As one of the major pollutants in the environment, chromium (Cr), a heavy metal, poses a serious threat to urban green spaces and human life and health. Cosmos bipinnatus is considered a potential accumulator of Cr, and the differences in cellular Cr distribution and compartmentalization may uncover the mechanisms involved in its tolerance to Cr. To elucidate the effects of Cr stress on C. bipinnatus and determine the mechanism of Cr tolerance in C. bipinnatus, we investigated the physiological indicators, subcellular distribution and chemical forms, cell wall fractions and their Cr contents, uronic acid content in the cell wall fractions, and Fourier transform infrared spectroscopy (FTIR) of the cell wall. The results showed that the antioxidant enzyme activities in C. bipinnatus under Cr stress and most of the Cr were fixed in the cell wall. Notably, changes in the content of pectin fractions in the cell wall affected the accumulation of Cr in the cell wall of C. bipinnatus and the stability of negatively charged groups. In addition, the carboxyl and hydroxyl groups played a role in fixing metal in various parts of the C. bipinnatus cell wall.