Protective Effect of Ventilago maderaspatana Bark Extract against various Ulcer Models in Rats

Preeti Shanbhag¹, Ramdas Bhat²*, A R Shabaraya³

¹PG Scholar, Department of Pharmacology, Srinivas College of Pharmacy,

Valachil, Post Farangipete, Mangalore, Karnataka, India-574143.

²Assistant Professor, Department of Pharmacology, Srinivas College of Pharmacy,

Valachil, Post Farangipete, Mangalore, Karnataka, India-574143.

³Principal and Head of the Department of Pharmaceutics, Srinivas College of Pharmacy,

Valachil, Post Farangipete, Mangalore, Karnataka, India-574143.

*Corresponding Author E-mail: ramdas21@gmail.com

ABSTRACT:

Background and Objective: Gastric ulcer is a common gastrointestinal disorder, and traditional medicines have been used for centuries to treat various ailments, including gastric ulcer. Ventilago maderaspatana bark is a traditional remedy used in Ayurvedic and Malay medicine for the treatment of gastric ulcer. This study aimed to investigate the anti-ulcer potential of the ethanolic extract of Ventilago maderaspatana bark against in vivo indomethacin- and cold restrained stress-induced gastric ulcer, and pylorus ligation assays. Methodology: The ethanolic extract of Ventilago maderaspatana bark was administered orally at doses of 100, 200 and 400 mg/kg. The anti-ulcer potential of the extract was evaluated against in vivo indomethacin- and cold restrained stress-induced gastric ulcer, and pylorus ligation assays. The ulcer index, pH of gastric acid, volume of gastric juice, free and total acidities were measured. Result: The extract of Ventilago maderaspatana bark (400 mg/kg) significantly (P<0.05) reduced the ulcer index in all assays used. Furthermore, the extract significantly increased the pH of gastric acid while reducing the volume of gastric juice and free and total acidities. Conclusion: The ethanolic extract of Ventilago maderaspatana bark has significant anti-ulcer potential against in vivo indomethacin- and cold restrained stress-induced gastric ulcer, and pylorus ligation assays. These findings support the traditional use of Ventilago maderaspatana bark in the treatment of gastric ulcer. Further studies are required to elucidate the mechanisms underlying its anti-ulcer potential.

KEYWORDS: Antiulcer activity, ethanolic bark extract of Ventilago maderaspatana (EBVM), Ranitidine, Ulcer index, indomethacin, Pylorus ligation.

INTRODUCTION:

Peptic ulcer disease is a common condition caused by an imbalance between aggressive and protective factors in the stomach¹. Factors such as lifestyle, medication use, H. pylori infections, and family history can contribute to its occurrence. Complications include bleeding and organ penetration². Treatment options include antibiotics, antacids, and proton pump inhibitors, but they can have unwanted side effects. Identifying the underlying cause and making lifestyle modifications are important preventive measures¹.

There is a significant amount of research on the phytochemical composition of plants and their potential use in treating various diseases³. Many plant-derived compounds such as alkaloids, flavonoids, steroids, glycosides, terpenes, tannins, and phenolic compounds have been identified for their biologically active properties⁴. In addition, the evaluation of antioxidant activity is important as antioxidants are essential in preventing oxidative damage in the body³.

Ventilago maderaspatana is a medicinal herb that belongs to the Rhamnaceae family and is widely distributed in forests of low elevations in South Greece, India, Indonesia, Myanmar, and Sri Lanka⁵. It has been traditionally used to treat various health conditions such as Kapha, dyspepsia, ulcer, colic disorder, leprosy, scabies, skin disorders, fever, inflammation and general disability⁶. The plant contains various biologically active compounds such as alkaloids, flavonoids, saponins, steroids, glycosides, terpenes, tannins, and phenolic compounds, which have been reported to possess antioxidant and nutraceutical effects⁶.

The present studies to evaluate the Ulcer protective activity of Ventilago maderaspatana on stress induced ulcers in rats.

MATERIALS AND METHODS:

Collection and authentication of plant:

The bark of Ventilago maderaspatana was collected from Mangalore, Karnataka, India and authenticated by Dr. H. S. Shenoy, MSc, M.Phil., Ph.D., Principal scientist and Head of Botany division.

