Four water-soluble benzylidene cyclopentanone dyes are synthesized for photodynamic therapy (PDT) by introducing different numbers of carboxylate groups on their periphery. Their photophysical and photochemical properties are investigated systematically. The results show that the water solubility of these dyes is significantly enhanced after modification. All dyes exhibit large two-photo absorption (TPA) cross-section in near infrared (IR) range, which can generate both superoxide anion radical and singlet oxygen species under visible illumination. One-photon PDT experiments indicate that four novel dyes displaye low dark cytotoxicity. However, only the dye Y1 presents excellent photodynamic activity, which highlights the role of moderate lipophilicity for PDT dyes. Further cellular experiments reveals that Y1 had obvious two-photon induced cytotoxic effect on human rectal cancer 1116 cells. The result indicates that Y1 is a potential photosensitizer for two-photon induced PDT.