Effects of ethanol on the aerobic plate count, weight loss, sensory evaluation, relative electrical conductivity, total phenol content, chlorophyll content, polyphenol oxidase (PPO) activity, and phenylalanine ammonia-lyase (PAL) activity of fresh-cut broccoli were determined in this study. Effects of different ethanol concentrations on the physio-biochemical metabolism were analyzed. Results showed that the ethanol treatment could maintain the texture of fresh-cut broccoli, delay the nutrients loss, and restrain the activities of PPO and PAL to effectively improve the storage quality and extend the shelf life of fresh-cut broccoli.