Study site:

The current study was conducted in Department of Pharmacology, Srinivas college of Pharmacy, Valachil, Mangalore.

Animals:

Rats of both sexes weighing between 150 and 200 g were utilized in the following investigation. Under a constant 12-hour cycle of light and dark, the animals were fed a standard pelleted diet and distilled water as required. All of the animals spent the five days preceding up to the experiment in a lab setting. Now there are 6 groups of 5 animals each made up of the animals. The ethical criteria were followed during the conduct of this experiment.

Plant extract:

Ventilago maderaspatana bark was powdered after being shade-dried for 15 days following collecting. A Soxhlet apparatus (Borosil, Mumbai, India) was used to extract about 0.98 kg of powdered drug material using 99% pure ethanol in a 1:2 (w/v) ratio. The resulting extract (EBVM) was dried using a rotavapor (Roteva-Equitron, Medica Instruments, Mumbai, India), and the dry mass was weighed and recorded. A yield % calculation was made. The obtained weight of dried crude extract was around 0.21 g, commemorating a yield of 21.16%.

Acute oral toxicity study and selection of doses:

According to the Organization for Economic Co-operation and Development's (OECD) 423 recommendations (OECD recommendations for the Testing of Chemicals, 2010), an acceptable oral dose of EBVM was established using an acute oral toxic test on rats. The EBVM was made by dissolving the extract in distilled water and adjusting the concentration so that it would not exceed 1 ml/100 g of the rat. It was administered at various doses up to 2000 mg/kg. Animals were monitored for behavioral changes, any toxicity, and death for up to 48 hours after the extract was delivered (p.o.). The acute toxicity assessment led to the selection of three different EBVM doses (100, 200, and 400 mg/kg, p.o.) for this investigation.

Anti-ulcer assays

Indomethacin-induced gastric ulcers

Indomethacin (IND; 5 mg/kg p.o.) was given for five days in order to develop ulcers in the stomach⁷. Following the induction of the ulcer, the animals were given either misoprostol (100 g/kg p.o.) or EBVM (100, 200, and 400 mg/kg) once daily for an additional five days, while the control group only received vehicle. The ulcer index was calculated after the rats were sacrificed on the fifth day following the injection of the test solutions⁸. Briefly, the animals were divided into six groups (n = 5) and treated with the respective test solutions as given below:

a) Group 1 (normal control group) – vehicle + vehicle.

b) Group 2 (negative control group) – 5 mg/kg IND + vehicle.

c) Group 3 - 5 mg/kg IND + 100 mg/kg misoprostol.

d) Group 4 - 5 mg/kg IND + 100 mg/kg EBVM.

e) Group 5 - 5 mg/kg IND + 200 mg/kg EBVM.

f) Group 6 - 5 mg/kg IND + 400 mg/kg EBVM.

Cold restraint stress-induced ulcers

The animals were subjected through cold restraint stress (CRS), which caused an ulcer. Thirty minutes prior to exposing the animals to cold stress, ranitidine (50 mg/kg) or EBVM (100, 200, and 400 mg/kg) were given orally. The animals were housed in a restraint cage that was kept at a temperature of 2°C for three hours. Three hours later, the animals were put to death, and the ulcer index was calculated⁸. In a nutshell, the animals were divided into six groups (n = 5) and given the appropriate test answers as described below:

a) Group 1 (normal control group) – vehicle + vehicle.

b) Group 2 (negative control group) – CRS + vehicle.

c) Group 3 - CRS + 50 mg/kg ranitidine.

d) Group 4 - CRS + 100 mg/kg EBVM.

e) Group 5 - CRS + 200 mg/kg EBVM.

f) Group 6 - CRS + 400 mg/kg EBVM.

