ANTI INFLAMMATORY PROSPECTIVE STUDY OF COMMIPHORA MUKUL (GUGGUL) ON WISTAR ALBINO RATS

4/27/2012KRISHNAMRAJU NALIMELA

ANTI INFLAMMATORY PROSPECTIVE STUDY OF ETHANOLIC HERBAL EXTRACT OF COMMIPHORA

MUKUL (GUGGUL) BARK BY ADOPTING CARRAGEENAN INDUCED PAW OEDEMA IN WISTAR ALBINO RATS

A Dissertation Submitted to theJAWAHARLAL NEHRU TECHNOLOGICAL UNIVERSITY

HYDERABAD.

In partial fulfillment of the requirements for the award of degree of

BACHELOR OF PHARMACYBy

N.KRISHNAM RAJU( 08Y41R0030 )

Under the guidance ofMr. V.S. GIRI PRASAD M.Pharm

Assistant Professor

R.G.R.SIDDHANTHI COLLEGE OF PHARMACYSECUNDRABAD-500034(2011-2012)

DECLARATION OF AUTHORSHIP

This research work on” Anti Inflammatory Prospective Study Of Herbal

Ethanolic Extract Of Commiphora Mukul (Guggul) Bark By Adopting Carrageenan

Induced Paw Oedema In Wistar Albino Rates “ is carried out by us in “RGR

SIDDHANTHI COLLEGE OF PHARMACY Secundrabad, Andhra Pradesh Submitted

to Jawaharlal Nehru Technological University, Hyderabad for the award of the degree

Bachelor of pharmacy. We solemnly declare that this thesis either in part or in full has not

been submitted to any other University or Institution for the award of any Degree or

Diploma.

KRISHNAM RAJU (08Y41R0030)

Date: 27-04-2012

Place: Secundrabad

Ref:

CERTIFICATE BY THE PRINCIPAL

This is to certify that the dissertation work entitled “Anti Inflammatory Prospective Study

Of Herbal Ethanolic Extract Of Commiphora Mukul (Guggul) Bark By Adopting

Carrageenan Induced Paw Oedema In Wistar Albino Rats” submitted to Jawaharlal

Nehru Technological University in partial fulfillment for the award of the degree of bachelor

of Pharmacy, has been successfully carried out by below mentioned students during the

academic year 2011-2012 under the guidance of Mr.V.S. GIRI PRASD

M.Pharm.Department of Pharmacology.

KRISHNAM RAJU (08Y41R0030)

Forwarded by

Mrs.Dr. K. VIJAYA, M.Pharm, PhD

Principal,

RGR Siddhanthi College of pharmacy

Secundrabad -500003.

Ref:

THESIS CERTIFICATE BY THE RESEARCH GUIDE

This is to certify that the dissertation work entitled “Anti Inflammatory Prospective Study

Of Herbal Ethanolic Extract Of Commiphora Mukul (Guggul) Bark By Adopting

Carrageenan Induced Paw Oedema In Wistar Albino Rats” submitted to Jawaharlal

Nehru Technological University in partial fulfillment for the award of the degree of bachelor

of Pharmacy, has been successfully carried out by below mentioned students during the

academic year 2011-2012 under the guidance of Mr.V.S. GIRI PRASD

M.Pharm.Department of Pharmacology.

KRISHNAM RAJU (08Y41R0030)

Forwarded by

Mr.V.S. GIRI PRASAD

Assistant Professor,

Department of pharmacology,

RGR Siddhanthi College of pharmacy

Secundrabad -500003.

BONAFIDE CERTIFICATE

This is to certify that the project titled Anti Inflammatory Prospective Study Of Herbal

Ethanolic Herbal Extract Of Commiphora Mukul (Guggul) Bark By Adopting

Carrageenan Induced Paw Oedema In Wistar Albino Rates is a bonafide record of the

work done by

KRISHNAM RAJU (08Y41R0030)

In partial fulfillment of the requirements for the award of the degree of Bachelor of

Pharmacy of the JAWAHARLAL NEHRU TECHNOLOGICAL UNIVERSITY,

HYDERABAD, during the year 2011-2012.

Project Viva-voce held on _____________________________

Internal Examiner External Examiner

Mr.V.S. GIRI PRASAD

Assistant Professor,

Department of pharmacology,

RGR Siddhanthi College of pharmacy

Secundrabad -500003

ACKNOWLEDGEMENT

I am thankful to the Almighty for blessings in successful completion of this

dissertation. The satisfaction that accompanies the successful completion of any task would

be incomplete without mention of the people who made it possible with constant guidance,

support and encouragement that crows all effort with success. I would like to express my

appreciation for all the efforts to everyone who have directly or indirectly contributed their

ideas and energies in successful completion of my project.

I take this golden opportunity to express my humble gratitude and respect to my

research guide Mr. V.S. GIRI PRASAD M.Pharm.  Asst.Professor, Department of

Pharmacology, RGR Siddhanthi College of pharmacy, A.P. Ranjit Kumar M.Pharm

Department of Pharmacology, RGR Siddhanthi College of pharmacy, for their

constant invaluable guidance, encouragement and support throughout the project work. I am

proud to say that it has been the most fruitful and enjoyable experience to work under their

untiring guidance. Scrupulousness discipline, principle, and provision of fearless work

environment will be cherished in all walks of my life.

I express my gratitude and sincere thanks to Principal Dr. K. Vijaya M.Pharm

PhD, RGR Siddhanthi College of pharmacy for providing facilities which enabled me to

complete this work successfully.

Then with deep gratitude I would like to thank Sri. P.L. Srinivas our beloved

secretary of RGR Siddhanthi institutions who gave me admission into B.Pharmacy course

there by making my dream of higher education possible.

