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Sulfated polysaccharides derived from seaweeds have important biological activities. The aim of this work was to demonstrate the anti-inflammatory effect of a sulfated polysaccharide extracted from the seaweed Digenea simplex (Wulfen) C. Agardh 1822 on 2,4,6-trinitrobenzene sulfonic acid-induced colitis in rats. After inducing colitis, rats were euthanized, and the colon was excised to determine macroscopic lesion scores and wet weight and to histologically evaluate and quantify glutathione, malondialdehyde, myeloperoxidase, nitrate/nitrite, and cytokines. The polysaccharide, composed of β-D-galactose and 3,6-α-L-anhydrogalactose residues, reduced wet weight as well as macroscopic and microscopic lesion scores. It also reduced myeloperoxidase activity; reduced proinflammatory cytokines, malondialdehyde, and nitrate/nitrite levels; and preserved glutathione consumption in the colon. According to these results, we can infer that the sulfated polysaccharide derived from D. simplex has an anti-inflammatory effect on induced colitis in vivo.