Abstract: Nano-Cu(OH)₂, one of the most used nano-pesticide formulations, can be applied to replace pesticide. However, the hazardous effects of nano-Cu(OH)₂ on biological plant are still unclear. This study investigated the effects of various concentrations of nano-Cu(OH)₂ on the growth of lettuce (Lactuca sativa L.) in hydroponic culture for 4 weeks. Cu dominantly accumulated in the old leaves (target organs) of lettuce during the first 2 weeks, which resulted in greenness loss and yellow-turning of old leaves. Meanwhile, the Cu partially transferred from old leaves to new leaves (P<0.05) and root (P>0.05). Nano-Cu(OH)₂ stress caused irreversible damage to old leaves of lettuce during the last 2 weeks, reflected by the drying and yellow-shedding old leaves. Again, the Cu continually transferred from old leaves to the new leaves or roots of lettuce. Lettuce mediated the content of SOD and POD to resist the hazardous damage caused by Cu stress. Low concentration of nano-Cu(OH)₂ promoted the growth of lettuce, and high concentrations of nano-Cu(OH)₂ and Cu²⁺ inhibited the growth and development of lettuce. The results indicated that the underling toxicity of nano-Cu(OH)₂ on lettuce growth was derived from the free Cu ions. The study provides a new perspective of potentially hazardous effects of nano-Cu(OH)₂ on lettuce, and scientific basis for the safe use of nano-Cu(OH)₂.