| Abstract|| |
Background: Natural honey is widely used all over the world as a complementary and alternative medicine in various disorders including gastrointestinal lesions. Aim: To evaluate the effects of combination of low dosage of honey (0.312g/kg) and sucralfate (0.125 or 0.250 g /kg) on gastric protection and to determine any potentiating interactions between them against ammonia-induced gastric lesions in rats. Material and Methods: Twenty-four hours fasted rats were given I ml of ammonium hydroxide 1 % intragastrically and they were killed one hour later under deep ether anesthesia. The gastric lesion index was calculated according to the method of Takaishi et al 1998. Non protein sulthydryls level was determined spectrophotometrically as described by Sedlak and Lindsay 1968. Results: Administration of ammonium hydroxide produced red and black linear lesions and significant depletion of gastric nonprotein sulthydryls level. Oral administration of honey (0.312g/kg) or sucralfate (0.125 and 0.250g/kg) 30min before ammonium hydroxide reduced the severity of gastric mucosal lesions by 1 I or 18 and 42 % respectively, and has shown the changes in nonprotein sulfhydryls level induced by ammonium hydroxide. Furthermore, pretreatment with a combination of a low dose of honey (0.312g /kg) and sucralfate (0.125g or 0.250g/kg) afforded significantly greater protection (58 and 77 %) than that obtained with either of them administered alone. Conclusion: The present results suggest potentiation of gastric protection effect of sucralfate by honey and this may have a clinical value in the treatment of peptic ulcer diseases in Helicobacter pylori positive patients
Keywords: Ammonia, gastric lesions, honey, potentiation, and sucralfate.
|How to cite this article:|
Mobarok Ali AM, Al Swayeh OA. Honey potentiates the gastric protection effects of sucralfate against ammonia-induced gastric lesions in rats. Saudi J Gastroenterol 2003;9:117-23
|How to cite this URL:|
Mobarok Ali AM, Al Swayeh OA. Honey potentiates the gastric protection effects of sucralfate against ammonia-induced gastric lesions in rats. Saudi J Gastroenterol [serial online] 2003 [cited 2020 Oct 23];9:117-23. Available from: https://www.saudijgastro.com/text.asp?2003/9/3/117/33352
Natural honey (honey) is a unique viscous liquid, prepared by the bees from the nectars of various plants. It is widely used all over the world and has occupied a prominent place in the traditional medicine. The revelation in the Holy Koran (Surat Al Nahl, verses 68, 69P)  and documentation in the Hadith clearly referred to the effectiveness of honey in the healing of diseases for mankind  .
Two thousand years ago, honey was listed in the first medical handbook to be used in burns, cuts, abscesses and boils  . Since then efforts have been made to establish the various qualities of honey. Unfortunately, the therapeutic potential of honey has so far been neglected in the modern medicine due to lack of systematic scientific studies. However, scientific support of the therapeutic potential of honey in diverse clinical and experimental conditions is beginning to emerge. Thus, honey has been reported to be effective in gastrointestinal disorders in humans , in the healing of wounds, burns and as antimicrobial agents ,,,,,,,,,[I5] and to have gastric protection against acute and chronic gastric lesions in animals ,,,,,,,,,, . These studies have shown that the mechanisms of action of honey may be due to restoration of essential mediators of gastric protection such as nitric oxide (NO) and nonprotein sulfhydryls (NP-SH), the antioxidant effects and neutralization of stimulated acidity ,,,,,,,,, . Furthermore, honey has the ability to inhibit the growth of Helicobacter pylori ri) in vitro , .
Sucralfate, a basic aluminum salt of sucrose octasulfate, is a unique, locally active ulcer-healing agent. It has been reported to afford protection against various types of experimentally-induced gastric lesions. The gastric protection effects has been suggested to be due to prostaglandin production, stimulation of mucus secretion, increase in blood flow, epithelial cell renewal and antioxidant effect ,,,,,,,,,, . Furthermore, sucralfate is a clinically effective drug in various gastrointestinal disorders including H.pylori-induced chronic gastritis ,,, .
Since honey contains sucrose and different metals including aluminum and sulfate ions  , it is possible that honey may contain a "sucralfate like" substance. Indeed, honey has been reported to share a number of common gastroprotective mechanisms with sucralfate against experimentally induced acute and chronic gastric lesions ,,,, . It has been reported that combination of sucralfate with antimicrobial agents like amoxicillin or tetracycline and metronidazole resulted in higher eradication rate of H.pylori than that of monotherapy with sucralfate  . H. pylori have high urease activity resulting in a high concentration of ammonia in the stomachs of H. pylori-infected patients  . Since ammonia is highly cytotoxic, it is an important aggressive factor in gastric mucosal injury. We have reported recently that both honey and sucralfate at high concentrations afford protection against ammonia-induced gastric lesions in rats  . Therefore, the present work was undertaken to investigate the effect(s) of combination of low doses of honey and sucralfate on ammonia-induced gastric lesions model in rats.
