A solid-phase extraction method was proposed for the preconcentration of Cu(II) in different samples in a mini-column packed with functionalized multi-walled carbon nanotubes (MWCNTs-COOH) as an effective sorbent, without using any complexing reagent, prior to its determination by flame atomic absorption spectrometry using response surface methodology. The experimental optimization step was performed by both a two-level full factorial design, with a center point, and a Box-Behnken design combined with response surface methodology. Three variables (pH, amount of Cu(II), and sample volume) were regarded as factors in the optimization. It was found that pH is the most significant factor affecting the preconcentration of Cu(II). The preconcentration factor was obtained as 100. The linear range was 1-5 mg L-1 (R-2 = 0.999). Under the optimized experimental conditions, the detection limit (3s) of the proposed method followed by FAAS was found to be 0.27 mu g L-1. The relative standard deviation for 10 replicate measurements of 50 and 100 mu g L-1 Cu(II) was 2.39% and 0.98%, respectively. The response surface methodology was successfully applied to the determination of Cu(II) in water samples and mussel samples, and in a certified standard reference material (BCR-320R, Channel sediment).