Most investigations on the laser generation and fragmentation of nanoparticles focus on Feret particle size, although the hydrodynamic size of nanoparticles is of great importance, for example in biotechnology for diffusion in living cells, or in engineering, for a tuned rheology of suspensions. In this sense, the formation and fragmentation of gold colloidal nanoparticles using femtosecond laser ablation at variable pulse repetition rates (100–5000 Hz) in deionized water were investigated through their plasmon resonance and hydrodynamic diameter, measured by Dynamic Light Scattering. The increment of the repetition rate does not influence the ablation efficiency, but produces a decrease of the hydrodynamic diameter and blue-shift of the plasmon resonance of the generated gold nanoparticles. Fragmentation, induced by inter-pulse irradiation of the colloids was measured online, showing to be more effective low repetition rates. The pulse repetition rate is shown to be an appropriate laser parameter for hydrodynamic size control of nanoparticles without further influence on the production efficiency.