We report the synthesis of small zinc oxide nanoparticles (ZnO NPs) based colloidal suspensions and the study of second-harmonic generation from aggregated ZnO NPs deposited on glass substrates. The colloidal suspensions were obtained using the laser ablation of solids in liquids technique, ablating a Zn solid target immersed in acetone as the liquid medium, with ns-laser pulses (1064 nm) of a Nd-YAG laser. The per pulse laser fluence, the laser repetition rate frequency and the ablation time were kept constant. The absorption evolution of the obtained suspensions was optically characterized through absorption spectroscopy until stabilization. Raman spectroscopy, SEM and HRTEM were used to provide evidence of the ZnO NPs structure. HRTEM results showed that 5–8 nm spheroids ZnO NPs were obtained. Strong second-harmonic signal is obtained from random ZnO monocrystalline NPs and from aggregated ZnO NPs, suggesting that the high efficiency of the nonlinear process may not depend on the NPs size or aggregation state.