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The simultaneous voltammetric determination of melatonin (MT) and pyridoxine (PY) has been carried out at a cathodically pretreated boron‐doped diamond electrode. By using cyclic voltammetry, a separation of the oxidation peak potentials of both compounds present in mixture was about 0.47 V in Britton‐Robinson buffer, pH 2. The results obtained by square‐wave voltammetry allowed a method to be developed for determination of MT and PY simultaneously in the ranges 1–100 μg mL−1 (4.3×10−6–4.3×10−4 mol L−1) and 10–175 μg mL−1 (4.9×10−5–8.5×10−4 mol L−1), with detection limits of 0.14 μg mL−1 (6.0×10−7 mol L−1) and 1.35 μg mL−1 (6.6×10−6 mol L−1), respectively. The proposed method was successfully to the dietary supplements samples containing these compounds for health‐caring purposes.

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