Pylorus ligation-induced ulcers in rats

The animals used in the study were fasted for 48 hours before pylorus ligation (PL), but were allowed access to water. To prevent cannibalism and coprophagy, the animals were housed individually in cages. During the procedure, an incision was made below the xiphoid process while the animals were under anesthesia, and the pylorus was carefully ligated without damaging its blood supply. Test solutions were administered orally, and the animals were placed in plastic cylinders. The animals were sacrificed approximately 18 hours after pyloric ligation, and their stomachs were removed. The contents of the stomach were collected in a graduated centrifuge tube and measured for volume, pH, and acidity after centrifugation through titration with 0.1 N NaOH. Total and free acidity were determined using established methods⁹. The stomach was cut open along its greater curvature and pinned onto a cork plate, and the inner surface was examined for ulcers using a binocular microscope. The severity of the ulcers was graded according to established criteria¹⁰, and the ulcer index was calculated. Ulcer severity is graded as: 0-No ulcer, 1-Superficial ulcer, 2-Deep ulcer and 3-Perforation The ulcer index (UI) was calculated by the following equation:

UI = UN + US + UP × 10-1 where, UN = Average number of ulcers/ animals, US = Average severity scores, UP = Percentage of animals with ulcers. Ulcer index and acidity of the gastric content of the treated animals were compared with the control.

Statistical analysis:

All the data were expressed in mean ± SEM. The significance of differences in mean between control and treated animals for different parameters determined by one way ANOVA followed by Dunnett’s multiple comparison test. Significance for difference between groups were evaluated for student’s t-test to come to final conclusion.

RESULT:

Table 1: Effect of EBFM on ulcer index in the three models of gastric ulcers.

Sl. no	Group	Dose (mg/kg)	IND induced gastric ulcers	CRS induced gastric ulcers	PL induced gastric ulcers
1.	Control	-	10.4	10.32	10.32
2.	Standard	Varying doses	6.14^*B	4.08*A	4.08*A
3.	Test 1	100	8.20	8.22	8.22
4.	Test 2	200	6.18^*	4.12*	6.16*
5.	Test 3	400	4.08^**	2.06**	4.10*

Each value represents the mean (Standard deviation for each of the test solution was less than 10% of the mean value), * Data differs significantly (P<0.05) when compared to the control of respective column, ** Data differs significantly (P<0.05) when compared to the control of respective column, A The standard drugs used was 50 mg/kg ranitidine, B The standard drugs used was 100 µg/kg misoprostol.

Table 2. Effect of EBVM on pylorus ligation induced gastric ulcers model.

Sl. no	Parameters	Control	Ranitidine	EBVM
Sl. no	Parameters	-	50 mg/kg	100 mg/kg	200 mg/kg	400 mg/kg
1.	Volume of gastric juice	3.36 ± 0.45	1.54 ± 0.17**	2.8 ± 0.34	1.86 ± 0.10**	1.7 ± 0.30**
2.	pH	2.14 ± 0.49	1.54 ± 0.17	1.88 ± 0.08	2.6 ± 0.40	3.0 ± 0.45
3.	Free acidity	42.8 ± 10.15	4 ± 0.71**	26.2 ± 4.10	11.8 ± 2.48**	3.2 ± 0.58**
4.	Total acidity	94.8 ± 17.59	10.2 ± 1.28**	56.4 ± 8.84*	24.8 ± 4.38**	6 ± 0.71**

Note: Data in each column is represented as Mean ± SEM; ** indicates P < 0.01 and * P indicates < 0.05 when compared to the control group.

Acute toxicity study carried out on EBVM up to the dose of 2000 mg/kg demonstrated that the extract did not show any sign of toxicity and mortality. However, there was a decrease in physical activity, which was observed only at the dose of 2000 mg/kg. Thus, the present doses regime (100, 200 and 400 mg/kg) was chosen for further studies. Based on the gross examination of the rats stomach, the control animals had ulcers and confirmed by the presence of hemorrhagic streaks. However, there was significant (P<0.05) reduction in ulcer development in IND- and CRS-induced gastric ulcer groups treated with 200 and 400 mg/kg EBVM as indicated by reduction in ulcer index (Table 1). In group underwent PL-induced gastric ulceration, the same doses of EBVM (200 and 400 mg/kg) were significantly (P<0.05) effective in reducing the ulcer index (Table 1). To determine the mechanisms of action of EBVM, the extract antiulcer potency was tested against PL-induced gastric ulcer. The macromorphological data obtained when ulceration of the gastric mucosa was provoked using PL are shown in Table 2. EBVM, at doses of 200 and 400 mg/kg, significantly (P<0.05) reduced the volume of gastric juice and free acidity while at the three doses used (100 to 400 mg/kg) significantly (P<0.05) reduced total acidity. However, at all doses tested, EBVM failed to affect the pH of gastric juice. The extract ability to influence the volume of gastric juice, free acidity and total acidity was similar to the effect of 50 mg/kg ranitidine.

