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| ORIGINAL ARTICLE |
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| Year : 2017 | Volume
: 1
| Issue : 3 | Page : 63-67 |
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Gastroprotective effect of Nigella sativa seed on ethanol-induced gastric ulcer in rats
Mustapha Ahmed Bukar, Helga Bedan Ishaya, Nathan Isaac Dibal, Martha Orendu O. Attah
Department of Human Anatomy, University of Maiduguri, Borno, Nigeria
| Date of Web Publication | 26-Dec-2017 |
Correspondence Address:
Mr. Nathan Isaac Dibal
Department of Human Anatomy, College of Medical Sciences, University of Maiduguri, P. M. B. 1069 Maiduguri, Borno
Nigeria
 Source of Support: None, Conflict of Interest: None  | 3 |
DOI: 10.4103/LJMS.LJMS_23_17
Background: Peptic ulcer is a gastrointestinal disease characterized by mucosal damage. The study is aimed at evaluating the gastroprotective effect of Nigella sativa seed extract in rats. Materials and Methods: Twenty rats were divided into four groups of five rats each and were fasted for 18 h. Rats in Groups 1, 2, 3, and 4 were pretreated with normal saline, gestid, and N. sativa extract at 320 mg/kg and 640 mg/kg, respectively, 30 min before administration of 80% ethanol. All the rats were sacrificed after 1 h and the stomachs were cut open. The stomachs were examined for macroscopic lesions and processed for light microscopic study. Results: There was a significant decrease in ulcer index of rats pretreated with gestid suspension, 320 and 640 mg/kg of N. sativa extract as compared to those of rats pretreated with normal saline at P < 0.05. The stomach of rats pretreated with normal saline showed mucosa full of lesions, those of rats pretreated with gestid suspension showed very few vascular congestion, while those of rats pretreated with 640 mg/kg of N. sativa showed few vascular congestion. Photomicrograph of the stomach of rats pretreated with normal saline showed vacuolation of basement epithelium while those of rats pretreated with gestid suspension and N. sativa extract showed normal basement epithelium. Conclusion: N. sativa seed extract was able to protect the stomach mucosa from gastric ulceration. Keywords: Gastric mucosa, Nigella sativa, peptic ulcer, stomach, ulcer index
How to cite this article:
Bukar MA, Ishaya HB, Dibal NI, Attah MO. Gastroprotective effect of Nigella sativa seed on ethanol-induced gastric ulcer in rats. Libyan J Med Sci 2017;1:63-7 |
How to cite this URL:
Bukar MA, Ishaya HB, Dibal NI, Attah MO. Gastroprotective effect of Nigella sativa seed on ethanol-induced gastric ulcer in rats. Libyan J Med Sci [serial online] 2017 [cited 2023 Mar 28];1:63-7. Available from: https://www.ljmsonline.com/text.asp?2017/1/3/63/221495 |
| Introduction | |  |
Peptic ulcer is a gastrointestinal disease characterized by mucosal damage as a result of pepsin and gastric acid secretions, Helicobacter pylori infection, and prolong use of nonsteroidal anti-inflammatory drugs.[1] The most common symptoms of peptic ulcer include epigastric and nocturnal pain which are alleviated by either food intake or antacids; other symptoms include indigestion, nausea, loss of appetite, fatty food intolerance, and heartburn.[2] The cost of treating peptic ulcer disease and its complications amounted to billions of US dollars annually and the worldwide prevalence of the disease is in millions.[3] Nitric oxide (NO) is one of the major substances that prevent gastrointestinal mucosal damage but can also contribute to gastric mucosal damage as a result of certain factors. This is because NO concentrations can cause opposite effects on the same tissue.[4],[5] Several gastrointestinal disorders are associated with oxidative stress and/or antioxidant property malfunction;[6],[7] free radicals and reactive oxygen species (ROS) play a major role in the progress of some diseases including gastric ulcer.[8] ROS instability results in either reduced antioxidant activity or increases ROS production which eventually leads to apoptosis.[9],[10] H2 antagonist and proton-pump inhibitors are used in the treatment of gastric ulcers, but despite a healing rate of >80% within 2 months, the average recurrence rate after treatment is about 60% within 1-year posttreatment.[11],[12] Due to the high recurrence rate after ulcer treatment,[13] coupled with the side effects associated with the prolonged use of certain drugs,[14],[15],[16] there is a need for a natural remedy with little or no side effects.
