The production of non-toxic and homogeneous colloidal solutions of nanoparticles (NPs) for biomedical applications is of extreme importance nowadays. Among the various methods for generation of NPs, pulsed laser ablation in liquids (PLAL) has proven itself as a powerful and efficient tool in biomedical fields, allowing chemically pure silicon nanoparticles to be obtained. For example, laser-synthesized silicon nanoparticles (Si NPs) are widely used as contrast agents for bio visualization, as effective sensitizers of radiofrequency hyperthermia for cancer theranostics, in photodynamic therapy, as carriers of therapeutic radionuclides in nuclear nanomedicine, etc. Due to a number of complex and interrelated processes involved in the laser ablation phenomenon, however, the final characteristics of the resulting particles are difficult to control, and the obtained colloidal solutions frequently have broad and multimodal size distribution. Therefore, the subsequent fragmentation of the obtained NPs in the colloidal solutions due to pulsed laser irradiation can be utilized. The resulting NPs’ characteristics, however, depend on the parameters of laser irradiation as well as on the irradiated material and surrounding media properties. Thus, reliable knowledge of the mechanism of NP fragmentation is necessary for generation of a colloidal solution with NPs of predesigned properties. To investigate the mechanism of a laser-assisted NP fragmentation process, in this work, we perform a large-scale molecular dynamics (MD) modeling of FS laser interaction with colloidal solution of Si NPs. The obtained NPs are then characterized by their shape and morphological properties. The corresponding conclusion about the relative input of the properties of different laser-induced processes and materials to the mechanism of NP generation is drawn.