To investigate the potential immunomodulatory mechanism of low molecular weight seleno aminopolysaccharide，an untargeted metabonomics method based on ultra high performance liquid chromatography-time of flight mass spectrometry(UPLC-TOF-MS) was developed for analysis of the variations of spleen endogenous metabolites in Acathopagrus schlegelii after they were extracted with frozen methanol,then isolated with a C18 column and a HILIC column,respectively.The potential biomarkers were screened out by non-targeted mass spectrometry with XCMSplus software and Metabolite HR MS2 library,and the related metabolic pathways were analyzed by Metabo Analyst 4.0 website.Results showed that the spleen metabolites in the group fed with the low molecular weight seleno aminopolysaccharide were significantly different from those in the blank group,and 36 biomarkers were found and identified.Low molecular weight seleno-aminopolysaccharide could enhance the immune function of Acathopagrus schlegelii through 9 metabolic pathways.This study provides a scientific basis for elucidating the immunoenhancement mechanism of low molecular weight seleno aminopolysaccharide.