Montmorillonite (MMT), a natural absorbent agent, has widely been accepted for its antidiarrhea function in human and farm animals; however, its specific physicochemical property limits its biological function in practical use. In the current study, raw MMT was loaded by andrographolide, namely andrographolide loaded montmorillonite (AGP-MMT). The microstructure of AGP-MMT was observed by scanning electron microscope (SEM) and X-ray diffraction (XRD). The effect of AGP-MMT on the growth performance, intestinal barrier and inflammation was investigated in an enterotoxigenic Escherichia coli (ETEC) challenged mice model. The results show that the microstructure of MMT was obviously changed after andrographolide modification: AGP-MMT exhibited a large number of spheroid particles, and floccule aggregates, but lower interplanar spacing compared with MMT. ETEC infection induced body weight losses and intestinal barrier function injury, as indicated by a lower villus height and ratio of villus height/crypt depth, whereas the serum levels of diamine oxidase (DAO), D-xylose and ETEC shedding were higher in the ETEC group compared with the CON group. Mice pretreated with AGP-MMT showed alleviated body weight losses and the intestinal barrier function injury induced by ETEC challenge. The villus height and the ratio of villus height/crypt depth, were higher in mice pretreated with AGP-MMT than those pretreated with equal levels of MMT. Pretreatment with AGP-MMT also alleviated the increased concentration of serum tumor necrosis factor-α (TNF-α) and interleukin-1β (IL-1β), and the corresponding genes in the jejunum induced by ETEC infection in mice. The protein and mRNA levels of IL-1β were lower in mice pretreated with AGP-MMT than those with equal levels of MMT. The results indicate that AGP-MMT was more effective in alleviating intestinal barrier injury and inflammation in mice with ETEC challenge than MMT.