In this present work, to minimise the risk of contamination by biofilm of Staphylococcus aureus (S. aureus), D-amino acid (D-AA) and nutmeg essential oil (NEO) were used to synergistically eliminate S. aureus biofilm formation. Chitosan nanoparticles (CNPs) were introduced to encapsulate D-Pro, D-Phe and NEO for getting better chemical stability during processing and storage. The optimal D-AA/NEO@CNPs possessed high stability (particle size 155.3 nm, zeta potential 42.2 mV), high uniformity (polydispersity index 0.267) and superior antibiofilm performance. Then, for the application in soya bean products, electrospun nanofibrous membranes were introduced to immobilise the D-AA/NEO@CNPs. And the antibiofilm assay of D-AA/NEO@CNP-loaded nanofibrous membranes exhibited excellent antibiofilm activity on soya bean products in this study.