KRISHNAM RAJU (08Y41R0030)

DEDICATION TO

We are lovingly dedicate this project to our PARENTS and my

research guide Mr. V.S. GIRI PRASAD M.Pharm. Asst.Professor,

Department of Pharmacology,

RGR Siddhanthi College of pharmacy.

ABBREVATIONS

COX : Cyclooxygenase

NSAID : Non-Steroidal Anti-Inflammatory Drug

ImSAIDs : Immune Selective Anti-Inflammatory Derivatives

WHO : World Health Organization

BSA : Bovine serum albumin

COX-1 : Cyclooxygenase 1

COX-2 : Cyclooxygenase 2

PGE : Prostaglandin E

UNIDO : United Nations Industrial Development Organization.

IL : Interleukin

RBC : Red Blood Corpuscle

WBC : White Blood Cells

CSM : committee on safety of medicines

Table of Contents

ABSTRACT............................................................................................................................................11

INTRODUCTION...................................................................................................................................13

TYPES OF INFLAMATION:.................................................................................................................14

1) Acute inflammation.............................................................................................................14

2) chronic inflammation:..........................................................................................................14

CAUSES :..........................................................................................................................................16

SYMPTOMS:.....................................................................................................................................16

INFLAMMATORY DISORDERS:..........................................................................................................17

THE MECHANISM OF ACTION OF ANTI-INFLAMMATORY DRUGS:...................................................17

ANTI INFLAMMATORY DRUGS:.......................................................................................................18

steroids........................................................................................................................................18

Classification of NSAIDs According to mechanism of action.....................................................18

Herbs :.........................................................................................................................................20

Adverse effects of NSAIDs............................................................................................................20

LITRERATURE REVIEW.........................................................................................................................22

PLANT PROFILE....................................................................................................................................26

INTRODUCTION...........................................................................................................................26

DESCRIPTION...................................................................................................................................27

CHEMICAL COMPOSITION / KEY ACTIVE CONSTITUENTS.......................................................27

REMEDIES FOR :..........................................................................................................................28

PHARMACOLOGY:......................................................................................................................28

Lipid-lowering effects:.................................................................................................................28

Antioxidant effects:..............................................................................................................29

Platelet effects:....................................................................................................................29

Anti-inflammatory:..............................................................................................................29

DOSAGE.........................................................................................................................................29

ADVERSE EFFECTS.....................................................................................................................29

USES & BENEFITS OF GUGGUL................................................................................................29

CAUTION.......................................................................................................................................30

AIMS AND OBJECTIVES.................................................................................................................31

AIM:................................................................................................................................................32

OBJECTIVES:................................................................................................................................32

MATERIALS AND METHODS.........................................................................................................34

MATERIALS:.................................................................................................................................34

Soxhlet extractor :........................................................................................................................34

Collection of plant materials :......................................................................................................35

Experimental animals:.................................................................................................................36

Inflammation inducers:................................................................................................................36

lab materials(plethysmo meter):..................................................................................................37

METHODS:.......................................................................................................................................37

Preparation of extracts :...............................................................................................................38

Extraction process:......................................................................................................................38

Liebermann – burchard test :.......................................................................................................40

Carrageenan-induced inflammation in Rats :..............................................................................40

Test for Anti inflammatory Activity:...........................................................................................41

EVALUATION OF ANTI-INFLAMMATORY ACTIVITY...........................................................................44

DISCUSSION:........................................................................................................................................47

CONCLUSION:......................................................................................................................................49

BIBLIOGRAPHY.....................................................................................................................................51

ABSTRACT

ABSTRACT

Guggul, the gum resin from Commiphora mukul, is one of the components of various

formulations of traditional Ayurvedic medicine to treat inflammation, obesity, and lipid

disorders. In most preparations of Ayurvedic medicine in India.

The Ethanolic extract of Guggul, consist of guggulusterone derivatives. The structures

of these compounds were confirmed by spectroscopic methods. These compounds were

assayed for cyclooxygenase (COX) enzyme inhibitory activities and inhibit the PGE

synthesis also. so these presence may be the cause in curing the inflammation

COX enzyme (COX-1 and COX-2) inhibitory activities exhibited by compounds

isolated from C. mukul may substantiate its use in traditional medicine.

Injection of carrageenan in the rat paw causes oedema. The increase in paw volume

was highest at 1hr after carrageenan injection.

Anti-inflammatory agents such as guggulusterone derivatives from Commiphora

mukul were administered orally at a daily dose of 200,400 and 600 mg/kg, respectively, for a

period of 2 hrs.

The significant difference was seen in rats treated with Ethanolic extract of

Commiphora mukul. The extract decreased the oedema of the rat paw during the course of

drug treatment. Anti-inflammatory activity of the Commiphora mukul was assessed by using

Plethysmograph.

These results indicate the beneficial role of gum Guggul in experimental anti

inflammatory activity.

INTRODUCTION

INTRODUCTION

Inflammation is the first response of the immune system to infection or irritation and

Inflammation a localized reaction that produces redness, warmth, swelling, and pain as a

result of infection, irritation, or injury. Inflammation can be external or internal.

 Inflammation is not a synonym for infection, even in cases where inflammation is caused by

infection. Although infection is caused by a microorganism, inflammation is one of the

responses of the organism to the pathogen. However it is considered as a mechanism

of innate immunity, as compared to adaptive immunity, which is specific for each pathogen.

