Synchronous fluorescence spectra was applied to assess tyrosine (Tyr) and tryptophane (Trp) residues in the plasma of patients suffering hepatocellular carcinoma, tumor-bearing mice, and cultured cells (HepG2 and HL-7702). The synchronous fluorescence peaks of Tyr and Trp residues were at 318 and 350 nm when Δλ=20 nm and Δλ=80 nm respectively, and did not shift regardless of the sample variations. The results showed that the fluorescent intensities of Tyr and Trp residues increased significantly in the plasma proteins of patients suffering hepatocellular carcinoma. There was a correlation between the increased fluorescent intensities of murine plasma Tyr or Trp residues and the increasing of time of tumor-bearing, indicating that the alterations of Tyr and Trp residues might be associated with tumor development. On the contrary, the fluorescent intensities of Tyr and Trp residues in tumor tissue or HepG2 cells decreased along with the increasing of time of tumor-bearing or culturing. Further experiments showed that a potential anti-tumor medicine-matrine which exerted an anti-tumor activity could enhance the fluorescence intensities of Tyr and Trp residues. These results demonstrated that synchronous fluorescence spectra could be used to determine Tyr and Trp residues in proteins. By this method, we found that Tyr and Trp residues increased in the plasma proteins of tumor bearing patients and mice which might be correlated with tumor development. On the contrary, Tyr and Trp residues in proteins of tumor tissue or cancer cells decreased. It remains unclear whether these changes acted as a biomarker for protein metabolism imbalance or they contributed to the tumor evolution in malignant diseases, which is worthy of further investigation.