Nigella sativa (black seed) is a flowering plant that belongs to the family Ranunculaceae.[17] It is native to Southern Europe, Southwest Asia, and North Africa and it is cultivated and widely used in many Southern European, Middle Eastern, and Asian countries as spice and food preservatives and in the treatment of disorders and diseases.[18],[19] The medicinal properties of N. sativa are as a result of its rich chemical components which include amino acids, carbohydrates, minerals, alkaloids, and saponins.[20],[21] In folk medicinal practice, N. sativa is either consumed with food or mixed with honey.[22],[23] Several therapeutic properties are attributed to N. sativa and they include anti-inflammatory, anticancer, hepatoprotective, antibacterial, and gastroprotective properties.[24],[25],[26],[27],[28],[29] This study is aimed at evaluating the gastroprotective effect of aqueous extract of N. sativa on ethanol-induced gastric ulcer in Wistar rats.
| Materials and Methods | | |
Plant material
N. sativa seed was purchased from a traditional herbalist in Damaturu, Yobe State, Nigeria, and was authenticated by a botanist in the Department of Biological Sciences, University of Maiduguri. The seeds were winnowed and grounded using mortar and pestle. Nearly 100 g of the crude extract was soaked in 1 L of distilled water for 24 h. The extract was then filtered and evaporated; it was poured on a tray for complete dryness. After drying, the yield was 35.4 g.
Animal husbandry
Twenty Wistar Rats (80–120 g) were obtained from the animal house, Department of Biochemistry University of Maiduguri. They were fed with grower mash (vital feed, grand cereal) and water ad libitum. They were kept in the animal house of Biochemistry Department for 2 weeks to acclimatize with the animal house conditions (a cross-ventilated room with temperature between 25°C and 32°C, 12 h light/12 h dark cycle) before the commencement of the study. The research was conducted in accordance with the University of Maiduguri Research and Ethical Committee guidelines, the ARRIVE guidelines (reporting of in vivo experiment), and the National Institutes of Health (NIH) guide for the CARE and use of laboratory animals (NIH Publications No. 8023, revised 1978). It was approved by the Department of Human Anatomy, University of Maiduguri, with code number UM/HA/UGP 16.17-002.
Experimental design
The rats were randomly divided into four groups of five rats each and fasted for 18 h before administration of extract. Rats in Group 1 were pretreated with 1 ml normal saline, the rats in Group 2 were pretreated with 1 ml gestid suspension, while those in Groups 3 and 4 were pretreated with aqueous extract of N. sativa at 320 mg/kg and 640 mg/kg, respectively, 30 min before the administration of 1 ml of 80% ethanol. All the rats were sacrificed 1 h after 80% ethanol administration using ketamine injection, and the stomachs were cut open, the stomach content were deposited in a beaker, and 3–5 drops of pH meter were added to determine the pH. Each stomach was washed with normal saline and examined for gross lesions using a hand lens. Severity of the gross lesions were estimated as: 0 for normal mucosa; 1 for vascular congestions; 2 for one or two lesions; 3 for severe lesions; 4 for very severe lesions; and 5 for mucosa full of lesions.[30] The total stomach area (TSA) and ulcerated areas of the stomachs were measured in mm2, and the ulcer index (UI) of the stomachs were calculated as UI = ulcerated area/TSA × 100.[31]
The percentage of ulcer inhibition rates (UIR) of each group was calculated using the formula UIR = UI (control rat–pretreated rat)/UI (control rat) ×100%.[31],[32] TSA was calculated using the formula: TSA = πr2 (mm2), where r = d/2, d is diameter of stomach and π = 3.14.[31]
Histopathology
The stomachs were fixed in 10% neutral buffered formalin, dehydrated in graded series of alcohol, cleared in xylene, and embedded in paraffin wax. The tissues were sectioned at 5 μm with a rotary microtome and stained with hematoxylin and eosin (H and E) and cresyl violet stain.[14]
Statistical analysis
The data were analyzed using a statistical package (InStat statistics version 3.1. GraphPad Inc., La Jolla, USA). They were expressed as mean ± standard error of mean and one-way ANOVA was used to compare the differences between groups, P < 0.05 was considered statistically significant.