TYPES OF INFLAMATION:

Inflammation can be classified as acute inflammation and chronic inflammation

1) Acute inflammation: can lead to chronic inflammation, but the latter can also exist

by itself. Acute forms that are common include:  headaches, stiff joints, back pain and

acne.  Acute inflammation will become chronic if the immune system is unable to rid

the body of the offending foreign agent or if the agent is constantly able to re-enter the

body. Acute inflammation is the initial tissue reaction to a wide range of injuries or

insults and may last from a few hours to a few days. The acute inflammatory response

is similar whatever the causative agent.

• Short term (minutesàdays)

• Exudation of fluid, plasma, proteins,and leukocytes (neutrophils).

• Phagocytosis and enzymatic release occurs.

• Activation of neutrophils and macrophages--digest foreign material--involves

recognition, attachment, engulfment, and degradation.

• Recognition and attachment is enhanced when serum factors, Opsonins present

immunoglobin G (Ig G), complement activated fragment C3b can adsorb to

biomaterials.

• Neutrophils and macrophages have receptors for these proteins.

2) chronic inflammation:The word 'chronic' applied to any process implies that the

process has extended over a long period of time. This is usually the case in chronic

inflammation, but here the term 'chronic' takes on a much more specific meaning, in that the

type of cellular reaction differs from that seen in acute inflammation. Chronic inflammation

may be defined as an inflammatory process in which lymphocytes, plasma cells and

macrophages predominate, and which is usually accompanied by the formation of granulation

tissue, resulting in fibrosis. Chronic inflammation is usually primary, but does occasionally

follow acute inflammation.

• Long term (≥≥ days).

• Characterized by the presence of macrophages, monocytes, and mononuclear cells

including lymphocytes and plasma cells.

• Accompanied by the proliferation of blood vessels and connective tissue.

• Lymphocytes and plasma cells are involved in the immune reactions- mediate

antibody production.

• Macrophages process and deliver antigen to immunocompetent cells— mediate

immune reactions.

Comparison between acute and chronic inflammation:

Acute Chronic

Causative agent Bacterial Pathogens, injured tissues

Persistent acute inflammation

due to non-degradable

pathogens,viral infection,

persistent foreign bodies, or

autoimmune reactions

Major cells involved

neutrophils (primarily), basophils

(inflammatory response), and

eosinophils (response to helminth

worms and parasites), mononuclear

cells (monocytes, macrophages)

Mononuclear cells

(monocytes, macrophages,

lymphocytes, plasma cells),

fibroblasts

Primary mediators Vasoactive amines, eicosanoids

IFN-γ and other cytokines,

growth factors, reactive

oxygen species, hydrolytic

enzymes

Onset Immediate Delayed

CAUSES :

Burns

Chemical irritants

Toxins

Infection by pathogens

Physical injury, or penetrating

Immune reactions due to hypersensitivity

Ionizing radiation

Foreign bodies, including splinters, dirt and debris

Trauma

SYMPTOMS:

Pain

Bruising

Redness

Swollen

Fever

Stuffy nose and head

Breathing problems (asthma)

Fluid retention

Blood clots.

INFLAMMATORY DISORDERS:

Examples of disorders associated with inflammation include:

Asthma

Autoimmune diseases

Glomerulo nephritis

Hypersensitivities

Inflammatory bowel diseases

Pelvic inflammatory disease

Rheumatoid arthritis

Transplant rejection

THE MECHANISM OF ACTION OF ANTI-INFLAMMATORY DRUGS:

When there is injury to any part of the body, the arterioles (minute blood vessels) in the

surrounding tissue dilate (widen). This allows an increased blood flow to the area

(redness). Vaso active substances also increase the permeability (increase pore size) of

these arterioles which allows blood cells, chemical mediators, blood proteins and fluid to

accumulate in the area. This fluid accumulation causes swelling and may compress nerves

in the area resulting in pain. Furthermore, the main chemical mediators of inflammation

like prostaglandins, which are produced by cells, may also cause ‘irritation’ of the nerves

and further contribute to pain.

Inflammation is caused by release of chemicals from tissues and migrating cells. Most

strongly implicated are the prostaglandins (PGs), leukotrienes (LTs), histamine,

bradykinin, and, more recently, platelet- activating factor (PAF) and interleukin-1.

ANTI INFLAMMATORY DRUGS:

steroids glucocorticoids

corticosteroids

Classification of NSAIDs According to mechanism of action

1) COX-1 selective inhibitors

    - Acetylsalicylic acid at low dosage

2) Non selective COX inhibitors

    - Acetylsalicylic acid at high dosage

    - Diclofenac

    - Ibuprofen

    - Indomethacin

    - Piroxicam

    - Naproxen

3) COX-2 selective inhibitors

    -Nimesulid

   - Meloxicam

   - Nabumeton

Therapeutic classification of NSAID s

A: Analgesics

Aspirin, paracetamol

B: Anti-inflammatory

Indomethacin, naproxen, ibuorofen

C: Anti-coagulants

Aspirin

D: Anti-pyretics

Aspirin, paracetamol, indomethacin, celecoxicv, ibuprofen

E: Anti-cancer drugs

Methotraxate

F: Anti-malarial

Chloroquine, hydroxychloroquine

G: Tissue transplantation

Cyclosporine

H: Anti-gout drugs

Indomethacin, ibuprofen

Herbs : Harpagophytum ,

hyssop ,

ginger ,

turmeric ,

Arnica montana

Adverse effects of NSAIDs

Gastrointestinal effects: abdominal pain, gastric and duodenal ulcer, diarrhea,

pancreatis

Gastrointestinal hemorrhage, hepatotoxicity

Renal effect

Disturbances of renal function with water and sodium retention

Inhibition of platelet aggregation

central symptoms: headache, decreased hearing, tinnitus, dizziness, confusion,

depression

Allergic reactions: asthma, rashes, photosensitivity

LITRERATURE REVIEW

LITRERATURE REVIEW

1. Department of Pharmacology and University of Pittsburgh Cancer Institute on z-

Guggulsterone, a constituent of Ayurvedic medicinal plant Commiphora mukul,

inhibits angiogenesis in vitro and in vivo:

