The invention relates to a preparation method and an application of a double-rate-type (ultraviolet/fluorescence) multifunctional (calorimetric/fluorescence/ultraviolet) high-sensitivity florescent probe for carboxylesterase detection. Firstly, 2,4-dihydroxybenzaldehyde, diethyl glutaconate and anhydrous piperidine react in ethyl alcohol to produce a product 1, the product 1 and acetic anhydride react in anhydrous pyridine to produce a product 2, the product 2 reacts with osmium tetroxide and sodium periodate in tetrahydrofuran to produce a product 3, and the product 3 reacts with anhydrous potassium carbonate in methyl alcohol to produce a product 4; then methylpyridine and methyl iodide react in absolute ether to produce a product 5; next, the product 4 and the product 5 are dissolved in absolute ethanol, piperidine is added for a reaction, and a product 6 is obtained; finally, the product 6 is dissolved in acetic anhydride, anhydrous sodium acetate is added, and the double-rate-type multifunctional florescent probe for carboxylesterase detection is obtained. The probe is applicable to qualitative and quantitative analysis of carboxylesterase in biological samples, is sensitive, accurate and quick in detection, and can be applied to related fields of analytical chemistry, life organic analytical chemistry, disease pre-diagnosis, clinical medical examination and the like.