Protective Effect of a Polyherbal Traditional Formula Consisting of Rosa damascena Mill., Glycyrrhiza glabra L. And Nardostachys jatamansi DC., Against Ethanol-induced Gastric Ulcer

Abstract

"VARD" formula consisting of Rosa damascena Mill. (Rosaceae) petals, and rhizomes of Glycyrrhiza glabra L. (Papilionaceae) and Nardostachys jatamansi DC. (Valerianaceae), has been proposed for gastric ulcer in Iranian traditional medicine. We investigated the antiulcer activity of each plant separately and in combination. The biochemical and molecular functions of extracts were also evaluated. Each plant hydroalcoholic extract was standardized via determination of total phenolic and flavonoid contents, also via some phenolic compounds determination and specially glycyrrhizic acid in G. glabra by using HPLC. Rats received orally extracts of the plants (20, 40 and 80 mg/Kg) and "VARD" (45 mg/Kg) 1 h before ethanol administration. Two h after receiving ethanol, animals were sacrificed; the stomach was removed for macroscopic and microscopic assessment. Also heme-oxygenase-1, glutathione, and catalase were measured in the gastric tissue of the rats pretreated by "VARD" and dose of 20 mg/Kg of extracts. Among three extracts, R. damascena and G. glabra contained more total phenolic and flavonoid content respectively. Gallic acid was prominent compound in R. damascena. The extracts of R. damascena, G. glabra, and N. jatamansi significantly decreased ulcer index. ED₅₀ values were 8.2, 31.86 and 25.08 mg/Kg respectively. "VARD" significantly decreased ulcer index compared to 20 mg/Kg of G. glabra (p < 0.0001) and N. jatamansi (p < 0.001). Pretreatment with "VARD" and each plant extracts (20 mg/Kg) increased glutathione, catalse and heme-oxygenase-1 significantly (p < 0.05) in comparison with control group. Our findings indicate that "VARD" partly via antioxidant activity can be considered as an effective antiulcer formula.

Keywords: Antioxidant; Glycyrrhiza glabra; Heme-oxygenase-1; Inflammation; Nardostachys jatamansi; Peptic ulcer; Rosa damascena.

Figure 1

HPLC chromatogram of phenolic compounds 1: Gallic acid, 2: Syringic acid, 3: Quercetin, 4: Caffeic acid, 5: p-Cumaric acid, 6: Ferulic acid detected in RDHE (a), GGHE (b) and NJHE (c) hydroalcoholic extracts

Figure 2.

Gastric ulcer index induced by ethanol in animal groups pre-treated by RDHE (A), GGHE (B) and NJHE (C). The results are expressed as mean ± SEM. (n = 7). The animals received: vehicle (water), ranitidine (50mg/kg), and doses of plant extract (20, 40 and 80 mg/kg, respectively). Statistical comparison was performed using analysis of variance (ANOVA) followed by post hoc Newman keuls test (* p < 0.05, ** p < 0.01 and *** p < 0.001

Figure 3

Protective effects of the RDHE (A), GGHE (B) and NJHE (C) against gastric lesions induced by ethanol. The results are expressed as mean ± SEM. (n = 7). The animals received ranitidine (50 mg/kg), and doses of plant extract (20, 40 and 80 mg/kg, respectively). Statistical comparison was performed using analysis of variance (ANOVA) followed by post hoc Newman keuls test (* p < 0.05, ** p < 0.01 and *** p < 0.001

Figure 4

Ulcer index in animal group pre-treated by VARD formula compared with all other groups. Statistical comparison was performed using analysis of variance (ANOVA) followed by post hoc bonferroni test. Difference from control group: * p < 0.05, *** p < 0.001 and **** p < 0.0001

Figure 5

GSH content of rat gastric tissues in RDHE, NJHE, GGHE (20 mg/kg), VARD (45 mg/kg) and ranitidine (50 mg/kg). ** p < 0.01 and *** p < 0.001 significantly different from the control

Figure 6

CAT activity in rat gastric tissues in RDHE, NJHE, GGHE (20 mg/kg), VARD (45 mg/kg) and ranitidine (50 mg/kg). * p < 0.05, ** p < 0.01 and *** p < 0.001 significantly different from the control

Figure 7

Western blot analysis of RDHE, NJHE, GGHE (20 mg/kg), VARD (45 mg/kg) and ranitidine (50 mg/kg) effects on HO-1 levels in gastric tissue. Proteins separation was conducted on SDS-PAGE, and then proteins blotted, probed with anti-HO-1 antibody and reprobed with anti-β-actin antibody. The densities of HO-1 bands on emerged films were measured and the ratio calculated. *** p < 0.001 significantly different from the control. # p < 0.05 significantly different from NJHE