Our previous studies have shown that z- guggulusterone, a constituent of Indian Ayurvedic

medicinal plant Commiphora mukul, inhibits the growth of human prostate cancer cells by

causing apoptosis. We now report a novel response to z-guggulusterone involving the

inhibition of angiogenesis in vitro and in vivo. The z-guggulusterone treatment inhibited

capillary-like tube formation (in vitro neovascularization) by human umbilical vein

endothelial cells (HUVEC) and migration by HUVEC and DU145 human prostate cancer

cells in a concentration- and time-dependent manner. The z- and E-isomers of

guggulusterone seemed equipotent as inhibitors of HUVEC tube formation. The z-

guggulusterone mediated inhibition of angiogenesis in vitro correlated with the suppression

of secretion of proangiogenic growth factors, down-regulation of VEGF receptor 2 (VEGF-

R2) protein level, and inactivation of Akt. The z-guggulusterone–mediated suppression of

DU145 cell migration was increased by knockdown of VEGF-R2 protein level. Ectopic

expression of constitutively active Akt in DU145 cells conferred protection against z-

guggulusterone mediated inhibition of cell migration. Oral gavage of 1 mg z-

guggulusterone (five times/wk) to male nude mice inhibited in vivo angiogenesis in DU145-

Matrigel plug assay as evidenced by a statistically significant decrease in tumor burden,

microvessel area (staining for angiogenic markers factor VIII and CD31), and VEGF-R2

protein expression. In conclusion, the present study reveals that z-guggulusterone inhibits

angiogenesis by suppressing the VEGF–VEGF-R2–Akt signaling axis. Together, our results

provide compelling rationale for further preclinical and clinical investigation of z-

guggulusterone for its efficacy against prostate cancer.

2. Mutsuhiro Ikuma, MD, Ph.D., FACP, AGAF, First Department of Medicine,

Hamamatsu University School of Medicine on Guggulusterone Suppresses Bile Acid-

induced and Constitutive Caudal-related Homeobox 2 Expression in Gut-derived

Adenocarcinoma Cells:

Guggulusterone, a plant polyphenol guggulipid, has several antitumour effects and acts as an antagonist for the farnesoid X receptor. Although bile acids induce caudal-related homeobox 2 (CdX2), a transcription factor essential for intestinal development and gut tumourigenesis, the effects of guggulusterone on regulation of CdX2 in the gut are unknown. Materials and Methods: Regulation of CdX2 expression by treatment with bile acids and/or guggulsterone was analysed by immunoblot analysis in human gut-derived adenocarcinoma, Bic-1 cells.

Nuclear factor-κB (NF-κB) activity and the cell cycle distribution were also examined. Results: Chenodeoxycholic acid and deoxycholic acid increased CdX2 expression in Bic-1 cells. Guggulsterone reduced bile acid-induced and constitutive CdX2 expression at 5μM. Guggulsterone (up to 5μM) did not affect cell viability or the cell cycle and did not attenuate bile acid-induced or constitutive NF-κB activation. Conclusion: Guggulsterone may be used as a novel drug to target CdX2 expression in certain gut adenocarcinomas.

3. lipid Lowering Activity of Guggulsterone from Commiphora mukul in

Hyperlipaemic Rats by Ramesh Chander, A. K. Khanna, N. K. Kapoor

The lipid lowering action of guggulusterone, the active constituent of guggulipid, has been

studied in triton and cholesterol fed hyperlipaemic rats. Serum lipids were found to be

lowered by guggulsterone (50 mg/kg, b.w.) in triton WR-1339 induced hyperlipaemia.

Chronic feeding of this drug (5 mg/kg, b.w.) in animals simultaneously fed with cholesterol

(25 mg/kg, b.w.) for 30 days, caused lowering in the lipid and apoprotein levels of very low

density and low density lipoproteins in experimental animals. Guggulsterone activates

lipolytic enzymes in plasma and liver as well as stimulated receptor mediated catabolism of

low density lipoprotein. The hypolipidaemic activity of this drug is mediated through

inhibition of hepatic cholesterol biosynthesis, increased faecal bile acid excretion and

enhanced plasma lecithin: cholesterol acyltransferase activity

4 .Rhabdomyolysis Caused by Commiphora mukul, a Natural Lipid-Lowering Agent by

Antonio Bianchi, MD

Director, Traditional Medicine Department, Centro di Orientamento Educativo, Barzio,

Italy

Paola Cantù, MD

Head, Nephrology Department, Gallarate Hospital, Gallarate, Italy

Fabio Firenzuoli, MD

Phytotherapist, Director, Centre for Natural Medicine, S. Giuseppe Hospital, Empoli, Italy

Gabriela Mazzanti, PhD

Associate Professor, Faculty of Pharmacy, University “La Sapienza,” Rome, Italy.

OBJECTIVE: To report a case of rhabdomyolysis caused by Commiphora mukul, a natural lipid-

lowering agent.

CASE SUMMARY : 55-year-old man was taking an extract of C. mukul 300 mg 3 times daily to

lower his cholesterol level. He developed rhabdomyolysis with hemoglobinuria after 2 weeks

of treatment. Laboratory tests showed creatine kinase 144 600 IU/L (reference range 24–

195), myoglobin >3000 ng/mL (28–72), lactate dehydrogenase 7157 IU/L (230–460),

aspartate aminotransferase 1115 IU/L (10–35), and alanine aminotransferase 205 IU/L (10–

35). Analysis of a urine sample was 2+ positive for hemoglobin. All parameters returned to

normal after the herbal preparation was discontinued.

