Purified polysaccharides from safflower (Carthamus tinctorius L.) post-extraction residues alleviate immunosuppression in mice by modulating gut microbiome.

A novel polysaccharide AES-S was extracted from safflower post-extraction residues using response surface methodology (RSM) and artificial neural network (ANN) optimization techniques, and was purified to obtain a homogeneous polysaccharide. Structural analysis revealed that AES-S is an RG-I-type pectic polysaccharide with a backbone of →4)-α-GalpA-(1 → 2)-α-Rhap-(1 → and a molecular weight (Mw) of 16.27 kDa. AES-S restored the immune homeostasis of cyclophosphamide (CTX)-induced immunosuppressed mice by promoting cytokine secretion, recovering T cell proliferation, increasing immunoglobulins and maintaining intestinal barrier integrity. Based on a pseudo-sterility mouse model and fecal microbiota transplantation (FMT), a crucial role of the gut microbiome and its associated metabolites in protective effects of AES-S against intestinal injury was confirmed. 16S ribosomal RNA (rRNA) and internal transcribed spacer (ITS) sequencing showed that AES-S remodeled the gut microbiome, enriching beneficial bacteriota (Corynebacterium) and mycobiota (Rhodotorula). Metabolomics further revealed key changes in immunomodulatory metabolites, including 5-methylthio-d-ribose and D-galactose 6-sulfate, as well as a marked increase in short-chain fatty acids (SCFAs). Correlation analyses linked these microbial shifts to metabolic recovery. These findings demonstrate that AES-S exerts immunomodulatory effects via the gut microbiome-metabolism axis, positioning it as a promising natural immunomodulatory agent.