https://doi.org/10.52973/rcfcv-e34423

Received: 28/03/2024 Accepted: 27/07/2024 Published: 27/10/2024

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Revista Científica, FCV-LUZ / Vol. XXXIV, rcfcv-e34423

ABSTRACT

Peptic ulcer disease poses a critical health risk, leading to considerable

morbidity and mortality. It arises from an imbalance between aggressive

factors that damage the stomach’s mucosal lining and protective

components that safeguard it. Traditional plant–based remedies have

gained traction as potential alternatives to conventional drugs. This

experiment aimed to assess the anti–ulcer action of the crude extract

of Paeonia mascula through two in vivo tests using male Wistar rats.

The preventive test involved inducing acute gastric lesions, while the

therapeutic test involved treating the lesions with crude extract and

in vitro quantifying stomach parameters such as gastric volume, pH,

total acidity, pepsin activity, and ulcer index. The results illustrated a

noteworthy reduction in ulcer index in preventive and healing tests.

These ndings suggest that the extract has an anti–ulcerogenic effect

by suppressing acid secretion and reducing gastric injuries.

Key words: Antiulcer; crude extract; Paeonia mascula; preventive;

curative

RESUMEN

La afección por úlcera péptica representa un riesgo sanitario crítico,

que provoca un sufrimiento y una mortalidad considerables. Surge

de un desequilibrio entre las variables de fuerza que dañan el

revestimiento mucoso del estómago y los componentes protectores

que lo aseguran. Las curas convencionales a base de plantas han

cobrado fuerza como opciones potenciales a los fármacos ordinarios.

Este experimento pretendía evaluar la acción antiulcerosa del extracto

crudo de Paeonia mascula con dos pruebas in vivo utilizando ratas

Wistar macho. La prueba preventiva consistió en inducir lesiones

gástricas agudas, mientras que la prueba terapéutica consistió en

tratar la lesión con extracto crudo y cuanticar in vitro parámetros

estomacales como el volumen gástrico, el pH, la acidez total, la

actividad de la pepsina y el índice de úlcera. Los resultados ilustraron

una notable reducción del índice de úlcera en las pruebas preventivas

y curativas. Estos descubrimientos recomiendan que el extracto

tiene un impacto antiulcerogénico al sofocar la secreción corrosiva

y disminuir las lesiones estomacales.

Palabras clave: Antiulceroso; extracto crudo; Paeonia mascula;

preventivo; curativo

Assessment of antiulcer activity in crude extract of Paeonia mascula

subsp. atlantica (Coss.) Greuter & Burdet

Actividad antiulcerosa en extracto crudo de Paeonia mascula

subsp. atlantica (Coss.) Greuter & Burdet

Wafa Nouioua

* , Soane Gaamoune

Ferhat Abbas University Setif 1, Faculty of Natural and Life Sciences, Department of Plant Ecology and Biology,

Laboratory of phytotherapy applied to chronic diseases. Setif, Algeria.

National Institute of Agriculture Research. Setif, Algeria.

*Corresponding Author: nouioua.wafa@yahoo.fr

Antiulcer activity in crude extract of Paeonia mascula / Nouioua and Gaamoune ___________________________________________________

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INTRODUCTION

Peptic ulcer disease is a group of ulcerative disorders in areas of

the upper gastrointestinal tract exposed to acid–pepsin secretions,

affecting 10% of the world population [1, 2]. Ulcers that affect the

gastrointestinal system are usually aggravated by an imbalance

between destructive and defensive factors in the stomach [3].

The major protective factors include adequate blood ow and the

secretion of prostaglandins, mucus, and bicarbonate by resident

mucosal cells. Aggressive agents include increased secretion of

hydrochloric acid and pepsin, inadequate dietary habits, consumption

of nonsteroidal anti–inflammatory drugs and alcohol, stressful

conditions, and infection by Helicobacter pylori [4].

