In this study, a deep eutectic solvent based on thymol as a hydrogen bond donor and 4-(dimethylamino)benzaldehyde as a hydrogen bond acceptor was synthesized and characterized by various methods for the first time. The new solvent is stable in the presence of water and can be considered as a hydrophobic extractant. In the presented work, this solvent was utilized for the derivatization and microextraction of urea for its determination in human urine and powdered milk samples. In the developed procedure, water-soluble analyte reacts with the hydrogen bond acceptor (4-(dimethylamino)benzaldehyde) with the formation of a colored derivative (Schiff base) followed by its mass-transfer into deep eutectic solvent phase. Combination of derivatization and extraction stages allowed to reduce sample pretreatment time and simplify the procedure for the determination of urea. The analysis time was 3 min. The limits of detection, calculated from the blank tests based on 3σ, were 0.1 mg L^-1 for human urine and 2 mg kg⁻¹ for powdered milk, RSD values were < 7 % and the enrichment factors were 24 and 23 for urine and milk samples. The procedure was successfully applied to the analysis of real samples.