| Results | | |
The rats pretreated with N. sativa extract at 640 mg/kg showed a significantly lower UI (11.196 ± 2.645) than those of rats pretreated with the extract at 320 mg/kg (18.486 ± 4.880) at P < 0.05. The ulcer indices of rats pretreated with N. sativa extract at 320 mg/kg and 640 mg/kg are significantly lower than those of rats pretreated with normal saline (55.91 ± 6.94) at P < 0.05. The UI of rats pretreated with gestid suspension (9.858 ± 2.135) is significantly lower than those of rats pretreated with N. sativa extract. UIR of rats pretreated with gestid suspension (77.626 ± 9.806) and aqueous extract of N. sativa at 640 mg/kg (73.96 ± 12.275) are significantly higher than those of rats pretreated with the extract at 320 mg/kg (61.11 ± 13.963) [Table 1]. | Table 1: Effect of aqueous extract of Nigella sativa on ulcer index and inhibition rate of rats
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The gross anatomy of the stomach of rats pretreated with normal saline showed mucosa full of lesions while those of rats pretreated with gestid suspension showed very few vascular congestion, the gross anatomy of the stomach of rats pretreated with N. sativa extract at 320 mg/kg showed few severe lesions, while those of rats pretreated with and N. sativa extract at 640 mg/kg showed few vascular congestion and few mild lesions [Figure 1]. | Figure 1: The photograph of gross anatomy of stomach of Wistar rats after posttreatment with 1 ml 80% ethanol, (a) (pretreated with normal saline), (b) (pretreated with gestid suspension), (c) (pretreated with 320 mg/kg Nigella sativa), and (d) (pretreated with 640 mg/kg)
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There was a significant decrease between the pH of stomach content of rats pretreated with 640 mg/kg of N. sativa extract and those of rats pretreated with gestid suspension [Table 2]. The pH of stomach content of rats pretreated with normal saline (7.90 ± 0.60) and those of rats pretreated with 320 mg/kg of N. sativa extract and gestid suspension (7.2 ± 0.96 and 8.1 ± 0.37, respectively) were all alkaline while those of rats pretreated with 640 mg/kg of N. sativa extract (6.5 ± 0.57) were slightly acidic [Table 2]. | Table 2: Effect of Nigella sativa on the pH of stomach content of Wistar rats
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Microscopic examination of the stomach of rats pretreated with normal saline showed loss of luminal mucosal epithelium, distortion of submucosal layer, and vacuolation of basement epithelium. The stomach of rats pretreated with gestid suspension showed normal muscularis mucosa with distorted submucosal layer and loss of some luminal mucosal epithelium. The stomach of rats pretreated with N. sativa extract at 320 mg/kg showed severe loss and distortion of luminal mucosal epithelium and distortion of submucosal layer. The stomach of rats pretreated with N. sativa extract at 640 mg/kg showed loss of some luminal mucosal epithelium and distortion of submucosal layer [Figure 2]a, [Figure 2]b, [Figure 2]c, [Figure 2]d. The stomach of rats pretreated with normal saline, gestid suspension, and aqueous extract of N. sativa at 320 mg/kg and 640 mg/kg each showed normal muscularis mucosa (basement membrane) with distorted submucosal layer [Figure 3]a, [Figure 3]b, [Figure 3]c, [Figure 3]d. | Figure 2: The photomicrograph of the stomach of rats pretreated with, (a) normal saline, showing vacuolated epithelium (white arrows) and loss of luminal epithelium (black arrows). (b) Gestid suspension showing distorted submucosal layer (blue arrows), normal muscularis mucosa (yellow arrows). (c) 320 mg/kg aqueous extract of Nigella sativa showing distorted submucosal layer (blue arrows) and loss of luminal epithelium (black arrows). (d) 640 mg/kg aqueous extract of Nigella sativa showing loss of luminal epithelium (black arrows) and distorted submucosal layer (blue arrows) H and E, ×100
