Phytotoxicity of copper (Cu) depends on the relative distribution of different chemical forms, which is a function of soil properties, such as soil pH and organic matter content. Sequential fractionation was conducted to partition the total Cu into exchangeable, sorbed, organically bound, precipitate, and residual forms. Three soils were sampled from existing citrus groves and 0 to 400 mg Cu kg−1 were added. The soils used were: Myakka fine sand (sandy, siliceous, hyperthermic Aeric Haplaquods; pH = 5.7), Candler fine sand (hyperthermic, uncoated, Typic Quartzipsamments; pH = 6.5), and Oldsmar fine sand (sandy, siliceous, hyperthermic Alfic Arenic Haplaquods; pH = 8.2). Phytotoxicity of added Cu was evaluated using citrus rootstock (Swingle citrumelo) seedlings grown for 330 d. In Cu-unamended soils, the major portion of the total Cu was in the organically bound form in the low pH soils. However, in the high pH soil, the precipitate form was the dominant form. As the rate of Cu increased, the concentration of the readily soluble Cu forms (exchangeable + sorbed forms) increased in the low pH soils, that is, from 0.8 to 89.5 mg kg−1 (8.4–25.3% of total Cu) in the Myakka soil, and from 2.2 to 70.3 mg kg−1 (3.1–20.3% of total Cu) in the Candler soil. In the high pH Oldsmar soil, however, the concentration of readily soluble Cu forms increased only from 1.1 to 5.3 mg kg−1 In relation to the total Cu content this was equivalent to a decrease from 5.2 to 1.5%. The citrus seedling growth was negatively correlated with Cu concentrations in the readily soluble forms and positively correlated with those of the precipitate form. A 20% decrease in the top and root weights occurred at 2.5 mg kg−1 of readily soluble Cu in the Candler soil (pH = 6.5). The critical concentration was lower (1.7 mg kg−1) for root growth on the Myakka soil (pH = 5.7). The critical Cu concentration in the leaves varied from 60 to 68 mg kg−1, while that in the roots was 62 mg kg−1 in the Myakka soil, but increased to 270 mg kg−1 in the Candler soil. This study demonstrated that the readily soluble form of Cu is the most phytotoxic, and an increase in the precipitate form is, thus, responsible for a reduction in Cu phytotoxicity.