Water-soluble colloidal ZnSe quantum dots (QDs) modified by mercaptoacetic acid (MAA) were used to determinate H + in aqueous solutions by fluorescence spectroscopic technique. The results showed that the fluorescence of the water-soluble colloidal ZnSe QDs could be quenched by H + and the fluorescence intensity of the water-soluble QDs decreased linearly as the pH varied from 4.5 to 7.0. Based on this phenomenon, a convenient, rapid and specific method to determination of enzyme reaction kinetics was proposed. The modified ZnSe QDs were successfully used as pH probes in monitoring the hydrolysis of glycidyl butyrate catalyzed by porcine pancreatic lipase (PPL). The proposed method was found to improve stability, sensitivity and a monitoring range for determination of H + as compared to the already described analytical methods based on p-nitrophenoxide (PNP).