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Background <p>Ulcerative colitis (UC) is a chronic inflammatory bowel disease involving oxidative stress and immune dysregulation. Natural phytochemicals with multi-target actions are gaining attention as complementary strategies. The aim of this study is to evaluate the protective effects of a combined ethanol extract of <i>Withania somnifera</i> (ashwagandha), <i>Glycyrrhiza glabra</i> (liquorice), and <i>Syzygium aromaticum</i> (clove) in dextran sulfate sodium (DSS)-induced UC in rats.</p> Methods <p>Seventy male Wistar rats were randomly allocated into seven groups: Negative Control, Positive Control (UC), Ashwagandha Extract (250&#xa0;mg/kg), Liquorice Extract (200&#xa0;mg/kg), Clove Extract (250&#xa0;mg/kg), Herbal Extract (ALC, 400&#xa0;mg/kg), and Reference Drug (sulfasalazine, SASP, 100&#xa0;mg/kg). High-performance liquid chromatography (HPLC) analysis was conducted to characterize major phenolic and flavonoid constituents present in the ethanol extracts of ashwagandha, liquorice, and clove. Disease activity index (DAI), colon length, oxidative stress markers (SOD, CAT, MDA), cytokines (IL-6, IL-1β, IL-10), serum biochemical markers, and histopathology were assessed. Molecular docking analysis of selected HPLC-identified phytochemicals against cyclooxygenase-2 (COX-2) was performed using CB-Dock2.</p> Results <p>DSS-induced UC caused significant weight loss, elevated DAI, colon shortening, impaired antioxidant defenses, increased pro-inflammatory cytokines (IL-6, IL-1β), elevated liver/kidney biomarkers, and severe histological injury. Individual herbal extracts demonstrated variable protective effects, whereas the combined extract demonstrated broader improvement across inflammatory, oxidative stress, biochemical, and histopathological parameters. The combined extract significantly reduced TNF-α, IL-1β, IL-6, MPO activity, MDA levels, and serum nitrite while restoring antioxidant enzyme activities, including SOD, CAT, and GPx. Histopathological examination revealed marked preservation of mucosal architecture and reduced inflammatory infiltration in the combined extract-treated group. ALC treatment significantly restored antioxidant enzymes, reduced lipid peroxidation and pro-inflammatory cytokines, increased IL-10, normalized biochemical markers, and preserved colonic architecture, with improvements approaching those observed in the SASP-treated group for several measured parameters. HPLC analysis identified several major phenolic and flavonoid compounds, including quercetin, rutin, kaempferol, apigenin, diosmin, and eugenol derivatives. Molecular docking demonstrated favorable predicted binding affinities of several identified phytochemicals toward the COX-2 active site.</p> Conclusion <p>The combined ALC extract showed antioxidant and anti-inflammatory protective effects in experimental UC and showed predicted in silico interactions with the COX-2 active site. These findings support its potential as a complementary experimental approach, while further mechanistic, toxicological, pharmacokinetic, and clinical investigations are required before therapeutic translation.</p> Graphical Abstract <p></p>

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Protective Role of Individual and Combined Ethanolic Extracts of Ashwagandha, Liquorice, and Clove in DSS-Induced Ulcerative Colitis in Rats

  • 期刊论文

  • 发表日期：

    2026年6月23日

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  • Dalia Elebeedy,
  • Dalal Sulaiman Alshaya,
  • Aml Ghanem,
  • Emad Gamil Khidr,
  • Aya Salem,
  • Maha Ali Alghamdi,
  • Haitham Ibrahim El-Mekkawy,
  • Amina Rashad Ali,
  • M. Yasser Alsedfy,
  • Eman Fayad,
  • Osama A. Mohammed,
  • Elwathiq Khalid Ibrahim,
  • Ahmed S. Doghish

摘要

Background

Ulcerative colitis (UC) is a chronic inflammatory bowel disease involving oxidative stress and immune dysregulation. Natural phytochemicals with multi-target actions are gaining attention as complementary strategies. The aim of this study is to evaluate the protective effects of a combined ethanol extract of Withania somnifera (ashwagandha), Glycyrrhiza glabra (liquorice), and Syzygium aromaticum (clove) in dextran sulfate sodium (DSS)-induced UC in rats.

Methods

Seventy male Wistar rats were randomly allocated into seven groups: Negative Control, Positive Control (UC), Ashwagandha Extract (250 mg/kg), Liquorice Extract (200 mg/kg), Clove Extract (250 mg/kg), Herbal Extract (ALC, 400 mg/kg), and Reference Drug (sulfasalazine, SASP, 100 mg/kg). High-performance liquid chromatography (HPLC) analysis was conducted to characterize major phenolic and flavonoid constituents present in the ethanol extracts of ashwagandha, liquorice, and clove. Disease activity index (DAI), colon length, oxidative stress markers (SOD, CAT, MDA), cytokines (IL-6, IL-1β, IL-10), serum biochemical markers, and histopathology were assessed. Molecular docking analysis of selected HPLC-identified phytochemicals against cyclooxygenase-2 (COX-2) was performed using CB-Dock2.

Results

DSS-induced UC caused significant weight loss, elevated DAI, colon shortening, impaired antioxidant defenses, increased pro-inflammatory cytokines (IL-6, IL-1β), elevated liver/kidney biomarkers, and severe histological injury. Individual herbal extracts demonstrated variable protective effects, whereas the combined extract demonstrated broader improvement across inflammatory, oxidative stress, biochemical, and histopathological parameters. The combined extract significantly reduced TNF-α, IL-1β, IL-6, MPO activity, MDA levels, and serum nitrite while restoring antioxidant enzyme activities, including SOD, CAT, and GPx. Histopathological examination revealed marked preservation of mucosal architecture and reduced inflammatory infiltration in the combined extract-treated group. ALC treatment significantly restored antioxidant enzymes, reduced lipid peroxidation and pro-inflammatory cytokines, increased IL-10, normalized biochemical markers, and preserved colonic architecture, with improvements approaching those observed in the SASP-treated group for several measured parameters. HPLC analysis identified several major phenolic and flavonoid compounds, including quercetin, rutin, kaempferol, apigenin, diosmin, and eugenol derivatives. Molecular docking demonstrated favorable predicted binding affinities of several identified phytochemicals toward the COX-2 active site.

Conclusion

The combined ALC extract showed antioxidant and anti-inflammatory protective effects in experimental UC and showed predicted in silico interactions with the COX-2 active site. These findings support its potential as a complementary experimental approach, while further mechanistic, toxicological, pharmacokinetic, and clinical investigations are required before therapeutic translation.

Graphical Abstract