| Materials and Methods|| |
A natural honey was purchased from the College of Agriculture, King Saud University, Saudi Arabia. The honey sample was obtained from Abha region of the Kingdom of Saudi Arabia. The probable sources of nectars for this honey were various plants such as acacia raddiana, acacia flava of the family Leguminosae, Juniperus polycarpos and Morus nigra and Morus alba of the family Moraceae. Sucralfate was donated by Roussel, Middx, England. Honey or sucralfate was diluted/suspended in distilled water (w/v) to obtain desired concentrations. Other chemicals were purchased from the sources indicated. Ammonium hydroxide (BDH), 5,5 dithio-bis-(2-nitrobenzoic acid) (Sigma), ethylenediaminetetraacetic acid (BDH), fructose (Fluka), glucose (BDH), maltose (Calbiochem), reduced glutathione (Sigma), sucrose (BDH), trichloroacetic acid (Sigma), and Tris buffer (Sigma). All chemicals were of analytical grade.
Male Wistar rats, approximately of same age, weighing between 220-240g, obtained from the Animal House, College of Medicine, King Saud University, Riyadh. The animals were maintained on standard rat chow diet with free access to drinking water. The animals were starved for 24h before the experiments, but water was allowed ad libitum. The fasted rats were housed individually in cages with a shield to prevent coprophagy. After 24h the rats were randomly coded in groups of 6 animals, and water was withdrawn. The groups were then subjected to one of the previously planned studies outlined below.
Induction of ammonia-induced acute gastric lesions: The method of Takaishi et al was followed with slight modification  . Twenty-four hours fasted animals were given 1ml of ammonium hydroxide (NH4OH 1%) orally by a plastic syringe fitted with a metallic feeding gastric tube. They were kept in the individual cages and killed lh later under deep ether anesthesia. The stomachs were removed after giving knots both at esophageal and pyloric ends. The separated stomachs were inflated by injecting 10m1 of 3% formalin and immersed in 3% formalin for 10min to fix both the inner and outer layers of the gastric wall. The stomachs were opened by cutting through the greater curvature and examined for lesions under a stereomicroscope with a square grid (X10) by two independent observers who were not aware of the protocol. The length (in millimeters) of bands of lesions was measured, summed per stomach, and used as lesion index.
Evaluation of the effects of honey or sucralfate alone or in combination on ammonia-induced gastric lesions: In this procedure, different doses of honey or sucralfate alone or in combination was given orally 30min prior to exposure to lml of ammonium hydroxide 1 % orally. The dose of honey or sucralfate alone or in combination was calculated as g/kg body weight and the calculated amounts were diluted/suspended in distilled water and administered orally in a volume of 0.5m1/100g body weight. Each control group received distilled water (0.5 mU100g body weight) orally 30min before ammonium hydroxide. Evaluation of gastric lesions in each group was carried out as described above.
Determination of nonprotein sulfhydryls (NP-SH): In some separate experiments, following the same protocol for induction of ammonia-induced gastric lesions and drug pretreatment schedules as described above, gastric glandular NP-SH levels were measured according to the method of Sedlak and Lindsay  . The glandular part of the stomachs was cut, weighed, and homogenized in ice-cold 0.02 M ethylene diamine tetraacetic acid (EDTA). Aliquots (5ml) of the homogenates were mixed in a 15m1 test tube with 4 ml of distilled water and 1 ml 50% trichloroacetic acid (TCA). The tubes were then shaken intermittently in a shaker for 15min and centrifuged at 3000 rpm for 10min. Two milliliters of supernatant were mixed with 4m1 Tris-buffer (pH 8.9); 0.lml of 5,5-dithio-bis-2(-nitro benzoic acid) (DTNB) was added, and the sample was shaken. The absorbency was read within 5min of addition of DTNB at 412nm against a reagent blank (with no homogenates) in a spectrophotometer (Perkin-Elmer, Lamda 5).
The final NP-SH concentrations per gram of tissue (in (mol) were calculated from a previously constructed standard curve with reduced glutathione.
The percentage changes produced by different treatments were calculated by comparing with the group that received water only.
Results are expressed as the mean + SEM. Comparisons between the means were carried out by means of the Unpaired Student's t test while comparing two groups and analysis of variance (ANOVA) and Dunnett's test for multiple comparisons. A difference of p<0.05 was considered significant
| Results|| |
Effects of sucralfate or honey or their combinations on ammonia-induced gastric lesions. Oral administration of 1 ml of ammonium hydroxide 1% has resulted in characteristics gastric mucosal lesions. Pretreatment with sucralfate alone (0.125g/kg) or honey (0.312g/kg) did not produce any significant protective effect. Sucralfate 0.250g/kg afforded 42% protection. However, coadministration of sucralfate (0.125g or 0.2508/kg) together with honey in a dose of 0.312g/kg resulted in significantly greater protective effects (58 and 77%) against ammonia-induced gastric lesions [Table - 1].
