Rapid Determination of 1-Deoxynojirimycin in Milk Powder by Precolumn Derivatization Combined withHigh Performance Liquid Chromatography with Fluorescence Detection
2025
DENG Sisi, QIN Hao, XIONG Meijin, ZHAO Hongyang, LIU Ling, HUANG Yuying, BAO Lei, LI Shaobo
A rapid precolumn derivatization combined with high performance liquid chromatography (HPLC) with fluorescence detection method for the determination of the 1-deoxynojirimycin (DNJ) content in functional milk powder. Samples were extracted with 60% methanol in water. In pH 8.5 borate buffer solution, o-phthalaldehyde was added as a derivatizing agent to remove the interference from primary amines. DNJ in samples reacted with 9-fluorenylmethoxycarbonyl chloride to form a complex with fluorescence absorption. The separation was performed on a C18 column (150 mm × 4.6 mm, 2.7 μm). Mobile phase A was 10 mmol/L sodium acetate buffer solution, and mobile phase B was acetonitrile. Gradient elution was performed at a flow rate of 1.5 mL/min. The column temperature was 45 ℃. The excitation wavelength was 275 nm. The emission wavelength was 320 nm. Good separation of DNJ from other components was achieved. The detection limit was 4 ng/mL, and the calibration curve was linear in the range of 0.01 to 50 μg/mL, with a determination coefficient (R2) > 0.999. The recovery rates ranged from 101.8% to 104.1%. The method was validated to be stable and reliable, and could be used as an effective method for determining the content of DNJ in milk powder.
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