A simple and environmentally friendly method to synthesize spiropyrazolo[3,4-b]pyridines derivatives starting from isatin, cyclic-1,3-diketone and 3-methyl-5-aminopyrazole was developed. With the optimized conditions for enzymes, solvents, enzyme loading and reaction time in hand, 9 compounds wereobtained in acceptable yields. Moreover, the investigation in fluorescent properties of these products showed their potential application in the field of new fluorescent material.