Stimuli-responsive microemulsions have recently attracted significant interest due to their unique properties. Here, we developed a novel surfactant-free microemulsion (SFME) in a nontoxic ternary mixture, in which dimethyl sulfoxide (DMSO) was used as an amphisolvent, n-butanol was used as a nonpolar phase, and water was used as a polar phase. The DLS results confirmed the presence of the preouzo zone, and the polarity experiment revealed that the single-phase region can be further divided into oil-in-water, bicontinuous, and water-in-oil subregions. The size of droplets increased upon increasing the water or n-butanol content but decreased with increasing DMSO content. With increasing temperature, the area of the single-phase region increased, accompanied by a decrease in the size of the droplets, and the critical point moved to the corner of n-butanol. No matter in what subregion the formulation was found, decreasing temperature to below the phase-transition temperature (PTT) will induce a transition from monophasic MEs to complete phase separation and vice versa. This is mainly attributed to the effect of temperature on the hydrogen-bond interaction. Ag nanoparticles (Ag NPs) can be prepared above the PTT and facilely separated below PTT. The Ag NPs obtained from the current SFME showed higher catalytic activity than that obtained from a common surfactant-based ME.