Two types of long-chained pyridinium N-chloramines based on 4-hydroxypyridine were synthesized via poly-step strategy. The structure of precursor and corresponding N-chloramine was confirmed by NMR and HRMS, and antibacterial activity of synthetic N-chloramines was preliminarily tested against E. coli (ATCC 25922) and S. aureus (ATCC 25923) using N-chloramine IIb as control. Antibacterial data showed that the antibacterial activity of para-derivative of pyridinium N-chloramine is superior to that of meta derivative counterpart, and that antibacterial activity showed an increasing trend with the attached chain lengthens. Specifically, the odecyl chained (-C12H25) pyridinium N-chloramine (XIIId/XVe) present the towering antibacterial efficacy, probably due to ‘synergistic’ biocidal action between N-chloramine moiety and long-chained pyridinium center. Besides, N-chloramine XVe derived from N-alkylation exhibits even higher bactericidal activity comparatively, achieving so far the most efficacious N-chloramine antimicrobial.