A rapid and high-throughput analytical method was developed for the determination of vitamin K1 using photo-induced chemiluminescence(PICL) detection.This method is based on ultraviolet irradiation(254 nm/15 W) of vitamin K1 to produce a fluorescent product which could be determined by peroxideoxalate chemiluminescence system using chemiluminescence detector.Influence of imidazole on photochemical product structure and chemiluminescence mechanism were studied.Under the optimum conditions，a good linear relationship between chemiluminescence intensity and concentration of VK1 over the range of 0.007-3.333 μg?mL-1 was obtained.The detection limit(S/N=3) for vitamin K1 was 2.2 ng?mL-1，and the relative standard deviation(c=01 μg?mL-1 ，n=11) was 2.7%.The method could be used to analyze 96 samples in 10 min,and was successfully applied in the determination of vitamin K1 in injection and human serum by standard addition calibration，with recoveries of 96.0%-104.0%.The results showed that the PICL method was rapid，high-throughput，simple，sensitive and selective,and was especially suitable for the analysis of a large number of samples.