DISCUSSION:

Peptic ulcer and gastritis are gastrointestinal diseases that arise from disturbances in the natural balance between aggressive factors (e.g., acid, bicarbonate, and pepsin) and the maintenance of the mucosal integrity through endogenous defense mechanisms¹¹. Available drugs for treating these conditions have been limited by the presence of side effects, and therefore, attention has been shifted towards natural products as a new source of antiulcer agents¹². Ventilago maderaspatana is traditionally used to treat various health conditions such as Kapha, dyspepsia, ulcer, colic disorder, leprosy, scabies, skin disorders, fever, inflammation and general disability⁶. In this study, the anti-ulcer potential of the EBVM was investigated. The results of the study showed that the extract significantly reduced gastric ulceration as indicated by the reduction in ulcer index in the induced assays. Further findings suggested that the extract exerts its anti-ulcer activity by reducing the volume of gastric juice secreted, gastric free and total acidities. It was also suggested that the anti-secretory effect of the extract was one of the mechanisms through which it was able to protect the stomach mucosa from NSAIDs-induced damage¹³. Additionally, the observed anti-ulcer activity was associated with Ventilago maderaspatana ability to exhibit antioxidant and anti-inflammatory activities⁵. Oxidative stress has been known to take part in the pathogenesis of various diseases, including gastric ulcer, and antioxidants help protect cells from damage elicited by oxidative stress while enhancing the body’s defense systems against degenerative diseases³.

Currently, various non-specific methods are used to restore the imbalances in aggressive factors and maintenance of mucosal integrity, including regular food intake, adequate rest, and avoidance of ulcerogenic agents, such as tobacco, alcohol, and coffee¹⁴. Moreover, there are drugs, such as pump inhibitors, histamine (H2)-antagonists, anticholinergics, and antacids, used in the treatment of ulcer. However, these drugs have been associated with several adverse effects, such as anaphylaxis reactions, gynecomastia, hematopoietic changes, thrombocytopenia, acute interstitial nephritis, nephrotoxicity, and hepatotoxicity. As a result, the focus has been shifted towards natural products as a new source of antiulcer agents¹⁵.

Medicinal plants have been known to be amongst the most attractive sources of new drugs, and have been shown to give promising results in the treatment of various diseases, including gastric and duodenal ulcers¹⁶. Ventilago maderaspatana has been reported to exert several pharmacological properties, and traditional knowledge of its pharmacological properties has been confirmed to be useful in treating and managing ulcer⁵. Despite claims of its potential in the treatment of gastric ulcer, Ventilago maderaspatana has not been screened for anti-ulcer activity.

CONCLUSION:

The present study provided preliminary data for the first time that the bark of Ventilago maderaspatana possesses significant anti-ulcer activity in animal models. It has a gastric antisecretory and acid neutralizing effect that are comparable to reference drug ranitidine. The anti-ulcer activity is probably due to the presence of bioactive compounds like flavonoids, saponin and tannins. Further studies are required to confirm the exact mechanism underlining the ulcer healing and protecting property of the extract and to identify the chemical constituents responsible for it.

CONFLICT OF INTEREST:

The authors have no conflicts of interest regarding this investigation.

ACKNOWLEDGEMENTS:

We are thankful to Staffs for their support and Management of Srinivas college of Pharmacy, Mangalore for providing all the necessary facilities to carry out this research work.

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Received on 23.05.2023 Modified on 23.09.2023

Asian J. Pharm. Res. 2024; 14(1):90-94.

DOI: 10.52711/2231-5691.2024.00015