Abstract: Objective: To investigate the alleviating effect of isochlorogenic acid (ICGA) from Stevia rebaudiana on oxidative stress-induced intestinal damage. Methods: The total polyphenol content in the S. rebaudiana extract was determined using the Folin-Ciocalteu colorimetric method, and the ICGA content was analyzed via high performance liquid chromatography (HPLC). The cell counting kit-8 (CCK-8) assay and biochemical kits were employed to measure the levels of malondialdehyde (MDA), total antioxidant capacity (T-AOC), catalase (CAT), superoxide dismutase (SOD), and glutathione peroxidase (GSH-Px). The optimal H2O2 concentration and treatment time were determined to establish an intestinal epithelial cell of oxidative stress injury. The safe concentration range of ICGA and its optimal protective concentration against H2O2-induced oxidative damage in intestinal porcine epithelial cells-jejunum 2 (IPEC-J2) were selected using the CCK-8 assay. The protective effects of S. rebaudiana ICGA on intestinal epithelial cells under oxidative stress were evaluated by assessing antioxidant indicators (MDA, T-AOC, CAT, SOD, and GSH-Px) and the relative mRNA expression levels of tight junction protein genes (Claudin-1, ZO-1, and Occludin), the anti-apoptotic gene Bcl-2, and the pro-apoptotic gene Caspase-3. Results: The total polyphenol content in the S. rebaudiana extract was 67.68%, and the total content of three ICGA isomers was 64.23%. An intestinal oxidative stress injury model with significantly decreased cell viability and antioxidant activity (P < 0.05) was established by treating the cells with 200 μmol/L H2O2 for 1 hour. Compared with the model group, the cell viability in the 50 and 100 μmol/L ICGA pretreatment groups increased from 77.49% to 90.23% and 95.99%, respectively. Additionally, the activities of CAT, SOD, and T-AOC were significantly enhanced, and the MDA content was markedly reduced (P < 0.05). ICGA pretreatment significantly up-regulated the relative mRNA expression of Claudin-1 and Occludin (P < 0.05), and significantly down-regulated the expression of Caspase-3 (P < 0.05), this effect being more pronounced at 100 μmol/L than at 50 μmol/L. In conclusion, S. rebaudiana ICGA may alleviate intestinal oxidative stress by enhancing antioxidant enzyme activity, reducing lipid peroxide accumulation, and improving intestinal permeability to enhance barrier function.

Key words: Stevia rebaudiana extract; isochlorogenic acid; gut health; hydrogen peroxide; oxidative stress