In this paper, two guanidine derivative polymers were synthesized by reacting polyhexamethylene guanidine hydrochloride (PHGH) with starch and epichlorohydrin (EP) respectively and used as potential functional additives in papermaking. The molecular structure of the guanidine derivative polymers were characterized by FTIR and 2D NMR. PHGH- modified starches exhibited high antimicrobial activities against E. coli and the growth inhibition reached almost 100% in the presence of 1.0 wt.% PHGH in wood fibers. UV260 absorption was applied to reveal the dynamic antimicrobial process of guanidine derivative polymer obtained with PHGH and EP. Results showed that most of bacteria was deactivated within a few mins and the intracellular components were leaked when the concentration of the guanidine derivative polymer were higher than the minimum inhibitory concentration (MIC).