To remove the interference from tyrosine and phenylalanine in the determination of tryptophan by fluorescence method,The author established the determination method of free tryptophan in serum by derivative synchronous fluorescence.The results showed,when was 90 nm,the peak of derivative synchronous fluorescence wasn't disturbed by tyrosine and phenylalanine at 285 nm. The linear rang is 0 to 400 mol/L(Y=0287X0096,r=0999).The intraand interassay CV are 257% and 342%,respectively.The analytical recovery is 938% to 972%. The determination wasn't disturbed by hemoglobin(02550 g/L)and bilirubin(4275342 mol/L).Reference value of the tryptophan in serum from 30 healthy individuals were 7398726( s ). The method is simple,rapid,and precise,so it is a reasonable method for detection of tryptophan in serum.