Innovative approaches for beetroot (Beta Vulgaris L.) aqueous extraction by cyclodextrins and its use to alleviate ethanol induced gastric ulcer in rats

Abstract

One of the most common chronic conditions of peptic ulcer is gastric ulcer (GU), which is recognized as a high-risk illness in the present-day lifestyle. Originating from Asia and Europe, Beetroot (Beta vulgaris L.) is packed with high amounts of bioactive compounds like betanin and phenolics. These contribute to its nutritional value and overall health benefits. In this work, varying concentrations (0.5% to 7% w/v) of beta-cyclodextrin (β-CD) and hydroxypropyl-beta-cyclodextrin (HP-β-CD) were used to improve the efficiency of extracting beetroot components with water, using both stirring and ultrasound techniques. The presence of 5% (w/v) HP-β-CD during extraction appeared the maximum values of total phenolic compounds and betanin (3.1 mcg/mL and 3.01 mg/mL, respectively). The extraction recoveries of betanin at 5% (w/v) HP-β-CD emerging with magnetic stirring were 73%, 36% and 50% against water, ethanol and β-CD, respectively. The reproducibility of extraction procedure was found to be 0.86% relative standard deviation (RSD) indicating the highest precision of the beetroot modified HP-β-CD extraction procedure. Then, rats that had been treated with either HP-β-CD-modified aqueous extract of beetroot, omeprazole (20 mg/kg, orally administered) or both were studied to assess whether they helped reduce ethanol-induced GU. The combined treatment of HP-β-CD modified beetroot extract with omeprazole brought a significant decrease in the increased levels of serum oxidative stress malonaldehyde and nitric oxide, inflammatory markers myeloperoxidase, interleukin-10, interleukin-6 and tumor necrosis factor-α. It also restored the decreased levels of antioxidant glutathione and cyclo-protective prostaglandin E2 in comparison to the positive control. Furthermore, the proposed combination of beetroot-modified aqueous extract and omeprazole exhibited less severe histopathological damage in comparison to the positive control. Therefore, a novel synergistic pharmaceutical treatment using HP-β-CD modified aqueous extract of beetroot and omeprazole was presented to enhance GU healing.

Keywords: Aqueous extraction; Beetroot (Beta Vulgaris L.); Cyclodextrins; Gastric ulcer; Rats.

Fig. 1

UV–Vis spectrum of the extracted beetroot samples

Fig. 2

Total betanin concentration of beetroot samples in β-CD presence with magnetic stirring and sonication. Data expressed as mean ± standard deviation (n = 3) and different letters show significant variation at 5% level in Duncan’s test (p < 0.05). W: Water, EtOH: Ethanol, CD1: 0.5% (w/v) β-CD, CD2: 1.0% (w/v) β-CD, CD3: 3.0% (w/v) β-CD, CD4: 5.0% (w/v) β-CD and CD5: 7.0% (w/v) β- CD

Fig. 3

Total betanin concentration of beetroot samples in the presence of HP-β-CD with magnetic stirring and sonication. Data are expressed as mean ± standard deviation (n = 3) and different letters show significant difference at the 5% level in Duncan’s test (p < 0.05). W: Water, EtOH: Ethanol, HPCD1: 0.5% (w/v) HP-β-CD, HPCD2: 1.0% (w/v) HP-β-CD, HPCD3: 3.0% (w/v) HP-β-CD, HPCD4: 5.0% (w/v) HP-β-CD and HPCD5: 7.0% (w/v) HP-β-CD

Fig. 4

Total phenolic content (TPC) of beetroot samples in the presence of β-CD with magnetic stirring and sonication. Data are expressed as mean ± standard deviation (n = 3) and different letters show significant difference at the 5% level in Duncan’s test (p < 0.05). Others as mentioned in Fig. 2

Fig. 5

Total phenolic content (TPC) of beetroot samples in the presence of HP-β-CD with magnetic stirring and sonication. Data are expressed as mean ± standard deviation (n = 3) and different letters show significant difference at the 5% level in Duncan’s test (p < 0.05). W: Water, EtOH: Ethanol, Others as mentioned in Fig. 3

Fig. 6

Light microscopic feature of gastric mucosa in different experimental gastric ulcer rat groups: Illustrative photomicrographs of gastric wall sections taken one hour after ethanol administration (H&E, X 400). A Gastric mucosa from normal rat showing normal surface columnar epithelia (black arrow), normal gastric gland (right bracket) with normal parietal cells (green arrow). The muscularis mucosa (double black arrow) appears normal, followed by submucosa with normal feature. B Gastric mucosa from ethanol ulcerated rat showing erosion of surface epithelia (black thin arrows), hemorrhagic necrosis of the gastric mucosa (white thick arrow), infiltration of inflammatory immune cells (orang arrows) and edema in submucosa (black thick arrow). C Gastric mucosa from ulcerated rat pretreated with beetroot extract showing slight improvement in histological appearance of gastric architecture in ethanol. The surface epithelia are still appeared with erosion (arrows) with greatly decrease in infiltration of inflammatory immune cells. D Gastric mucosa from ulcerated rat pretreated with omeprazole showing great improvement in gastric tissue architecture as shown by decrease in necrotic cells and gastric mucosa damage. E Gastric mucosa from ulcerated rat pretreated with the combination of beetroot extract and omeprazole showing more or less normal gastric tissue architecture. F Quantitative statistical analysis of histopathological HE staining. Values are expressed as mean ± SE of 5 rats. a: significance versus normal group; b: significance versus ethanol group; c: significance versus ethanol + beetroot, ***P < 0.0001, *P < 0.01