Objective To investigate the protective effects of Ziziphus jujuba polysaccharides (ZJP) on immune suppression and intestinal barrier injury in hindlimb-unloaded (HU) mice. Methods Male ICR mice were randomly divided into six groups: control, HU model, levamisole, and ZJP low-, medium-, and high-dose groups (200, 400, and 800 mg/kg). All mice were orally administered for 14 consecutive days. Body weight and spleen index were recorded; histopathological changes in the spleen and colon were examined by hematoxylin eosin (HE) and alcian blue periodic acid Schiff (AB-PAS) staining. Serum levels of Interleukin-1β (IL-1β), Tumor Necrosis Factor-α (TNF-α), Interleukin-10 (IL-10), Immunoglobulin M (IgM), and Immunoglobulin G (IgG) were determined using enzyme-linked immunosorbent assay (ELISA). The messenger RNA (mRNA) expression of inflammation-related genes (Tlr4, Nlrp3, Nos2, Il1b, and Tnf ) and barrier-related genes (Tjp1 and Cldn1) in colon tissues was analyzed by real-time quantitative polymerase chain reaction (RT-qPCR). Results Compared with HU mice, ZJP treatment improved the reductions in spleen index (p<0.05). The medium ZJP dose markedly reduced IL-1β (48%, p<0.001) and TNF-α (43%, p<0.05), while the low dose increased IL-10 by 12.79% (p<0.001). Low and medium ZJP doses elevated IgM (22.54%, 50%) and IgG (70%, 262.61%), respectively (all p<0.05). All ZJP doses significantly downregulated inflammation-axis genes (Tlr4, Nlrp3, Nos2, Il1b, Tnf) and upregulated barrier-axis genes Tjp1 and Cldn1 (p<0.05). Conclusions ZJP markedly alleviates HU-induced immune suppression and intestinal barrier dysfunction potentially by inhibiting the Toll-like receptor 4/NOD-like receptor pyrin domain-containing 3 (TLR4/NLRP3) inflammatory signaling pathway, reducing proinflammatory cytokine production, upregulating tight junction protein-related gene expression, and restoring immunoglobulin levels. These findings provide experimental evidence for the potential application of jujube polysaccharides in spaceflight immune protection and functional food development.