The modified natural biopolymers, recognized as environmentally friendly flocculants, are gaining tremendous attention in the field of water treatment. In this study, a novel hybrid flocculant with antibacterial activity, cinnamic acid (CA) conjugated N-(2-hydroxy)-propyl-3-trimethylammonium chitosan chloride (HTCC) derivative (HTCC-CA), was prepared via quaternary ammonium and amide reactions. The characterization, flocculation, and antibacterial activities were carried out to access the structural properties and potential application. The results of UV-vis, FT-TR, and 1H NMR confirmed the successful synthesis of HTCC-CA1-3 derivatives. XRD and TGA revealed the lower crystallinity and thermal stability of HTCC-CA1-3 derivatives than chitosan (CS). Bacterial flocculation and antibacterial tests indicated the excellent flocculation effect of HTCC-CA1-3 derivatives and showed high antibacterial activity for Escherichia coli suspension. Moreover, the mechanism of action of the derivatives was investigated via zeta potential measurements and scanning electron microscope, which can be summed up as the effective interaction between charges. The results proved that HTCC-CA derivatives are promising agents for wastewater treatment.