The purpose of the present study was to compare the changes of infrared spectrum of breast cancer cell line MCF-7 treated by 5-fluorouracil with various concentrations and treating time. The results indicate that the relative intensity ratios of I2 920/I1 460 increased in the FTIR spectra of cell line MCF-7 while I1 400/I1 460, I1 080/I1 550 and I1 240/I1 550 decreased significantly within forty-eight hours, however, no regular change was observed during 48～72 hours. The relative intensity ratio of I1 640/I1 550 decreases along with the increase in the concentration of 5-fluorouracil. The changes of infrared spectra are consistent with the biochemical changes in breast cancer cell, and it can be the reference designators when assessing curative effect of breast cancer cell by FTIR spectrometry.