DISCUSSION: The Naranjo probability scale indicates C. mukul as the possible cause of

rhabdomyolysis in our patient. Drug-induced rhabdomyolysis is an established but rare

adverse effect of high doses of cholesterol-lowering agents (statins) or interactions between

drugs (eg, statins and fibrates). As of May 28, 2004, to our knowledge, this is the first

reported case of rhabdomyolysis following C. mukul ingestion.

CONCLUSIONS: Our report describes a case of rhabdomyolysis possibly caused by C.

mukul and underlines the need for active surveillance of natural products.

5. Cardio protective effect of Commiphora mukul resin extract in

isoproterenol induced myocardial infarction in rats by Dr.K.Kannabiran,

Ms.R.Sangeetha.

The present study investigates cardio protective effect of Commiphora mukul (Guggul) resin

extract on LDH, CPK, SGOT and Na+- K+ levels against isoproterenol induced myocardial

infarction (MI) in rats. The male Wistar rats were randomized into four groups. Group-I rats

were administered with normal saline (0.9% NaCl), Group-II rats were administered with

isoprenaline (2 mg/kg body wt) whereas Group-III rats were administered with Verapamil

(5µmole/kg body wt) as standard drug intravenously followed by isoproterenol after 10mins

of administration. Group-IV rats were administered with Commiphora mukul (Guggul) resin

extract followed by isoproterenol after 10mins of administration. After 48 hours of drug

administration, it showed significant reduction in serum cardiac enzymes like LDH, CPK,

SGOT and Na+- K+ levels compared with isoproterenol induced group rats which showed

increase in cardiac enzymes with myofibrillar damage. Hence, the present study is the

effective remedy for the myocardial infarction in rats using Commiphora mukul

PLANT PROFILE

PLANT PROFILE

Indian Name :Guggul

Botanical Name(s): Commiphora Mukul,

Family Name: Burseraceae

Kingdom: Plantae

Division: Magnoliophyta

Class: Magnoliopsida

Order: Sapindales

Family: Burseraceae

Genus: Commiphora

Popular Name(s): Commiphora, Mukul, Indian Bedellium Tree, Gugulipid, Mukul myrrh

tree, , guggul.

Parts Used: Whole Plant

Habitat: Rocky tracks of Western India and Eastern Himalayas.

INTRODUCTION 

Commiphora mukul is one of the most respectable herbs known in Indian Herbal

System (Ayurveda). Commiphora mukul gum resin is used in treatment of

rheumatism, neurological disorders, obesity and related disorders, hypothyroidism,

syphilis, skin and urinary disorders. Commiphora mukul resin oil possesses a

powerful stimulant, anti-inflammatory, antioxidant, astringent.

DESCRIPTION 

Guggul is a yellowish resin, which is secreted by a small, thorny mukul myrrh tree

called Commiphora mukul. The shrub reaches a maximum height of 4 to 6 feet and

bears thorns on its branches. The leaves are small similar to those of neem. The

flowers are red and the fruit is oval in shape and pulpy in nature. The gum resin

excreted by the bark of the plant is called guggul. The plant is grown throughout the

north India. The herb has been playing a major role in the traditional medicine of

India. It is also known as guggul gum, guggul, gugglesterone.

ORIGIN

The mukul myrrh (Commiphora mukul) tree is a small, thorny plant distributed

throughout India. Guggul and gum guggul are the names given to a yellowish resin

produced by the stem of the plant.

CHEMICAL COMPOSITION / KEY ACTIVE CONSTITUENTS

Ketone fraction that is extracted from the resin contains the most potent

cholesterol lowering components. This is composed of C21 or C27 steroids, with

the major components being Z and E-guggulsterone. Guggul contains resin,

volatile oils, and gum. The extract isolates ketonic steroid compounds known as

Guggulsterones. These compounds have been shown to provide the lipid-lowering

actions noted for guggul.

Guggul significantly lowers serum triglycerides and cholesterol as well as LDL

and VLDL cholesterols (the "bad" cholesterols). At the same time, it raises levels

of HDL cholesterol (the "good" cholesterol), inhibits platelet aggregation, and may

increase thermogenesis through stimulation of the thyroid, potentially resulting in

weight loss. In Ayurvedic medicine, it is used to remove "Ama", or deposits of

waste or toxic material in the body, including mucus and mineral deposits in the

joints, Thus reducing a possible cause of sluggishness, inflamed joints, and many

other conditions. A direct anti-inflammatory effect has been observed for

guggulsterones. Guggul has also been shown to reduce the stickiness of platelets-

another effect that lowers the risk of coronary artery disease.

REMEDIES FOR :

High cholesterol levels,

High triglyceride levels & Atherosclerosis,

Obesity. Guglipid offers considerable benefit in preventing and treating

atherosclerotic vascular disease (Heart disease).

It is most effective in lowering LDL cholesterol and triglycerides.

It also raises the level of good cholesterol (HDL).

It is used as expectorant i.e. in cough problems.

This guggul has great effect in preventing heart atherosclerosis.

PHARMACOLOGY:

Lipid-lowering effects: Guggul (gum guggul) is a resin produced by the mukul

mirth tree. Guggulipid is extracted from guggul using ethyl acetate. The

preparation produced by extraction with petroleum ether is called a fraction A.

Typical guggulipid preparations contain 2.5-5% of the plant sterols guggulsterones

E and Z. These two components have been reported to exert effects on

lipids.Several hypotheses have been advanced to explain these effects on lipids.

