Rapid, accurate and efficient detection of acidic pH and citric acid (CA) changes is of great significance for predicting environmental and food safety problems by fluorescence analysis technique. Herein, a small molecule ratiometric fluorescent probe (BICL) based on benzoindole derivatives is successfully synthesized and characterized and used for quantitatively and qualitatively “turn-on” detection acid pH and CA changes in solution and environment by ultraviolet spectrum and fluorescence emission spectrum. On the one hand, the probe has a good linear relation to acidic pH in the pH range 3.1–4.5 (I604/I550 = 13.088–2.3878pH, R2 = 0.9986). On the other hand, the probe has a good linear relationship in the range of CA concentration of 14.0–23.0 μM (I604/I550 = 0.5324 [CA]-5.2628, R2 = 0.9993) and a low detection limit of 2.967 μM. BICL has a good recovery rate in the range of 114.6 ~ 101.0% and a low relative standard deviation (RSD) (0.0011 ~ 0.0092) in the determination of CA in real samples (water, drinks and fruits), which holds great potential for application in determination of CA in real samples. Importantly, the probe has good blood compatibility, and it has been successfully applied to detect exogenously induced changes in acidic pH and CA in zebrafish with great time-stability by using fluorescence imaging technology, respectively.