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目的:建立固相萃取结合HPLC-荧光法测定人血浆中维生素B2含量的方法。方法:采用SiroccoTM96孔蛋白沉淀板和正压固相萃取装置处理样品,采用SymmetryshieldTMRP-C18(250 mm×4.6 mm,5μm)色谱柱,流动相为甲醇-5 mmol.L-1乙酸铵(35∶65),流速为0.6 mL.min-1,荧光检测器检测(激发波长:450 nm,发射波长:520 nm)。结果:维生素B2线性范围为0.2～10 ng.mL-1(r=0.9937)。定量下限为0.17 ng.mL1,日内精密度(RSD)为2.0%～4.8%,日间精密度(RSD)为3.8%,方法回收率为101.5%～116.0%(n=3),提取回收率为91.8%～115.1%。结论:所建方法采用96孔蛋白沉淀装置自动处理样品,高效液相色谱-荧光法测定,可用于临床血浆样品中维生素B2含量的测定。
Objective: To establish a method for the determination of vitamin B2 in human plasma by solid phase extraction combined with HPLC-fluorescence method. METHODS: The samples were treated with Sirocco 96 porcine protein pellet and positive pressure solid-phase extraction device. The mobile phase was Symmetryshield TMRP-C18 (250 mm × 4.6 mm, 5 μm) with methanol-5 mmol·L-1 ammonium acetate 65) at a flow rate of 0.6 mL.min-1. Fluorescence detector detection (excitation wavelength: 450 nm, emission wavelength: 520 nm). Results: The linear range of vitamin B2 was 0.2 ~ 10 ng.mL-1 (r = 0.9937). The limit of quantification was 0.17 ng.mL1. The intra-day precision (RSD) ranged from 2.0% to 4.8%. The intraday precision was 3.8%. The method recoveries ranged from 101.5% to 116.0% (n = 3) 91.8% ~ 115.1%. CONCLUSION: The established method uses a 96-well protein precipitation device to automatically process samples and determine the contents of vitamin B2 in clinical plasma samples by HPLC-fluorescence method.