Microemulsion composed of Span20 + Tween20 isopropyl myristate (IPM) + H2O were investigated as potential drug delivery systems for eye drops. The system is important in that all its components are food grade so that the microemulsion is almost free of toxicity and irritation. The phase transition was investigated using the electrical conductivity measurements. The chloramphenicol is used to treat the eye diseases such as trachoma and keratitis. However, this drug in the common eye drops hydrolyzes easily. The main product of the hydrolysis is glycol. Here, the chloramphenicol was trapped into the oil-in-water (o/w) microemulsions free of alcohols. Its stability was investigated by the high performance liquid chromatography (HPLC) assays in the accelerated experiments of 3 months. The location of the chloramphenicol molecules in the microemulsion formulations was determined by means of dynamic light scattering (DLS) and 1H NMR spectroscopy. The results of HPLC revealed that the content of the glycols in the microemulsion formulation was much lower than that in the commercial eye drops at the end of the accelerated experiments. It implied that the stability of the chloramphenicol in the microemulsion formulations was increased remarkably. The results of DLS and NMR confirmed that the chloramphenicol molecules should be trapped into the hydrophilic shells of the microemulsion drops, which were composed of many oxyethylene groups. The benzene rings of the chloramphenicol molecules were near the group of alpha2-CH2 and the oxyethylene groups of the surfactant molecules. It was this reason that enabled the chloramphenicol molecules in the microemulsions to be screened from the bulk water and its stability to be increased remarkably.