The treatment of gastric ulcers includes antacids, muscarinic

antagonists, histamine receptor antagonists, and proton pump

inhibitors. However, long–term use of these drugs can cause side

effects on human health, including hypersensitivity, arrhythmia,

hematopoietic disorders, impotence, and gynecomastia [5]. This

underscores the need for alternative means of control. Currently,

plants and plant–based products appear promising in the renewed

search for better ulcer treatment. Some medicinal plants have been

reported to possess anti–ulcer properties based on studies using

experimental animal models [6].

In this study, we aim to scientifically validate the anti–ulcer

properties of the methanol extract of Paeonia mascula. Peptic ulcer

disease is a prevalent gastrointestinal disorder characterised by

mucosal disintegration and ulcer formation. Our goal is to enhance

the body of knowledge advocating for the use of natural products in

treating peptic ulcers by investigating the anti–ulcer effects of the

methanol extract of Paeonia mascula.

MATERIALS AND METHODS

Plant material

Aerial parts of Paeonia algeriensis Chabert = Paeonia mascula

subsp. atlantica (Coss.) Greuter & Burdet were harvested from Kefrida

Forest at 36°34'14" N, 5°17'24" E, and dried in the shade for later use.

Methanol extraction

The method of Motamed and Naghibi [7] was employed. Briey, 10 g

of powdered areal parts were macerated in 100 mL of 80% methanol for

24 h at laboratory temperature. The resulting solution was evaporated

under a vacuum until dry. The extract was stored at -18 until use.

Animals

Experiments were conducted using adult male Wistar albino

rats (Rattus norvegicus) (180–240 g). The animals were housed in

standard metal cages (33 Wistar albino rats in total, with 15 for testing

preventive effects and 18 for testing curative effects), kept at ambient

temperature (20–25°C), and illuminated on a 12:12 h dark/light cycle.

They were provided with standard food pellets and tap water until

the night before the test or sacrice.

Preventive effects of crude extract against acute gastric lesions

For the determination of anti–ulcerative activity, fteen male Wistar

rats were divided into three groups:

•

The first group was orally administered 2.5 mL of water

(control).

•

The second group was orally administered 2.5 mL of a

suspension containing 200 mg of the extract.

•

The third group was orally administered 80 mg·kg

-1

Omeprazole 30 min before gastric ulcer induction.

The gastric mucosal lesions leading to an acute gastric ulcer

were induced by the oral administration of 1.5 mL of a 150 mM HCl/

ethanol (40:60, v/v) solution [8]. The animals were sacriced under

anaesthesia 60 min after the HCl/ethanol administration. Stomachs

were removed, opened along the greater curvature, rinsed with

physiological saline solution, and stretched on polystyrene boards. The

degree of gastric mucosal damage was evaluated from digital pictures

using a computerised image analysis system (Digimizer version

4.0.0.0). The percentage of the total lesion area (hemorrhagic sites)

to the total surface area of the stomach, excluding the forestomach,

was dened as the ulcer index.

Curative effects of crude extract against acute gastric lesions

Male Wistar rats weighing about 180–220 g were divided into three

groups of at least six rats each. Ulcers were induced in all rats by oral

gavage of 1 mL of absolute ethanol. The experimental groups were

then treated as follows:

• Group I was the control, treated with 1 mL of water only.

• Group II was treated with 200 mg·kg

-1

of crude extract.

• Group III was treated with 30 mg·kg

-1

of Omeprazole for each

of 8 successive days [9].

On the 9

day, all animals were sacriced under anaesthesia. The

stomachs were dissected out and opened along the greater curvature.

The stomachs were gently rinsed with water to remove the gastric

contents and blood clots for subsequent ulcer scoring.

Calculation of ulcer index and percentage ulcer inhibition

The ulcer index has been calculated by adding the total number of

ulcers per stomach and the total severity of ulcers per stomach [10].

The score for the ulcer was made as follows:

0: normal–coloured stomach.

0.5: red coloration.