Guggulsterones, particularly guggulsterone (4,17(20)-pregnadiene-3,16-dione),

have been reported to function as antagonists of the farsenoid X receptor (FXR)

and the bile acid receptor (BAR), nuclear hormones which are involved with

cholesterol metabolism and bile acid regulation. It has been reported that

guggulsterone does not exert its lipid effects on mice lacking FXR. Other

publications have proposed that guggul may inhibit lipogenic enzymes and HMG-

Co A reductase in the liver. increase uptake of cholesterol by the liver via

stimulation of LDL receptor binding. directly activate the thyroid gland. and/or

increase biliary and fecal excretion of cholesterol.

Antioxidant effects: Guggul extracts have been reported to possess

antioxidant properties possibly mediating protection against myocardial

necrosis

Platelet effects: Guggulipid has been found to inhibit platelet aggregation

and increase fibrinolysis.

Anti-inflammatory: the results of several studies suggest possible anti-

inflammatory and antiarthritic activities of guggul. On a per-microgram

basis, guggulipid appears to be significantly less potent than Indomethacin

or hydrocortisone.

DOSAGE

Daily recommendations for guggul are typically based on the amount of

guggulsterones in the extract. A common intake of guggulsterones is 25 mg three

times per day. Most extracts contain 5-10% guggulsterones. For a 5 percent

guggulsterone extract, this translates to an effective dose of 500mg 3 times/day.

ADVERSE EFFECTS Studies with the crude oleoresin reported numerous side

effects, including diarrhea, anorexia, abdominal pain, and skin rash. Hence,

Ayurveda has not only cautioned the use of Guggul but also given many combined

formulary with other herbs which when used as medicine causes no side effects

USES & BENEFITS OF GUGGUL

Guggul possesses strong disinfecting properties and is a weight loss and fat burning

agent.

It lowers elevated serum cholesterol and triglycerides, while maintaining or

improving the HDL to LDL ratio.

The herb increases white blood cells count and reduces the risk of coronary artery

disease.

Guggul was historically used for a condition called medoroga, a disease similar to the

modern atherosclerosis.

Studies report that it increases the production of thyroid hormone. Since this hormone

breaks down cells of protein, fat and carbohydrates, this herb is believed to be

promoting weight loss.

The herb is used as an anti-inflammatory and pain-relieving herb by many Indian

herbal doctors.  

Guggul is useful in arthritic pains and helps in reversing the degenerative changes

that occur in joints and bones.

The herb is widely used in diseases like rheumatoid arthritis, gout, osteoarthritis,

sciatica, paralysis, hemiplegia, lymphadenopathy, etc.

Since it acts as a blood purifier, it is widely used in skin diseases.

Guggul helps in promoting the production of red blood corpuscles (RBC) and

improving the action of white blood corpuscles (WBC).

It helps in strengthening the digestive system, easy secretion of digestive juices,

works as an appetizer and avoids indigestion and constipation.

The herb is beneficial in hemorrhoids and colitis and relieves from hyperacidity and

belching.

Regular use of guggul helps in improving sexual ability, sperm count and sperm

quality.

Guggul helps in treating leprosy and eczema. Its extract also helps in fighting tumors.

The herb is useful in menstrual disturbances and painful menses.

CAUTION

Guggul extract can generate side effects like headache, nausea, diarrhea, anorexia,

abdominal pain and skin irritation in some individuals.

Thyroid patients should consult a doctor before using guggul extract.

AIMS AND OBJECTIVES

AIMS AND OBJECTIVES

AIM:

Anti inflammatory prospective study of herbal extract of Commiphora mukkul (guggul) bark

by adopting carrageenan induced paw oedama in wistar albino rats.

OBJECTIVES:

To evaluate the inflammation caused by carrageen, formalin, acetic acid.

To evaluate the paw oedema by plethysmometer.

To evaluate the efficacy of the Commiphora mukul.

To study the anti-inflammatory activity of alcoholic extract of acetone extract of

Commiphora mukul on carrageen induced hind paw oedema in rats.

To study the (control) anti-inflammatory activity of the extracts with the standard

marketing drugs.

MATERIALS & METHODS

MATERIALS AND METHODS

MATERIALS:

1) Soxhlet extractor  

2) Collection of plant materials

3) Experimental animals

4) Inflammation inducers

5) Lab materials(Plethysmometer)

Soxhlet extractor :

1: Stirrer bar/anti-bumping granules

2: Still pot (extraction pot)

3: Distillation path

4: Soxhlet Thimble

5: Extraction solid (residue solid)

6: Syphon arm inlet

7: Syphon arm outlet

8: Reduction adapter

9: Condenser

10: Cooling water in

11: Cooling water out

Collection of plant materials : The stem of commiphora mukkul were collected during the months of

March and April from the hobby hub located at nacharam (Hyderabad).

The samples were washed with distilled water and dried under shade.

mechanically pounded to get coarse powder and passed through 40

number sieve meshes.

The sample powders were processed in such a way that they are suitable

for both powder studies and phytochemical analysis.

All the stem were Shade dried at room temperature

until they were free from moisture pulverized in electric grinder.

The  powder  was  obtained  and  extracted  separately  by 

continuous hot  extraction  process  using  soxhlet  extractor.

Experimental animals:  

Healthy albino rats of either sex (Wistar strain) weighing 150‐160 g

belonging to Wistar strain were  used  for  present  study. 

The  animals  had  free  access  to  food  and  water  and  were 

maintained under controlled temperature (27±20c) and 12 h: 12 h light and d

ark cycle.

 Initial body weight of each animal was recorded.

Inflammation inducers: Carrageenan

Histamine

Formalin

Acetic acid

lab materials(plethysmo meter):

The instrument designed to measure of small volume changes. For precise and

rapid screening of small rodents for inflammation or edema of the paw. 

