In the present work, polylactic acid (PLA) microcapsules as novel drug delivery systems were successfully fabricated by one-step processing using an electrospray technique. Curcumin (Cur) was chosen as model drug with satisfactory loading capacity (LC%) and entrapment efficiency (EE%) greater than 95%. By judiciously adjusting spinning solvents, flow rates, polymer and drug concentrations, the monodisperse and spherical structures of Cur/PLA microcapsules observed using scanning electron microscopy (SEM) and optical microscopy were successfully generated with diameter distribution ranging from 3.8 μm to 4.4 μm. The physical-chemical characterization including, FTIR, XRD, TG, and DTA, are explored and in vitro release profiles described by Ritger–Peppas models were also investigated, showing sustained release for 200 h after a burst release in the initial 12 h. The drug-loaded microcapsules showed excellent anti-bacterial activities towards Escherichia coli (E. coli) and Staphylococcus aureus (S. aureus) by using both disk diffusion and minimal inhibitory concentration methods. The anti-oxidant performance was also evaluated by using DPPH assays. In vitro cell tests, including Cell Counting Kit-8, hemolysis experiments, and cell adhesion revealed that PLA-based microcapsules had significant biocompatibility and low cytotoxicity. The study showed that the PLA-based electrospray strategy combined with spherical microcapsules has the potential for a broad range of applications in medical fields, especially in drug delivery.