Indoxyl is an intermediate product in the production of indigo and indirubin. There are no studies on the inhibitory effect of indoxyl on microorganism. In this study, it was found that temperature-stress induced the synthesis of intracellular β-glucosidase, which had a great influence on the ratio of indican, indoxyl, indigo and indirubin in Strobilanthes cusia. At 100 ℃, only indican was detected, whereas a large amount of indoxyl was found at 50–70 ℃. HPLC analysis indicated that the use of indoxyl reduced glucose consumption and metabolite production by Staphylococcus aureus. The cells treated with indoxyl had irregular shapes, and fragments of cells were found in SEM images. The images of CLSM suggest that cell morphological changes would be caused by the inhibition of indoxyl on EMs formation. Moreover, indoxyl decreased intracellular ATP content and increased the NAD+/NADH ratio, which promoted the generation of H2O2, damaging cells.