Abstract: In Kolthoff buffer at pH 10.7,Pd(Ⅱ) and acridine red could form a complex,which quenches the fluorescence intensity of acridine red.After adding L-isoleucine,a more stable complex was formed and was free acridine red release,which significantly enhanced the fluorescence intensity of the system.Based on this fact,a fluorospectrophotometric method with fluorescence probe of complex of Pd(Ⅱ) and acridine red was proposed for the determination of L-isoleucine.The excitation slit width was 5 nm and emission slit width was 10 nm.The reaction time was 120 min.The linearity range of L-isoleucine 0.125-1.375 mg·L-1,with detection limit (3s/k) of 0.27 μg·L-1.The complexation ratio of Pd(Ⅱ) to L-isoleucine was 3∶1.The recovery rates measured by standard addition method were in the range of 106%-114%,with RSD (n=6) less than 4.0%.