An analytical method, solid-phase extraction combined with dispersive liquid–liquid microextraction (SPE-DLLME), is established to determine carbamazepine in biological fluids and water samples. After concentration and purification of the samples in SPE C₁₈ sorbent, 1.5 mL of acetonitrile containing 60.0 μL of chloroform was injected into 5.0 mL pure water. After extraction and centrifuging, the sedimented phase was evaporated and the residue was dissolved in 30 μL methanol and injected into the HPLC system. Some important extraction parameters, such as sample solution flow rate, sample pH, type and volume of extraction and disperser solvents as well as the salt addition, are studied and optimized. The new method (SPE-DLLME) provides detection limits of 0.8 μg L⁻¹ and 1.7 μg L⁻¹ in urine and plasma samples, respectively. The calibration graphs are linear in the range of 2.5–500 μg L⁻¹ and 5.0–500 μg L⁻¹ in urine and plasma, respectively. The results show that SPE-DLLME is a suitable method for the determination of carbamazepine in biological and water samples.