Model (Rat&Mouse)

Measurement Range 0-80 ml

Resolution 0.01ml

Display LCD

Output Printer and RS232C

Power Requirements 100V 50/60Hz

METHODS:

Preparation of extracts 

Extraction process

Carrageenan-induced inflammation in Rats

Test for anti-inflammatory activity 

Preparation of extracts :The coarse powder 150 grams of the given samples were extracted with 600 ml of

ethanol (95%) by continuous hot percolation using Soxhlet apparatus until the

completion of extraction procedure. The successive extractions were done separately

for each solvent namely Chloroform, Ethanol, Ether, Benzene, Hexane and water. The

powder solvent ratio employed for the present study was 1:4. On completion, the

extracts were filtered and the solvents were removed by distillation and dried under

reduced pressure and controlled temperature 50°-60°. They were refrigerated until

use. Both stem and bark powder samples were subjected to various analyses such as

organoleptic characters ,fluorescence studies physico-chemical properties ,preliminary

phytochemical screening.

Extraction process:Ethanol extract:

Dried stem of commiphora mukkul were reduced to a fine powder with a mechanical

grinder. The powder plant material (200 g) was soaked in 3 l of 80% ethanol and

stand for 3 days. The extract was concentrated to dryness and stored at a temperature

of – 4oC until use.

Diagrammatic representation of a soxhlet extractor: A Soxhlet extractor is a piece

of laboratory apparatus invented in 1879 by Franz von Soxhlet. It was originally

designed for the extraction of a lipid from a solid material. However, a Soxhlet

extractor is not limited to the extraction of lipids. Typically, a Soxhlet extraction is

only required where the desired compound has a limited solubility in a solvent, and

the impurity is insoluble in that solvent. If the desired compound has a significant

solubility in a solvent then a simple filtration can be used to separate the compound

from the insoluble substance.

In order to remove the contaminants from the blubber samples, we used an extraction

protocol which put them, and the lipid material, into solution. First we weighed each

sample for later reference, and cut the blubber samples into small pieces. These pieces

were then homogenized with a mortar and pestle in sodium sulphate, to remove any

water.

The homogenate was transferred to a cellulose extraction thimble, and covered with

glass wool. We ran these samples in a Soxhlet extractor for 4 hours; reflux events

occurred every 7-10 minutes.

The sample, which has been homogenized in sodium sulphate, is covered with glass

wool and contained in a porous cellulose thimble. The thimble is placed in

the extraction tube, which itself sits on a flask containing an organic solvent (like

hexane).

The solvent is boiled, and its vapor travels upward through the extraction tube into

the condenser tube. The cool water flowing around the outside of the condenser tube

condenses the vapor, which then drips into the thimble, containing the sample.

Because the contaminants and lipid are soluble in organic solvents, they move into the

condensed solvent as it accumulated in the thimble. The solution, now containing the

contaminants and dissolved lipid, build up in the thimble. Once the liquid reaches the

level of the bypass arm, it is siphoned back into the flask.  This continuous

condensation, buildup, and siphoning is known as the reflux event.

The advantage of the soxhlet is that once the contaminants and lipid material are

brought into solution, and siphoned back into the flask, they stay in the flask--so that

the sample in the extraction thimble is continuously re-exposed to fresh, heated

solvent--thus greatly increasing the extraction rate.

After extraction, we reduced the volume of solvent in our samples by rapid rotary

evaporation down to about 2mL, using a Rotovap.

After extraction the solvent is removed, typically by means of a rotary evaporator,

yielding the extracted compound. The non-soluble portion of the extracted solid

remains in the thimble, and is usually discarded.

 Liebermann – burchard test :

The Liebermann–Burchard or acetic anhydride test is used for the detection

of cholesterol. The formation of a green or green-blue colour after a few minutes is positive

Method : Dissolve few drops of Commiphora mukul extrac in dry chloroform in a dry test

tube. Add several drops of acetic anhydride and then 2 drops of conc.H2SO4 and mix

carefully. After the reaction finished, the concentration of cholesterol can be measured

using spectrophotometry.

Carrageenan-induced inflammation in Rats :

Carrageenan is a sulphated polysaccharide obtained from sea weed

(Rhodophyta). It causes inflammation by releasing histamine, 5HT,

bradykinins and prostaglandins that produce inflammation and Oedema

Injection of carrageenan 1% (50 μl) in the rat paw causes a biphasic response:

an early inflammatory response that lasts 6 h and a second late response that

peaks at 72 h, declining at 96 h.

Western blot analysis showed that cyclooxygenase 2 (COX-2) at 24 h point.

In conclusion, rat paw oedema is biphasic and age-weight dependent. The

present results are the first report on the differential expressions of COX-1 and

COX-2 in response to carrageenan injection in the two phases of the rat paw

oedema.

Carrageenan solution injected in the albino rat:

Test for Anti inflammatory Activity: These extracts were tested for anti‐inflammatory activity by carrageenan

(inflamagens) induced rat paw edema method.  

Different groups of animals were taken for experiment is as follows:

Group I served as control (5 ml/kg Normal Saline)

Group II received ethanol extract of Commiphora mukkul (100 mg/kg body

weight)

Group III served as reference standard drug (10 mg/kg b.w).

The mean paw oedema volume of the extract treated groups was compared

both with control group and the standard drug Indomethacin treated groups.

Thus, oedema volume in control (Vc) and extract treated groups (Vt) was

computed. The percentage inhibition was calculated using the formula as

shown below.

where, 

Vc-oedema volume of control group 

Vt-oedema volume of test group

The results are tabulated and analysed using student's 't' test to know the level of statistical

significance.

