Researchers have explored the ability of chitosan to form nanoparticles, to suit varying applications, ranging from wound-healing to gene delivery. Ionic gelation is a widely used method for formulating chitosan nanoparticles, where self-assembly plays a crucial role. This self-assembly is initially promoted by hydrophilic-hydrophobic parity amongst individual chitosan residues, along with electrostatic and Van der Waals interactions with the cross-linker. However, until now the intrinsic ability of chitosan to self-assemble is not widely studied; hence, we investigate the self-assembly of chitosan, based on proton balance between its protonated and deprotonated residues, to promote facile nanoparticle synthesis. This is one of the first reports that highlights subtle but critical influence of proton balance in the chitosan polymer on the formation of chitosan nanoparticles.