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A sensitive and facile method for the determination of l-cysteine having important role in human health was reported.The structure of the prepared nano-adsorbent was investigated by FT-IR, XRD and SEM.The optimized conditions were obtained by experimental design methodology.In this research, a facile and selective method was described to extract l-cysteine (l-Cys), an essential α-amino acid for anti-ageing playing an important role in human health, from human blood plasma sample. The importance of this research was the mild and time-consuming synthesis of zinc organic polymer (Zn-MOP) as an adsorbent and evaluation of its ability for efficient enrichment of l-Cys by ultrasound-assisted dispersive micro solid-phase extraction (UA-DMSPE) method. The structure of Zn-MOP was investigated by FT-IR, XRD and SEM. Analysis of variance (ANOVA) was applied for the experimental data to reach the best optimum conditions. The quantification of l-Cys was carried out by high performance liquid chromatography with UV detection set at λ = 230 nm. The calibration graph showed reasonable linear responses towards l-Cys concentrations in the range of 4.0–1000 μg/L (r2 = 0.999) with low limit of detection (0.76 μg/L, S/N = 3) and RSD ≤ 2.18 (n = 3). The results revealed the applicability and high performance of this novel strategy in detecting trace l-Cys by Zn-MOP in complicated matrices.