anti ulcer property of dgh

7/31/2019 anti ulcer property of dgh

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Study of ulcer healing property of

traditional deglycyrrhizinated

licorice compared with flavonoid de

glycyrrhizinated licorice


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ULCER :

A break in skin or mucous membrane with loss of surface tissue,disintegration and necrosis of epithelialtissue

Causes :Due to the imbalance of aggressive and defensive factors.

Aggressive factors:HCl, Pepsin, Bile and H.pyloi

Defensive factors :Gastric mucosa, bi carbonate secretion, prostaglandins,

nitricoxide, innate resistant of mucosal cells


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ETIOLOGY:

H.Pylori

Acid and Pepsin

NSAIDs

Smoking

caffeine

alcohol

Stress


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Clinical Features :

Abdominal pain, classically epigastric with severity relating to

mealtimes (duodenal ulcers are classically relieved by food,

while gastric ulcers are exacerbated by it);

Bloating and abdominal fullness

Water brash (bitter regurgitation)

Nausea, and sometimes vomiting


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Loss of appetite and weight loss;

Hematemesis (vomiting of blood);

Melena (tarry, foul-smelling feces due to oxidized iron

from hemoglobin);

Rarely, an ulcer can lead to a gastric or duodenal

perforation. This is extremely painful and requires

immediate surgery.


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DIAGNOSIS :

Biopsy during EGD;

Breath testing (does not require EGD);

Direct culture from an EGD biopsy specimen;

Direct detection of urease activity in a biopsy specimen;

Measurement of antibody levels in blood (does not require

EGD). It is still somewhat controversial whether a positive

antibody without EGD is enough to warrant eradication

therapy.


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TREATMENT :

The drug treatment of peptic ulcer is targeted at either

counteracting aggressive factors or stimulating the mucosal defenses.

The ideal aims of treatment are to relieve pain, heal the ulcer and

delay the ulcer recurrence.

Reduction of gastric acid secretion:

H2anti histamines

Proton pump inhibitors


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Anti cholinergics

Prostaglandin analogues

Neutralization of gastric acid secretion:

Systemic

Non systemic

Ulcer protective


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DRUGS :

H2-blocker

Ranitidine

Famotidine

Cimetidine

Nizatidine

Mechanism of action

Histamine H2-receptor antagonists inhibit secretion of acid by

the parietal cells in the stomach lining by blocking H2 receptors.


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Proton Pump Inhibitors :

Omeprazole

Lansoprazole

Esomeprazole

Rabeprazole

Pantoprazole.

Mechanism of action :

These proton pumps move hydrogen ions across cellmembranes into the stomach cavity, thereby lowering pH in the

stomach.


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PLANT REVIEW 17

Kingdom : Plantae

Division : Angiospermae

Class : Dicotyledoneae

Order : Fabales

Family : Fabaceae,Leguminosae

Genus : Glycyrrhiza

Species : glabra


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CHEMICALCONSTITUENTS:

1. GlycosideSaponin group : Glycyrrhizin 50 time sweeter

than sugar .

2. FlavonoidsLiquiritin & Isoliquiritin

3. Proteins .

4. Sugars (glucose ,sucrose ).


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USES:

1. Antihistaminic

2. Expectorant

3. Flavoringagentfor Aloe , Quinine , NH4CL , Chocolates

and others

4. Anti-inflammatory activity

5. Demulcent

6. Soft drink

7. Anti tumor activity:prosate cancer


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Anti ulcer.

Licorice helps heal ulcers by inactivating 15-

hydroxyprostaglandin dehydrogenase in the stomach lining.As with cortisol in the kidney, licorice locally extends thelife of prostaglandins that protect the stomach wall.

