Spherical phospholipid-bilayers, vesicles, were prepared using the layer-by-layer double emulsion technique, which allows the bilayer to be formed asymmetrically. On the outer layer of the vesicles, the phospholipase D (PLD) reacted to convert phosphatidylcholine (PC) to phosphatidic acid (PA). The reaction induced the curvature change of the vesicles, which eventually led to rupture. The response time from the time of PLD injection to the time of rupture was measured against different vesicle curvatures and the outer layer phase, using the fluorescence intensity change of a pH-sensitive dye encapsulated within the vesicles. The effect of the vesicle curvature on the response was observed to be more significantly dramatic at the solid phase, compared to the liquid phase. Furthermore, in the solid phase, the response time was faster for 80 and 155 nm vesicles and, slower for 605 nm vesicles than similarly sized vesicles in the liquid phase vesicles. This difference in the response time was thought to result from the configuration determined by the phase difference and the PLD behavior.