An efficient and inexpensive method using vortex-assisted surfactant-enhanced emulsification microextraction (VASEME) based on solidification of floating organic droplet coupled with ultraperformance liquid chromatography–tandem mass spectrometry is proposed for the analysis of glucocorticoids in water samples (river water and hospital wastewater). VASEME was optimized by the experimental validation of Plackett–Burman design and central composite design, which has been co-related to experimental design. Plackett–Burman design showed that factors such as vortex time, surfactant concentration, and pH significantly affect the extraction efficiency of the method. Method validation was characterized by an acceptable calibration range of 1–1000 ng L−1, and the limit of detection was in the range from 2.20 to 8.12 ng L−1 for glucocorticoids. The proposed method was applied to determine glucocorticoids in river water and hospital wastewater in Lucknow, India. It is reliable and rapid and has potential application for analysis of glucocorticoids in environmental aqueous samples.