1: spot ulcers.

1.5: hemorrhagic streak.

2: ulcers.

3: perforation

The mean ulcer score for each animal was expressed as an ulcer

index. The percentage of ulcer inhibition was determined as follows:

(%)

()

Inhibition of UlcerIndex

Control mean index

Control mean indexTestmeanindex

Collection of gastric juice

The stomach was excised carefully by keeping the oesophagus closed

and opened along the greater curvature, and the luminal content was

removed. The samples were collected and centrifuged (Sigma 3–30K,

Germany) at 1000 G for 10 min. The volume of the supernatant was

TABLE I

Preventive eect of Paeonia mascula, crude extract

Ulcer Index Inhibition (%)

Omeprazole 0.15 ± 0.02 83.80 ± 7.20

Crude extract 200 mg·Kg

-1

0.15 ± 0.05 70.25 ± 14.24

Control 0.66 ± 0.27 –

FIGURE 1. A: Gastric lesion observed in crude extract treated

group induced by HCl – Ethanol. B: Gastric lesion observed

in standard group induced by HCl–Ethanol. C: Gastric lesion

observed in control group induced by HCl–Ethanol

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expressed as mL·100 g

-1

of body weight, and the centrifuged samples

were decanted and analysed for gastric volume, pH, and total acidity [11].

Estimation of total acidity

It was measured by the method of Hawk et al. [12]. Briefly, 1 mL

of the supernatant liquid was pipetted out and diluted to 10 mL with

distilled water. The pH of this solution was noted with the help of a pH

meter. The solution was titrated against 0.01N sodium hydroxide using a

phenolphthalein reagent as an indicator. The endpoint was titrated when

the solution turned orange–pink. The volume of NaOH was noted, which

corresponds to total acidity. Acidity was expressed as a formula [13]:

Acidity

Volume of NaOH Normality mEq

100��

Estimation of pepsin activity:

Aliquots of 20 μL of the gastric contents were incubated with 500μL

of albumin solution (5 mg·mL

-1

, 0.06 N hydrochloric acid) at 37°C for

10 min. The reaction was stopped with 200 μL of 10% trichloroacetic

acid, and the samples were centrifuged (Sigma 3–30K, Germany) at

1500 G for 20 min. The supernatant was alkalinized with 2.5 mL of 0.55

M sodium carbonate, and 400 μL of 0.1 N Folin reagent was added to

the tubes, which were then incubated for 30 min at room temperature.

The absorbance (spectrophotometer UV visible VWR) of the sample

was determined at 660 nm. A standard curve of tyrosine was used for

the determination of the concentration of pepsin. The pepsin content

of the gastric uid was expressed as μg of tyrosine·mL

-1

(Eq. tyr) [14].

Statistical analysis

The values are expressed as the mean ± SEM for six rats in each

group. All the data were analysed statistically by the t–test of the

student, followed by the Fisher test. The difference was considered

signicant at P<0.05.

RESULTS AND DISCUSSION

Preventive effect

The anti–ulcerative preventive effect of crude extract of Paeonia

mascula is shown in TABLE I below

with a percentage inhibition of 75.30 ± 17.64% in comparison to the

standard drug (80 mg·kg

-1

), which provided 83.80 ± 7.20% protection.

Oxidative stress and mucosal inammation are the main factors

associated with the pathogenesis of the hydrochloric acid (HCl) and

ethanol (EtOH)–induced gastric ulcer model [15, 16], a commonly

used in vivo model that induces gastric mucosal erosion, bleeding,

Results showed that rats pre–treated with omeprazole or crude

extract before being given HCl/ethanol solution had signicantly

reduced areas of gastric ulcer formation compared with the ulcer

control group (FIG. 1).