9.Skin problems:

emollient,eczema and psoriasis anti allergic

10.Harmonal Action:

pseudo aldosterone activity

Cortisone action estrogenic activity


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2. OBJECTIVE :

a. Procurement of Flavonoid-rich DGL & Traditional DGL

b. Standardization of anti-ulcer models.

c. Evaluation of anti-ulcer activity of Flavonoid-rich DGL and

Traditional DGL.

d. Comparative evaluation of anti ulcer potential of

Flavonoidrich DGL vs. Traditional DGL


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Animal Models :

Surgical: Pylorus Ligation (PL);

Physical: Cold Stress ulcer (CSU)

Chemical: Indomethacin Induced ulcer (IND)


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SCREENING PROCEDURES:

Pylorus ligation induced ulcer

NSAIDs induced ulcer (Indomethacin, Aspirin, Ibuprofen)

Cold stress induced ulcer

Alcohol induced ulcer

Sub acute gastric ulcer in rats

Gastric ischemia reperfusion injury in rats


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METHODOLOGY

ANIMALS

Species: Albino rats.

Strain: Wistar.

Sex: Either sex.

Source: Bred and reared at Natural Remedies Pvt. Ltd.

Body weight range: 180 - 200 g Identification: By cage card and corresponding picric acid colour

body markings.

Number of animals per dose group: 3 per sex.

Acclimation: One week in experimental room.


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Dosing of tests and standards

Dose formulation :

The test substance will be formulated as solution/suspension.

Formulations as above will be freshly prepared before dosing.

Administration of test substance

The test substance will be administered by oral gavage to each

rat as a single dose, using an intubation needle fitted onto a

syringe of appropriate size. The dose administered to

individual rat will be calculated according to its body weight

recorded on the day of test substance administration.


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Statistics

Values will be expressed as mean SEM. The data will be

analyzed by one way ANOVA. The statistical significance will

be set at p 0.05 followed by Dunnets T3.


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PYLORUS LIGATION INDUCED ULCER MODEL

Principle of the test

Pyloric ligation (PL) induced ulcers were caused due to

Imbalance between offensive and defensive mucosal factors.

PL-induced gastric ulcers occur because of an increase inacid-pepsin accumulation due to pyloric obstruction and

subsequent mucosal digestion

Anti-ulcer agents like omeprazole inhibit the acid

secretion and prevent the ulcers.


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Drugs used:

Omeprazole

Chemicals used:

Sodium hydroxidewas used at the conc. of 0.01M for

titration of gastric juices.

Phenolphthalein as indicator in acid base titration.

Observations: Ulcer index, pH of gastric content, total acidity and gastric

content.


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Pyrocedure:

Female, wister rats of 150-170gm, starved 48hr.

Avoid coprophagy.

six animals per dose and as control.

Midline abdominal incision, pylorus is ligated.

Closed by sutures , test compound given orally route.


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Placed 19hr in plastic cylinder d-45mm,closed ends

Animals are sacrificed in co2 anesthesia

Abdomen is opened and ligature is placed around

esophagus

Stomach is removed and contents are drained to centrifuge

tube

Stomach is opened along the greater curvature pined to

cork plate


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Scoring:

0 - no ulcer

1 - petechial hemorrhages

2ulcer24mm

Ulcer index(UI)=UN+US+UP*10-1


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GROUP TREATMENT

i Vehicle control(10ml/kgb.w)

ii Pylorus ligation control (4 h)

Iii Standard control (Omeprazole 10 mg/kg)

Iv Traditional DGL (37.5 mg/kg b.w.)

V Traditional DGL (75 mg/kg b.w.)

Vi Traditional DGL (150 mg/kg b.w.)

Vii Flavonoid-rich DGL (37.5 mg/kg b.w.)

Viii Flavonoid-rich DGL (75 mg/kg b.w.)

ix Flavonoid-rich DGL (150 mg/kg b.w.)


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COLD STRESS INDUCED ULCER MODEL

Principle of the test: Cold stress ulcer (CSU) is a well-accepted model for the

induction of gastric ulcer in which peripheral sympathetic

activation plays an important role in induction of ulcers.

Stress has been reported to have an important role in

etiopathology of gastro-duodenal ulceration, increase in gastric

motility, vagal over activity, mast cell degranulation; decreased

gastric mucosal blood flow and decreased prostaglandin

synthesis


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Significantly, when source of stress is cold as in CSU,

incidence of ulcers is mainly due to increased acid secretion

and generation of free radicals etc

Drugs used:

Omeprazole

Observations

Ulcer index, pH of gastric content.


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STRESS ULCER BY COLD WATER IMMERSION

PROCEDURE:

Groups of 6 wistard rats(150-200).

