Colon cancer is associated with a high incidence and a poor prognosis. The aim of the present study was to determine whether Fourier transform infrared (FTIR) microspectroscopy can be used to monitor the chemotherapy drug‑induced apoptosis of SW620 colon cancer cells. The 50% inhibitory concentration (IC50) of 5‑fluorouracil (5‑FU), the main chemotherapeutic agent used for the treatment of colorectal cancer, was determined as the inhibition of growth of the SW620 cells using an MTT assay. Cell starvation and 5‑FU treatment synergized to arrest the cells in the G1 and S phases of the cell cycle. FTIR combined with fluorescence activated cell sorting (FACS) analysis were used to analyze the SW620 cells following treatment with 5‑FU for 12, 24 and 48 h. The apoptotic cells had several spectral characteristics. The relative peak intensity ratio (I1740/I1460) was significantly increased (P<0.05), the I1740/I1460 ratio, associated with a band of amino acid residues at 1,410 cm‑1 was significantly increased at the early and late phases of cell death (P<0.05), the peaks at 1,240 cm‑1 increased in wave number, a band at 1,040 cm‑1, associated with polysaccharides, appeared at 24 and 48 h and then moved to a higher wave number and the I1040/I1460 ratio increased at the late stage of apoptosis. These results demonstrated that FTIR can be used as a label‑free technique to monitor cancer cell apoptosis and to understand the spectral fingerprints of apoptotic cells. This suggested that FTIR spectral features have potential as a powerful tool to monitor cancer cell apoptosis.