Plant polysaccharides can be used as bioactive natural polymers that provide health benefits, however high molecular weight neutral polysaccharides have not shown good bioactivity. In this study, high molecular weight neutral arabinogalactan was isolated and structurally characterized to investigate it antioxidant activity against IEC-6 cells. In this study, a neutral polysaccharide (AG-40-I-II) was obtained from the roots of Larix gmelinii (Rupr.) Kuzen. and purified using ethanol fractional precipitation and purification on a DEAE-52 cellulose column and a Superose 12 gel filtration column. The structural characteristics of AG-40-I-II was detected by chemical and spectroscopic methods. The results showed that the average molecular weight of AG-40-I-II was 18.6 kDa, the main chain was composed of →4)-β-D-Gal-(1, → 4, 6)-β-D-Gal-(1 and →4)-β- D-Glc-(1, the side chain is composed of T-β-L-Araf(1 → 6). The effect of AG-40-I-II on H2O2-induced IEC-6 cell injury was determined by MTT method. Besides, AG-40-I-II could reduce the level of MDA and increase SOD activity on IEC-6 cells, which could significantly inhibit the production of ROS. Importantly, AG-40-I-II inhibited the splicing of XBP1 by IRE1α through the ERS pathway and reduced the cell apoptosis induced by H2O2. In summary, the results of this study indicate that AG-40-I-II, as a natural source of plant polysaccharides, has good antioxidant activity, and is expected to become a safe plant source of natural antioxidants, which has great potential in biomedicine potential.