Oral test drug, placed in restraint cages, 22degc/1hr

Inject i.v via tail vein with 30 mg/kg evans blue.

After 10 min stomach is removed by co2 anaesthesia Formol saline is injected, storage overnight

Then next day stomach are opend with greatercurvature, washed.

lesions(lengths) are measuerd.

EVALUATION: inhibition of the lesion porduction isexpressed as

percentage value.


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INDOMETHACIN INDUCED ULCER MODEL

Principle of the test

Anti-inflammatory drugs like Indomethacin when administered

produce visible gastric ulcers in animals.

Indomethacin is a potent inhibitor of prostaglandin biosynthesis

and prostaglandins are known to play an important role in

maintaining mucosal integrity.


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Drugs used:

Omeprazole

Chemicals used:

Indomethacin was used at the dose levels of 40 mg/kg rat body

weight.

Sodium Carbonate

Observations

Ulcer index, pH of gastric content.


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Indomethacin induced ulcer:

6 wister rats(150-200g)

Test-20mg/kg indomethacin,

After 10 min oraly, test drug in 0.1%tween 80

6hr later , sacrifice in co2,stomaach is removed

Stomach Inject formal saline, kept for over night

Stomach is opened, examined under microscope


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RESULTS

Pylorus ligation induced ulcer

DGL at the dose level of 75 mg/kg and 150 mg/kg. Totalacidity was inhibited by Flavanoid rich DGL at all the doselevels.

Total acidity was reduced by 45.30% and 30.66% byflavonoid-rich DGL and traditional DGL respectively at their

highest dose.

Flavanoid rich DGL at the dose level of 75 mg/kg and 150mg/kg also inhibited ulcerogenic activity of pylorus ligationsignificantly.

Flavonoid-rich DGL at a dose of 150 mg/kg b.w inhibitedulcer index by 88.33% compared to traditional DGL, which atthe same dose level inhibited ulcer index by 77.8%.

Ph t h PYLORUS LIGATION INDUCED ULCER


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Photograph 5.1: PYLORUS LIGATION INDUCED ULCER

NORMAL CONTROL PYLORUS LIGATION CONTROL 4hOMEPRAZOLECONTROL 10mg/kg

TRADITIONAL DGL 150mg/kgFLAVONOID-RICH DGL 150mg/kg


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GRP

S

TREATMENT MEAN

pH

SEM

MEAN

G.CONT

ENT

SEM

MEAN

TOTAL

ACIDITY

SEM

MEAN

ULCER

INDEX

SEM

%INHIBITI

ON OF

ULCER

INDEX

IVehicle control (10 ml/kg)

2 0 0 0 0 0 3.16 0 -

II Pylorus ligation control (4

h)

2.14 0.05 6.33 0.3 120.66 5.55 30 4.32 -

III Omeprazole (10 mg/kg) 6.89* 0.13 2.41* 0.23 15.33* 0.91 3.33* 1.38 88.9

IV Traditional DGL (37.5

mg/kg)

2.2 0.06 5.41 0.35 113.33 2.99 18.66 1.54 37.80

V

Traditional DGL (75 mg/kg)

2.33 0.12 4.58 0.5 103 9.36 7.66 1.3 74.46

VI Traditional DGL (150

mg/kg)

2.39 0.07 4.83 0.33 83.66 8.98 6.66 1.52 77.80

VII

Flavonoid DGL (37.5 mg/kg)

2.45 0.2 4.25 0.4 87.33* 1.97 8.5 1.97 71.66

VIII

Flavonoid DGL (75 mg/kg)

2.52 0.17 3.83* 0.3 72* 7.42 5.66* 0.8 81.66

IX


2.72 0.21 3.33* 0.35 66* 7.16 3.5* 1.5 88.33

Graph 5.1: Volume of gastric content of treated groups in pylorus ligation induced ulcer model

i lbi Wi t t


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using albino Wistar rats

0

1

2

3

4

5

6

7

Treatment groups

Volu

meofgastriccontent(ml)

I Vehicle control (10 ml/kg) II Pylorus ligation control (4 h) III Omeprazole (10 mg/kg) IV Traditional DGL (37.5 mg/kg) V Traditional DGL (75 mg/kg)

