叶黄素、β-隐黄质及其异构体检测方法的研究

Separation and Identification of Lutein, β-Cryptoxanthin and Their Isomers

应用C30-HPLC-DAD技术,建立一种同时分离全反式叶黄素、全反式β-隐黄质及其异构体的检测方法。采用APCI-MS和光谱技术对分离出化合物进行定性分析。最佳色谱条件：1.5%乙酸铵水-甲基叔丁基醚（MTBE）-甲醇（5：25：70,V/V/V）为流动相A,1.5%乙酸铵水-甲基叔丁基醚（MTBE）-甲醇（5：85：10,V/V/V）为流动相B,线性梯度洗脱;流速：0.6 m L/min;洗脱时间：24min;进样量：20μL;柱温：25℃。混合物中的全反式叶黄素及其15-顺式、13/13’-顺式、9-顺式、9’-顺式异构体,全反式β-隐黄质及其15-顺式、13-和13’-顺式、9-顺式、9’-顺式异构体均得到较好分离。全反式叶黄素、全反式β-隐黄质分别在2~150 ng和5~250 ng范围内峰面积与进样量呈良好线性关系。该方法能够快速、准确地同时分离全反式叶黄素、全反式β-隐黄质及它们的异构体,可用于果蔬中全反式叶黄素和全反式β-隐黄质及其异构体混合物的定性定量分析。

A method to simultaneously detect all-trans-Lutein, all-trans-β-cryptoxanthin and their stereoisomers was developed using C30-column, high-performance liquid chromatography, diode array（C30-HPLC-DAD） technique. The structures of isolated compounds were elucidated by atmospheric pressure chemical ionization-mass spectrometry（APCI-MS） and ultraviolet/visible（UV/Vis） spectroscopy. The optimal chromatographic condition was as follows： mobile phase A： 1.5% ammonium acetate：water：methyl tert-butyl ether（MTBE）-Me OH（5：25：70, V/V/V）, mobile phase B： 1.5% ammonium acetate：water：MTBE-Me OH（5：85：10, V/V/V）; flow rate： 0.6 m L/min; elution time： 24 min; injection volume： 20 μL; column temperature： 25 ℃. Using this novel method, 15-cis-lutein, 13/13′-cis-lutein, 9-cis-lutein, 9′-cis-lutein, 15-cis-β-cryptoxanthin, 13- and 13′-cis-β-cryptoxanthin, 9-cis-β-cryptoxanthin and 9′-cis-β-cryptoxanthin were successfully separated from the mixture. The peak areas of all-trans lutein and all-trans β-cryptoxanthin showed a good linear relationship with their injection volumes in the range of 2 to 150 ng and 5 to 250 ng, respectively. The proposed method accurately and rapidly separated all-trans lutein, all-trans β-cryptoxanthin, and their cis-isomers simultaneously. This method has potential applications in qualitative and quantitative analysis of all-trans lutein, all-trans β-cryptoxanthin, and their stereoisomers in fruits and vegetables.