Gold nanoparticles (NPs) (GNPs) had been a hot spot in recent years and had been widely used in drug carriers. GNPs could be directly synthesized and showed very good physical and chemical properties. To analyze the mechanism of ulinastatin (UTI) based on GNPs in the proliferation and migration of breast cancer cells (BCC), UTI@GNPs was prepared and applied in the treatment of sub-cultured MDA-MB-231 human BCC. According to the different dosage regimens, human BCCs was divided into S0 group (normal saline), S1 group (200 U/mL UTI@GNPs), S2 group (700 U/mL UTI@GNPs) and S3 group (1,200 U/mL UTI@GNPs). The cell proliferative activity (PA), transfer ability (TA), heparanase (Hpa), insulin-like growth factor receptor 1 (IGFR1), platelet activating factor receptor (PAFR), type 1 hyaluronidase (HYAL1), and CD44v6 were detected IP: 182.75.50.34 On: Thu, 03 Feb 2022 07:18:15 and compared before and after administration. The results showed that the absorption peak of UTI@GNPs Copyright: American Scientific Publishers did not change much over time, basically floating around 535. After administration of UTI@GNPs, the levels Delivered by Ingenta of Hpa, HYAL1, IGFR1, CD44v6, and PAFR in the S1 group were much higher than those in the S2 and S3 group (P < 0.05), and the PA and TA of BCC decreased greatly. In addition, the cell apoptosis rate and invasion inhibition rate of S1 group at 24 and 48 hours after administration were greatly lower than those of S2 and S3 group, and the differences were meaningful in statistical level (P < 0.05). In short, UTI@GNPs showed small particle sizes and high stability, which was completely suitable for tumor drug delivery treatment. UTI@GNPs could effectively inhibit the PA and TA of human BCC, and its anti-tumor mechanism may be related to Hpa, and PAFR.