Effects of sucralfate or honey and their combinations on glandular NP-SH levels:
The amount of NP-SH in the glandular stomach of control rats receiving water as inducing agent was 7.68 ± 0.55 µmol/g of tissue. Oral administration of ImI of ammonium hydroxide 1% caused significant depletion of NP-SH level by 46% i.e. they were decreased from 7.68 + 0.55 to 4.15 f 0.25 gmol/g of tissue. The reduced NP-SH levels were significantly reversed and restored towards control level by coadministration of sucralfate (0.125g or 0.250g/kg) together with honey (0.312g/kg) [Table - 2] .
| Discussion|| |
This study demonstrates that combination of a low dose (which itself has no significant effect) of honey with sucralfate, a clinically effective antiulcerogenic drug, potentates the gastro-protective effects of sucralfate against ammonia-induced gastric lesions. Furthermore, this gastric protection effects is associated with restoration of NP-SH level towards the normal control.
Our previous studies have shown that some of the mechanisms of action of honey against experimentally-induced acute and chronic gastric lesions are identical to those of sucralfate ,,,,, . Because of these similarities, it was of interest to establish whether sucralfate and honey might interact synergistically. In this context, synergism has been suggested to include either additive effect (combined effect of two drugs is equal to sum of their individual effect) or potentiation (where one drug increases the action of another drug -both of the drugs used may be active or one of the drug lacks any significant effect)  . Our recent observation that a very high dose of sucralfate (lg/kg) or honey (1.25g/kg) is required to achieve 63 and 69 % gastric protection, respectively, against ammonia-induced gastric lesions in rats  . The objective of this study was to investigate the effect of combination of a minimum amount of honey that may be acceptable to use in combination with sucralfate. Therefore, a single minimum effective dose of honey (0.312g/kg) and two sub maximal effective doses of sucralfate (0.125g/kg and 0.250g/kg) were selected from our previous study  and to observe the effect of combination on this model. For example, only 22g or 22ml of honey will be required for an adult weighing 70kg and that amount was thought to be acceptable in clinical use. Thus, combined use of a low dose of honey (0.312g/kg) and low doses of sucralfate (0.125g/kg or 0.250g/kg) resulted in potentiation of gastric protection effect of sucralfate by 58% and 77%, respectively. This finding may suggest that a suspension of relatively low dose of sucralfate (than that are used clinically) in honey may be beneficial in the management of peptic ulcer diseases in humans.
Sucralfate molecule contains, in addition to sucrose, eight molecules of sulfate and aluminum hydroxide. Since all the components of sucralfate i.e. sucrose, sulfate and aluminum are present in honey  , it is possible that honey may contain 'sucralfate-like' substance. However, there is no evidence to support this hypothesis at the present time. The enhancement of the gastro protective effects of sucralfate may be due to the combined effect of each free individual component of sucralfate that are present in honey since sucrose octasulfate, sulfate and aluminum chloride have been reported to possess gastric protection effects  . On the other hand, potentiation may also be due to the occurrence of `sucralfate - like' substance present in honey and thus increasing the concentration of sucralfate in the stomach. Furthermore, both honey and sucralfate are known to afford protection against ethanol or ammonia-induced gastric lesions by preventing depletion of NP-SH ,,,, . Thus, significantly higher NP-SH level attained by the combination of honey and sucralfate than those obtainable with either of them alone may suggest the possible role of NP-SH in the potentiation of sucralfate effect. Moreover, one of the suggested mechanism in the pathogenesis of ammonia-induced gastric lesions is the formation of cytotoxic oxidant monochloramine by the reaction of ammonia with hypochlorous acid . Previously, both honey and sucralfate have been shown to possess antioxidant effects in vitro  .
A concentration of 125(g/ml sucralfate caused 51% and a concentration of 3.12mg/mI honey produced 71% inhibition of generation of super oxide in vitro  Thus, it is possible that their antioxidant properties may also be involved in producing potentiation of the gastric protection effects of sucralfate .
| Conclusion|| |
In conclusion, the present study shows potentiation of gastroprotective effects of sucralfate by honey against ammonia-induced gastric lesions in rats. Since ammonia is considered to be one of the factors in the pathogenesis of H. pylori-induced gastric lesions, this finding may be of clinical value in the treatment of patients with peptic ulcer disease.
Acknowledgement: This work was supported by a grant from the College of Medicine Research Center, King Saud University, Riyadh, Saudi Arabia. We would like to thank Mr. Mazhar Khan for technical assistance and Mr. Afzal Hossain for secretarial help
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Abu Taib Mohammad Mobarok Ali
Department of Pharmacology, College of Medicine, King Saud University, P.O. Box 2925, Riyadh 11461
Source of Support: None, Conflict of Interest: None
[Table - 1], [Table - 2]