As a kind of chelate with high affinity to Fe3+ produced by microorganisms and gramineous plants under iron deficiency conditions, the siderophore helps to promote the transformation of insoluble Fe3+ into soluble Fe2+ in the environment. In this current research, a high-yield siderophore microorganism, namely Ti-11, is isolated from the rhizosphere soil of plants, which is identified as Penicillium oxalicum capable of producing carboxylic acid siderophore with a maximum yield of 84.13% on the ninth day. By optimizing the single factor experimental conditions of microbial siderophore cultivation, the optimal carbon source is determined to be glucose 15 g/L, and the optimal nitrogen source is determined to be acid hydrolyzed casein 6 g/L with an optimal initial pH of 8.0. Finally, taking Escherichia coli and Staphylococcus aureus as experimental subjects, an investigation is made of the inhibitory effect of siderophores on common pathogenic bacteria. It is found that the supernatant of the siderophore can significantly inhibit Staphylococcus aureus, with the best antibacterial effect when treated for 24 hours.