Rats paw oedema:

Data are expressed as mean ± s.e.m. The level of statistical significance was determined by

one-way analysis of variance (ANOVA) followed by Bonferroni's t-test for multiple

comparisons, using the Graph Pad Prism software.

Each group of animals received subplantar administration of 50 μl of saline or 50 μl of

carrageenan 1% (w /v) in saline . The paw was marked in order to immerge it always at the

same extent in the measurement chamber. The volume was measured by using a

hydropletismometer specially modified for small volumes immediately before subplantar

injection, and 2, 4, 6, 24, 48, 72 and 96 h thereafter. The assessment of paw volume was

performed always in double blind and by the same operator. The increase in paw volume was

calculated by subtracting the initial paw volume (basal) to the paw volume measured at each

time point.

After Carrageenan induced in left Paw of albino rat:

RESULTS

EVALUATION OF ANTI-INFLAMMATORY ACTIVITY

Anti-inflammatory activity of the commiphora mukkul was assessed by using

Plethysmograph. It is a glass tube of 20 mm internal diameter and one end fabricated to a

glass tube with0.5 mm bore. This tube is fused to a flexible tube and a pump (glass -

syringe) and fixed to other end of the tube. This pump is used to adjust the level of mercury

in both the flexible tube and graduated glass tube up to zero level.

For the evaluation of anti inflammatory activity of commiphora.carrageenan induced paw

model rats were selected and divided into four groups of 6 in each group. The paw oedema

was produced by sub-plantar administration of 0.1 ml of a 1% freshly prepared solution of

carrageenan into the right hind paw of rats of each group. The paw volume was recorded

before (0h) and 1 h after histamine injection.

The percent inhibition of the inflammation is calculated using the formula and compared

with control group.

% Inhibition = Vc - Vt /Vc x 100

Vt - edema volume in the drug treated groups

Vc- edema volume in the drug control groups

TABLE: 1

Effect of alcoholic extract of commiphora mukul on inflammation produced in the

paw of rats using carrageenan

DRUG DOSE

(mg/kg)

%oede

ma

omin

15min 30min 45min 60min 75min 90min 120mi

n

Control ----- 25±1.2

(24.9)

29±1.9

(28.0)

32.68±2

.5

(30.0)

35.89±4

.2

(31.5)

39.8 ±

5.6

(32.7)

44.9±5.

8

(28.08)

52.22±6.8

(27.56)

40±6.

8

(25.9

)

GUGGU

L

200mg/

kg

25±5.4

(65.2)

21.0±5

.4

(65.2)

22.0±6.

2 (60.0)

22.4±4.

2

(48.98)

25.0±2.5�(38.6)

34.8±4.

2

(36.87)

44.79±5.8

(32.84)

44 ±

2.5

(28.5

)

GUGGU

L

400mg/

kg

25±5.4

(64.2)

22.0±6

.2

(60.0)

21.56±4

.4

(64.65)

21.0±5.

4

(65.2)

21.0±5.4

(65.2)

21.0±5

.4

(65.2)

25.0±2.5�(38.6)

33.5

± 6.8

(48.0

)

GUGGU

L

600mg/

kg

25±5.4

(65.2)

22.0±6

.2

(60.0)

22.0±2.

2 (58.0)

22.87±5

.4

(59.56)

22.2±6.2

(60.0)

26±5.4

(54.0)

29±1.9

(42.0)

32.4

±2.8

(46.0

)

The increase in paw volume was highest at 1hr after carrageenan injection. The significant

difference was seen in rats treated with Ethanolic extract of Commiphora mukul. The

difference is seen at 1:15hr and 1:45hr which reflex the decrease of release of inflammatory

mediators in the rats which were treated with Ethanolic extract of Commiphora The anti-

inflammatory activity at 0:30min, 11/2h, 2hrs respectively.

DISCUSSION:

DISCUSSION:

The traditional and conventional allopathic treatments suffers from severe effects of ulcers

and perforations, already the anti ulcer activity of plant was reported previously. Thus the

extract of this plant helps in reinforcing the anti-inflammatory activity without causing any

side effects via ulceration and perforation. Thus the use of ethanolic extract of commiphora

mukkul shows the superior effect in treating inflammation without side effect. The anti-

inflammatory activity of extract is mainly due to active principle like sterolsalkaloids,

flavonoids tannins etc. Already are reported to inhibit the COX-2 enzyme and inhibit PGE

synthesis so these presence may be the cause in curing the inflammation. Hence ethanolic

extract of commiphora mukkul can be of great potential in treating inflammation without

causing ulceration hence promoting the health of people.

According to vineagar et al (1987), the development of paw edema is derived from

the release of cytoplasmic enzymes and serotonin from mast cells and the increase of

Postaglandins(PGE) in the inflammatory area. The macrophage in carrageenan inserted

dermal tissue release interleukin-1 causing accumulation of polymorphic nuclear cells

(PMNs) in the inflammatory area and then releases lysosomal enzymes and active oxygen

which induces paw swelling.

The Ethanolic extract of commiphora mukul were evaluated in wistar rats. Using

carragenan Induced hind paw Oedema test after oral administration. The results show the

promising anti inflammatory activity both against acute and chronic inflammation.

CONCLUSION:

CONCLUSION:

The results of the present study shows that commiphora mukul formulation possesses

significant anti inflammatory activity in the wistar rat such as Carragenan Induced hind paw

edema .

Synthetic products are rich source of free radicals which are carcinogenic 6 but plant

extracts are rich source of antioxidants, which can prevent and even help in curing the

cancer and also possesses anti-ulcer activity.

But anti-inflammatory activity may not yet proved or documented therefore we have

selected these herbals to investigate its anti-inflammatory activity.

BIBLIOGRAPHY

BIBLIOGRAPHY

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