VI Traditional DGL (150 mg/kg) VII Flavonoid DGL (37.5 mg/kg) VIII Flavonoid DGL (75 mg/kg) IX Flavonoid DGL (150 mg/kg)

# p < 0.05 Pylorus ligation control Vs Vehicle control*p < 0.05 Treated groups Vs Pylorus ligation control

#

*

*

*

Graph 5.2: pH values of treated groups in pylorus ligation induced ulcer model using albino Wistar rats


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0

1

2

3

4

5

6

7

8

Treatment groups

pHvalues



*p < 0.05 Treated groups Vs Pylorus ligation control

*


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Graph 5.3: Total acidity of gastric juice of treated groups in pylorus ligation induced ulcer model

using albino Wistar rats

0

10

20

30

40

50

60

70

80

90

100

110

120

130

Treatment groups

Totalacidity(mEq/L/100g)

I Vehicle control (10 ml/kg) II Pylorus l igation control (4 h) III Omeprazole (10 mg/kg) IV Tradit ional DGL (37.5 mg/kg) V Tradit ional DGL (75 mg/kg)

VI Tradit ional DGL (150 mg/kg) VII Flavonoid DGL (37.5 mg/kg) VIII Flavonoid DGL (75 mg/kg) IX Flavonoid DGL (150 mg/kg)


#

*

*

*

*

Graph 5.4: Ulcer index of treated groups in pylorus ligation induced ulcer model using albino Wistar rats


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0

5

10

15

20

25

30

35

Treatment groups

U

lcerindex

(scores)




#

*

*

*


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Cold stress induced ulcer:

Flavanoid rich DGL at all dose levels inhibited ulcerogenicactivity of cold stress significantly.

Flavonoid-rich DGL at a dose of 150 mg/kg b.w inhibited

ulcer index by 92.71% compared to traditional DGL, which at

the same dose level inhibited ulcer index by 86.51%.

COLD STRESS ULCER (CSU)


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NORMAL CONTROL COLD STRESS CONTROL 4h OMEPRAZOlCONTRol10mg/kg

TRADITIONAL DGL 150mg/kg FLAVONOID-RICH DGL 150mg/kg

Graph 5.7: pH values of treated groups in cold stress induced ulcer model using albino Wistar rats


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0

1

2

3

4

5

6

7

Treatment groups

pHvalues

I Vehicle control (10 ml/kg) II Cold stress control (4 h) III Omeprazole (10 mg/kg)

IV Traditional DGL (37.5 mg/kg) V Traditional DGL (75 mg/kg) VI Traditional DGL (150 mg/kg)

VII Flavonoid DGL (37.5 mg/kg) VIII Flavonoid DGL (75 mg/kg) IX Flavonoid DGL (150 mg/kg)

*

*p < 0.05 Treated groups Vs Cold stress control

Graph 5.8: Ulcer index of treated groups in cold stress induced ulcer model using albino Wistar rats


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0

5

10

15

20

25

30

35

Treatment groups

U

lcerindex(scores)

I Vehicle control (10 ml/kg) II Cold stress control (4 h) III Omeprazole (10 mg/kg)



*

# p < 0.05 Cold stress control Vs Vehicle control*p < 0.05 Treated groups Vs Cold stress control

#

*

*

*

*


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Indomethacin induced ulcer:

Traditional DGL at the dose level of 37.5 mg/kg and 75 mg/kgsignificantly inhibited ulcer induction. Flavanoid rich DGL at

the dose level of 75 mg/kg and 150 mg/kg inhibited

ulcerogenic activity of indomethacin significantly.




Photograph 5.2: INDOMETHACIN INDUCED ULCER


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VEHICLE CONTROL INDOMETHACIN CONTROL 40mg/kg OMEPRAZOLE CONTROL 10mg/kg

TRADITIONAL DGL 150mg/kg FLAVONOID-RICH DGL 150mg/kg

TABLE 5.2: Anti-ulcer activity of DGL in Indomethacin induced ulcer model using albino Wistar rats.