Pre–treated animals with crude extract (200 mg·kg

-1

) signicantly

reduced the formation of ulcers induced by the HCl/ethanol mixture,

TABLE II

Curative eect of Paeonia mascula, crude extract

Treatment

Dose (mg·kg

-1

)

p.o

Volume of gastric

juice (ml·100 g

-1

)

Total acidity

(meq·L

-1

)

Ulcer index

Inhibition

(%)

Pepsin (µg·ml

-1

)

Eq.tyr

Control 10 ml·kg

-1

0 1.84 ± 0.14 2.80 ± 0.14 70.16 ± 0.30 9.00 ± 1.22 – 67.96 ± 1.71

Standard omeprazole 50 1.25 ± 0.05 5.24 ± 0.25 22.5 ± 7.5 4.83 ± 0.20 48.77 ± 5.85 11.15 ± 2.90

Crude extract 200 1.24 ± 1.12 4.50 ± 1.20 24.22 ± 1.12 4.12 ± 0.89 50.11 ± 3.22 9.54 ± 4.14

Antiulcer activity in crude extract of Paeonia mascula / Nouioua and Gaamoune ___________________________________________________

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perforation, and other damage [17, 18]. Driven by oxidative stress,

the nuclear factor κB (NF–κB) pathway is regulated to amplify the

inammatory response by increasing the release of pro–inammatory

cytokines, including tumor necrosis factor–α (TNF–α) and interleukin–6

(IL–6), and the expression of the downstream inammatory mediators

cyclooxygenase–2 (COX–2) and inducible nitric oxide synthase (iNOS)

[19]. These agents break the mucosal barrier, provoke an increase

in gastric mucosal permeability to H+ and Na+ ions, reduce the

transmucosal potential difference, and induce the formation of

erosions and ulcers [20]. The crude extract of Paeonia mascula was

effective in signicantly reducing the ulcer index and also increased

the mucus content. This model demonstrates the cytoprotective

effect of Paeonia mascula extract.

Curative effect

The results of the potential curative action against experimentally

induced ulcers caused by the administration of HCl–Ethanol are

demonstrated in TABLE II.

Oral administration of the crude extract (200 mg·kg

-1

) for 8 d

signicantly (P<0.05) reduced the ulcer index in ethanol–induced

experimental ulcers in rats, with a percentage of 50.11 ± 3.22%; this

result was better than the standard drug, which was 48.77 ± 5.85%.

These results demonstrate that the crude extract of Paeonia

mascula signicantly reduced ethanol–induced ulcers. The extract

at a dose of 200 mg·kg-1 signicantly decreased the ulcer index,

pepsin activity, and total acidity when compared to their respective

controls, which indicates the healing of the induced ulcer.

The anti–ulcer activity shown by the crude extract of Paeonia

mascula in ethanol–induced ulcers suggests that the extract has a

cytoprotective effect, meaning that it protects the gastric mucosa

through mechanisms other than inhibiting gastric acid secretion.

[21]. Such mechanisms include inhibition of leukotrienes [22],

pepsinogen [23], and substance P [24], free radical scavenging [25],

increasing gastric mucosal blood ow [26], increasing the protective

glycoprotein content and thereby strengthening the gastric mucosa,

and prevention of oxidation of the mucosal xanthine dehydrogenase.

CONCLUSION

The results of this research show a potential curative effect of the

crude extract of Paeonia mascula.

The methanol extract of Paeonia mascula appears to have a

signicant anti–ulcer effect, prominently inhibiting acid secretion

and the formation of stomach lesions. These ndings suggest that the

phytoconstituents in the extract may protect against ethanol/HCl

and

absolute ethanol–induced gastric mucosal damage by inhibiting gastric

acid, pepsin, histamine, and free radicals while promoting mucus

secretion. Additionally, the scavenging of reactive oxygen species

(ROS) by polyphenols is considered an important mechanism in the

healing of gastric ulcers. These multifaceted properties indicate the

extract’s potential in developing future medications. However, further

research is crucial to investigate the extract’s toxicity and chemical

composition, which are essential for its safe and effective utilization.

Conict of interests

The authors declare that they have no conicting interests.

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