Rapid screening of two microcystins (i.e., microcystin-YR (MC-YR) and microcystin-LR (MC-LR)) in surface water samples was performed by a simple and eco-friendly procedure using deep eutectic solvent-based vortex-assisted liquid-liquid microextraction (DES-based VALLME) combined with ultrahigh-performance liquid chromatography and electrospray ionization (+)-quadrupole time-of-flight mass spectrometry (UHPLC-ESI(+)-qTOF-MS) detection. To obtain an efficient water-miscible DES, choline chloride and phenol at a molar ratio of 1 : 2 were used as an extractant for VALLME. To optimize factors of DES-based VALLME, response surface design alongside Box-Behnken design was used. The limits of quantitation (LOQs) were 0.5 ng mL-1 and 0.4 ng mL-1 for MC-YR and MC-LR, respectively, which is sensitive enough to meet the World Health Organization (WHO) maximum guideline level for MC-LR in water of 1.0 ng mL-1. Moreover, satisfactory precision with relative standard deviations (RSD) for both intra- and inter-day analysis lower than 11%, and trueness (also known as mean extraction recovery) ranged from 85.5 to 113% based on the ICH method validation guideline.