T/BDAS 002-2023 Determination of Melatonin Concentration in Serum by Liquid Chromatography-Tandem Mass Spectrometry (LC-MS/MS)

Scope

Scope: This document describes the liquid chromatography-tandem mass spectrometry method for the determination of melatonin content in serum. This document is applicable to the determination of melatonin in serum. The detection limit of the detection method described in this document is 0.01 μg/L, and the quantification limit is 0.05 μg/L. Main technical content: This standard describes the liquid chromatography-tandem mass spectrometry method for the determination of melatonin content in serum, which is suitable for the determination of melatonin in serum. Extraction method: Accurately transfer 1 mL of the sample to a 15 mL centrifuge tube, add 50 μL of deuterated melatonin standard working solution, add 4 mL of methanol, and shake and extract with a vortex mixer for 5 minutes. Centrifuge at 10000 g and 4 °C for 5 minutes. Take the supernatant and filter through a 0.22 μm filter membrane for liquid chromatography-tandem mass spectrometry determination. Technical point 1: Methanol as a solvent for melatonin not only does not affect the stability of the target substance in the sample, but also can cause protein denaturation and precipitation in serum. The experimental operation is simple, and the recovery rate can reach more than 60%, which can well meet the requirements for the detection of melatonin in serum. Technical point 2: After melatonin in serum is extracted with methanol, it is determined by liquid chromatography-tandem mass spectrometry using an internal standard method. This standard specifies the detection limit of melatonin as 0.01 μg/L, and the quantification limit is 0.05 μg/L; the recovery rate of melatonin in this standard method is 60~120%, which meets the recovery range at a certain concentration level according to GB/T 27404; and the relative deviation (RSD) is ≤20%.

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