Abstract: In the present work, starch/thymol nano-emulsions (S-T-NE) were prepared by enzymatic hydrolysis combined with dynamic high-pressure microfluidization technology. Three different methods including starch-DHPM-thymol-magnetic stirring (SDTM), starch-thymol-magnetic stirring-DHPM (STMD), and starch-thymol-DHPM-magnetic stirring (STDM) were selected to discuss the multi-dimensional structure and property discrepancies of S-T-NE. Results showed that S-T-NE particles exhibited the minimal aggregation with uniformly adsorption of thymol by STMD, which was benefited to ensure the excellent storage stability of S-T-NE. In the meantime, the S-T-NE prepared by STMD displayed the highest thymol loading rate (78.30%±1.73%) and drug loading capacity (223.71±3.02 μg/mg) compared with other methods. Furthermore, the growth of Escherichia coli and Staphylococcus aureus was obviously inhibited by S-T-NE, showing inhibition zone diameters with 16.15±0.19 mm and 13.98±0.26 mm, respectively. For another, the S-T-NE prepared by STMD exhibited remarkable iron reduction capability (1.36±0.04 mmol Trolox/L) and DPPH, ABTS⁺ free radical scavenging abilities (0.11±0.00, 0.43±0.00 mmol Trolox/L). These findings provided valuable technical support on the preparation and application of starch-based nanoemulison loaded with bio-active compounds.