应用制备高效液相色谱同时制备6种大豆异黄酮单体的方法研究

Simultaneous Preparation of Six Soybean Isoflavones Monomers by Preparative HPLC

经大孔吸附树脂纯化大豆异黄酮粗提物后,以制备型高效液相色谱法(PHPLC)分离得到高纯度的大豆异黄酮单体。以SHIM-pack PRC-ODS(20 mm×250 mm,5μm)制备柱,考察了流动相组成及流速、进样量对分离度的影响,确定了最佳色谱条件为乙腈-水流动相梯度洗脱,进样量800μL,流速10 mL/min,在120 min内实现了6种异黄酮单体的基线分离及制备。经超高效液相色谱-串联质谱(UPLC-MS/MS)鉴定,6种异黄酮单体依次为大豆苷、黄豆黄苷、染料木苷、大豆素、黄豆黄素、染料木素。6种产品的纯度分别为95.54%、90.14%、100%、100%、96.27%、100%。方法具有简便易行、稳定性好、产品纯度高等特点,适用于大豆异黄酮标准品的制备。

A preparative high performance liquid chromatographic （PHPLC） method was established for the simultaneous preparation of six soybean isoflavones monomers with high purity from soybean crude extracts purified with macroporous adsorption resin.The effects of experimental conditions,including mobile phase composition,flow rate and sample quantity on resolution were investigated on a SHIM-pack PRC-ODS preparative column （20 mm × 250 mm,5 μm）.The optimal conditions were as follows：mobile phase：acetonitrile-water by gradient elution,injection volume：800 μL,flow rate：l0 mL/min.Under the optimal conditions,the baseline separation and preparation of six isoflavones monomers were achieved within 120 min.Six compounds were identified by ultra high performance liquid chromatography-tendem mass spectrometry （UPLC-MS/MS） to be daidzin,glycitin,genistin,daidzein,glycitein and genistein with purities of 95.54％,90.14％,100％,100％,96.27％ and 100％,respectively.With the benefits of simplicity,good stability and high products purity,the developed method show a good application prospect in the production of soybean isoflavones standard materials.