Two novel ferrocene-appended coumarin derivatives, FcL1 and FcL2, were synthesized and characterized using various analytical techniques which includes IR, UV, 1H NMR, 13C NMR and mass spectroscopy. 1H NMR revealed signals for the substituted cyclopentadiene ring at δ 4.80 and δ 4.61 ppm (FcL1) and δ 4.81 and 4.60 ppm (FcL2), while 13C NMR confirmed azomethine carbons at δ 162.95 and 160.43 ppm, respectively. UV–Vis analysis confirmed the successful molecular design of FcL1 and FcL2, achieving the desired electronic properties. Cytotoxic and antiproliferative effects of FcL1 and FcL2 were evaluated against metastatic colorectal carcinoma (CRC) DLD-1 cells and healthy colon epithelium CCD-18Co cells using the Alamar Blue assay. Both compounds inhibited DLD-1 cell proliferation in a dose-dependent manner, with IC50 values of 64.3 ± 2.4 µM (FcL1) and 67.1 ± 3.1 µM (FcL2). Minimal cytotoxic effects were observed on healthy colon epithelial cells at the same concentrations (p < 0.001). Fluorescence microscopy revealed the localization of both compounds in the cytoplasm and nucleus of cells. These findings suggest FcL1 and FcL2 as potential candidates for colorectal cancer therapy, exhibiting selective cytotoxicity toward cancer cells with low toxicity to healthy cells.