Gastroprotective and anti-diarrheic activities of menthol

AL Rozza, DAS Cesar, CH Pellizzon. UNESP, Morphology 18618-970, Brazil

Gastrointestinal diseases are global health problems, in which the gastric ulcer, diarrhea and constipation are the prevalent diseases. The present work aimed to evaluate and caractherize the gastroprotective and anti-diarrheic effects of (-)-Menthol (ME).

Male Wistar rats (n=7) weighing 200-250 g were used in all bioassays. Rats were orally treated with vehicle, carbenoxolone 100 mg/kg or ME 25, 50 or 100 mg/kg. After one hour, the rats were treated with 1 mL of ethanol, and after one additional hour the rats were euthanized and the stomachs were removed. The ulcer areas (mm2) were determined by the software AVSoft BioView and the lowest effective dose was chosen for the subsequent assays. Samples of these stomachs were destined to immunohistochemistry analysis (heat-shock protein-70 HSP-70 and vasoactive intestinal peptide VIP) and glutathione (GSH) measurement by ELISA, which included a sham group. In the next assay, the involvement of nitric oxide (NO), sulfhydryl compounds (SH), alpha-2 receptors and K+ ATP channels in the ME gastroprotective effect was determined by the intraperitoneal administration of the respective antagonists N-Nitro-L-Arginine-Methyl Ester (70 mg/kg), N-Methyl-Maleinide (7 mg/kg), yohimbine (3 mg/kg) and glibenclamide (3 mg/kg) 30 minutes before the oral treatments administration (vehicle or ME 50 mg/kg). The ulcer areas (mm2) were determined by the software AVSoft BioView. To evaluate the anti-diarrheic activity, rats were orally treated with vehicle, loperamide 3 mg/kg or ME 50 mg/kg 30 minutes before 1 mL of castor oil oral administration. Each group received an evacuation index (EI) according to the formula EI = 1× (number of solid feces) + 2× (number of semi-solid feces) + 3× (number of wet feces). To evaluate the intestinal motility, rats were orally treated with charcoal meal after treatment with vehicle, loperamide 5 mg/kg or ME 50 mg/kg. After 30 minutes, the rats were killed, the intestines were removed and the distance moved by the charcoal meal was measured (cm) and expressed as a percentage of the distance from the pylorus to the caecum. Results were expressed as mean ± s.e.m. and analyzed by Anova followed by Dunnett or unpaired t test, ap<0.01. All assays were approved by the UNESP Institutional Animal Care and Use Committee.

ME 50 mg/kg was the lowest effective dose, presenting 81.83% of gastroprotection (ulcer area 68.35 ± 28.30a) in comparison to the vehicle group (ulcer area 376.27 ± 14.99). The immunolabeled area for HSP-70 and VIP was higher in ME treated group than in control groups. ME (1264.87 ± 44.20a nmol/g) and the control groups presented decreased GSH levels in comparison to the sham group level (1774.18 ± 88.22). The gastroprotective effect of ME was reversed when the SH compounds were inhibited and the K+ ATP channels were blocked but not when the NO was inhibited or the alpha-2 receptors were blocked. The EI for ME was 8.3 ± 0.42a, inhibiting the evacuation index in 36.15% in comparison to the vehicle group (13.0 ± 0.84). ME inhibited the intestinal motility in 56.18% (17.55 ± 3.13a) in comparison to the vehicle group (40.05 ± 2.94).

ME presented gastroprotective activity against ethanol and the lowest effective dose was 50 mg/kg. The HSP-70, VIP, SH compounds and K+ ATP channels are part of the gastroprotective mechanism of action of ME, but GSH, NO and alpha-2 receptors are not involved. ME presented anti-diarrheic activity, inhibiting the intestinal motility.

Financial Support: CAPES and FAPESP