Sea cucumber is a food with nutritional benefits distributing mainly in Asia, and Apostichopus japonicus (A. japonicus) is a kind of sea cucumber whose quality is better than any others. However, different processing methods make various effect on its quality. In this study, we evaluated the protection effect of A. japonicus with different processing methods on mice with ulcerative colitis induced by trinitrobenzene sulfonic acid (TNBS), especially on the intestinal microflora. The expression of IFN-γ/IL-4 and IL-1β in gut, and intestinal microbiota were discussed. The results revealed that three different processing methods of A. japonicus could decrease the expression of inflammatory cytokines, except for the expression of IFN-γ/IL-4 treated with enzymatic, and dried A. japonicus was the most efficient. A. japonicus could change the microbiotic imblance relatively back to normal in terms of bacterial diversity and composition, meanwhile increase the abundance of Bifidobacteria, Lactobacillus and Clostridium leptum. The elements of protein, polysaccharide in dried, instant, enzymatic A. japonicus are 73.09%, 65.06%, 57.42% and 6.72%, 5.46%, 5.45% respectively. This study indicated that A. japonicus have a good improving effect on ulcerative colitis, especially on the microbiome, and processing methods had an effect on alleviation of ulcerative colitis, which might be associated with content of protein and polysaccharide.