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 | Figure 3: The photomicrograph of the stomach of rats pretreated with, (a) normal saline showing normal muscularis mucosa (yellow arrows) and submucosal layer (blue arrows). (b) Gestid suspension showing normal muscularis mucosa (yellow arrows) and submucosal layer (blue arrows). (c) 320 mg/kg aqueous extract of Nigella sativa showing normal muscularis mucosa (yellow arrows) and submucosal layer (blue arrows). (d) 640 mg/kg aqueous extract of Nigella sativa showing normal muscularis mucosa (yellow arrows) and submucosal layer (blue arrows) cresyl violet stain, ×100
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| Discussion | | |
The significant decrease in UI of rats pretreated with N. sativa extract at both 320 mg/kg and 640 mg/kg as compared to those of rats pretreated with normal saline is an indication that the extract can protect the stomach lining from ethanol-induced mucosal damage. This is attributed to the active compounds present in N. sativa seed such as thymoquinone, thymohyroquinone, pyrazol alkaloids, alpha-hederin, saponin, and citronellol.[18],[33],[34] The significant decrease in UI of rats pretreated with gestid suspension as compared to those of rats pretreated with N. sativa extract at 320 mg/kg and 640 mg/kg is an indication that gestid suspension offers better protection than N. sativa extract. Aqueous extract of N. sativa presents better result in terms of gastric mucosal protection at 640 mg/kg than at 320 mg/kg; this is due to the fact that varying concentration of the same substance may exert a different effect on the same tissue.[4],[5] The significant decrease in UIR of rats pretreated with gestid suspension and N. sativa extract at 640 mg/kg as compared to those of rats pretreated with N. sativa extract at 320 mg/kg is an indication that N. sativa extract at 640 mg/kg is as effective as gestid suspension in terms of UIR.
The few vascular congestion and mild lesions observed in the gross appearance of the stomach of rats pretreated with gestid suspension and N. sativa extract at 640 mg/kg as compared with mucosa full of lesions in the stomach of rats pretreated with normal saline and some severe lesions in the stomach of rats pretreated with N. sativa extract at 320 mg/kg is an indication that N. sativa extract is more effective at a higher dose (640 mg/kg) than lower dose (320 mg/kg). Thymoquinone content of N. sativa seed is reported to significantly decrease malondialdehyde, aspartate aminotransferase, and alanine aminotransferase levels which is as a result of its antioxidant property.[35] The acidic pH of stomach content of rats pretreated with N. sativa at 640 mg/kg might be due to the high protein and carbohydrate content which will trigger off the secretion of gastric juice for digestion.
Six hundred and forty mg/kg of N. sativa seed extract showed decreased rate of luminal mucosal epithelial damage, which might be due to the antioxidant activities of the phenolic compounds present in N. sativa extract.[36],[37]
| Conclusion | | |
Eight percent ethanol was able to inflict gastric mucosal damage in rats, 640 mg/kg of N. sativa seed extract was able to protect the gastric mucosa from ethanol-induced ulcers by decreasing UI, increasing UIR, preventing gastric mucosal lesions, and loss of gastric luminal epithelium.
Acknowledgment
We wish to acknowledge the effort and contributions of Mustapha Ali, Freda Nathan Abwage, Johnson Adamu, Ladi Yahi Mingila, Moloh Immaculate, Emue Nebaari Rapheal, and Mohammed Musa Abba.
Financial support and sponsorship
Nil.
Conflicts of interest
There are no conflicts of interest.
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[Figure 1], [Figure 2], [Figure 3]
[Table 1], [Table 2]
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