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GRPS TREATMENT MEAN

pH

SEM

MEAN

ULCER

INDEX

SEM

% INHIBITION

OF ULCER INDEX

IVehicle control (10 ml/kg) 2 0 0.83 0.4

-

IIIndomethacin control (40

mg/kg)2

0 71.5 3.87-

IIIOmeprazole (10 mg/kg)

6.5*0.22 2.83* 1.45

96.04

IVTraditional DGL (37.5 mg/kg)

20 27.83* 8.02

61.07

V Traditional DGL (75 mg/kg) 2.33 0.33 27.66* 5.54 61.31

VITraditional DGL (150 mg/kg)

2.660.42 40 6.06

44.05

VIIFlavonoid DGL (37.5 mg/kg)

2.660.42 54.5 4.02

23.77

VIIIFlavonoid DGL (75 mg/kg)

3.330.42 32.16* 6.23

55.02

IX


3.33

0.42 24.66* 2.13

65.51


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Graph 5.5: pH values of treated groups in indomethacin induced ulcer model using albino Wistar rats

0

1

2

3

4

5

6

7

Treatment group

pHvalues

I Vehicle control (10 ml/kg) II Indomethacin control (40 mg/kg) III Omeprazole (10 mg/kg)



*p < 0.05 Treated groups Vs Indomethacin control

*

Graph 5.6: Ulcer index of treated groups in indomethacin induced ulcer model using albino Wistar rats

#


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0

5

10

15

20

25

30

35

40

45

50

55

60

65

70

75

Treatment group

Ulcerindex(scores)

I Vehicle control (10 ml/kg) II Indomethacin control (40 mg/kg) III Omeprazole (10 mg/kg)



# p < 0.05 Indomethacin control Vs Vehicle control

*p < 0.05 Treated groups Vs Indomethacin control

*

*

*

*

#

*


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Conclusion:

Hypersecretion of gastric acid is a pathological condition, which

occurs due to uncontrolled secretion of hydrochloric acid from the

parietal cells of the gastric mucosa through the proton pumping H+

K+ ATPase

Flavonoids are known to inhibit several enzymes e.g. alkaline

phosphatase, cAMP phosphodiesterase, lipases, hydrolases,

lysosomal H+ - ATPase and Na+ / K+ - ATPase.

Licorice extracts contained several potent antioxidant constituents asdetermined by their ability to inhibit -carotene consumption and

LDL oxidation.


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Pyloric ligation induced ulcers caused due to imbalance

between offensive and defensive mucosal factors

DGL with multiple mechanisms Total acidity was reduced by

45.30% and 30.66% by flavonoid-rich DGL and traditional

DGL respectively at their highest dose.





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The indomethacin-induced ulcer model was employed because

the model shows drugs effect on cytoprotection and gastric

acid secretion.

Flavonoid-rich DGL inhibited ulcer index by 65.51% compared

to traditional DGL, which at the same dose level inhibited ulcerindex by 44.05%.

Stress can cause abnormalities in acid secretion, bile and

pancreatic juice reflux; these are factors that can lead to ulcer

formation


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Flavonoid-rich DGL at a dose of 150 mg/kg b.w inhibited ulcer

index by 92.71% compared to traditional DGL, which at the

same dose level inhibited ulcer index by 86.51%.

On the basis of the data presented here, it can be concluded that

the gastro protective effect elucidated by DGL could be mainly

due to the modulation of defensive factors through an

improvement of gastric cytoprotection and partly due to acid

inhibition and free radical scavenging properties.


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CONCLUSIONS

Flavonoid-rich DGL at a dose of 150 mg/kg b.w was found toinhibit ulcers in PL (88.33%), IND (65.51%), and CSU

(92.71%). While traditional DGL at the same dose level of 150 mg/kg b.w

reduced ulcers in PL (77.8%), IND (44.05%), and CSU(86.51%) induced ulcer models.

Total acidity was reduced by 45.30% and 30.66% by flavonoid-rich DGL and traditional DGL respectively in PL induced ulcermodel.

Conclusively, the ulcer protective effect of Flavonoid-rich DGLmay be due to its anti-oxidant27 along with cytoprotective25

mechanism. Therefore Flavonoid-rich DGL have more potentanti-ulcerogenic as well as ulcer-healing properties thantraditional DGL and could act as a potent therapeutic agentagainst peptic ulcer disease.


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anti ulcer property of dgh