ABSTRACT: The optimization of wheat starch esterification (acetylation) with a high degree of substitution was performed through response surface methodology (RSM) via various concentrations of reagents (acetic anhydride), pHs, and temperatures under various ultrasonication frequencies (25, 40, and 25 + 40 kHz). According to RSM methodology, optimized samples were selected by achieving high degrees of substitution at various frequencies, temperatures, and pHs. Solubility, swelling, X-ray, RVA, DSC, freeze-thaw stability, texture, and SEM analysis of the optimized samples were performed at three frequencies. X-ray pattern exhibited a more significant reduction in the crystallinity percentage of esterified starch at frequency 25 + 40 kHz compared with 25 kHz, 40 kHz, and native starch. According to DSC analysis, To, Tp, Tc, and enthalpy of gelatinization (?H gel) were lower in AC at frequency 25 + 40 kHz compared with AC at frequency 25 and 40 kHz and N starches. According to morphology analysis, in acetylated starches at 25 and 40 kHz, the surfaces and small granules underwent more damage, whereas in 25 + 40 kHz, large granules were more affected than small granules. Upon acetylation, freeze-thaw stability and textural properties of the starch significantly increased and decreased, respectively. The peak and final viscosity of acetylated starch increased (25 + 40 kHz ? 25 kHz ? 40 kHz ? N starch).