM, Basavaraj NR, and Vijayalaxmi A . Antiinflammatory activity of

different fractions of Leucas aspera spreng. Indian Journal of Pharmacology 2003;

35: 397-398.

25. Krishnaraju AV, Rao TVN, Sundararaju D, Vanisree M, Tsay HS, and Subbaraju

GV. Assessment of bioactivity of Indian medicinal plants using brine shrimp

(Artemia salina) lethality assay. International Journal of Applied Science and

Engineering 2005; 3: 125-134.

26. Maheswaran R, Sathish S, and S Ignacimuthu S. Larvicidal activity of Leucas aspera

(Willd) against the larvae of Culex quinquefasciatus Say. and Aedes aegypti L.

International Journal of Integrative Biology 2008; 2 : 214-217.

27. Mannan A, Das S, Rahman M, Jesmin J, Siddika A, Rahman M, Rahman S,

Chowdhury M and Rahmatullah M. Antihyperglycemic Activity Evaluation of

Leucas Aspera (Willd.) Link Leaf and Stem and Lannea Coromandelica (Houtt.)

Merr. Bark Extract in Mice. Advances in Natural and Applied Sciences 2010; 4(3):

385-388.