A rapid and facile preparation of APTS-labeled N-glycans by combination of ion pair-assisted extraction and HILIC-SPE for routine glycan analysis
A rapid and facile preparation of APTS-labeled N-glycans by combination of ion pair-assisted extraction and HILIC-SPE for routine glycan analysis
复制标题
通过离子对辅助萃取和 HILIC-SPE 相结合，快速、简便地制备 APTS 标记的 N-聚糖，用于常规聚糖分析
DOI：
10.1016/j.jpba.2020.113875
复制
发表时间：
2021
影响因子：
3.4
通讯作者：
Suzuki Shigeo
中科院分区：
文献类型：
--
作者：
Kinoshita Mitsuhiro;Nakatani Yumi;Yamada Keita;Yamamoto Sachio;Suzuki Shigeo
糖分析技术广泛应用于各种科学研究，包括基础糖生物学制药和生物标志物研究。尽管一些创新的分析技术已经被开发出来，但基于电泳分离的定量聚糖分析经常受到缺乏成本效益和简便的样品制备方法的阻碍。在这里，我们开发了一种快速简便的样品制备工作流程，用于具有成本效益的聚糖分析，并演示了其与全自动微芯片电泳（ME）的使用。采用离子对辅助萃取（IPAE）和亲水性相互作用色谱-固相萃取（HILIC-SPE）相结合的方法纯化8-氨基芘-1,3,6-三磺酸酯（APTS）标记的多糖。与常用的样品制备方法相比，IPAE/HILIC-SPE方法在纯化步骤中对n -聚糖的非特异性损失和不良降解最小。此外，我们的方法只需要10 min，整个工作流程，包括聚糖释放、标记和浓缩过程在4 h内完成。虽然目前的系统需要改进以分析更复杂的混合物，但基于me的apts标记n -聚糖分离提供了一个完全自动化的操作，包括调节，样品装载，分离，并且可以在并行过程中以120 s的样品对样品吞吐量进行分析。目前的工作流程易于实施，不需要昂贵的试剂和仪器，可能对跨学科的糖科学家有用。
Glycoanalytical technology is required for a wide variety of scientific research, including basic glycobiological pharmaceutical, and biomarker research. Although several innovative analytical techniques have been developed for these purposes, quantitative glycan analysis based on electrophoretic separation, has often been impeded by the lack of cost-effective and facile sample preparation approaches. Here, we developed a rapid and facile sample preparation workflow for cost-effective glycan analysis and demonstrated its use with fully automated microchip electrophoresis (ME). Purification of 8-aminopyrene-1,3,6-trisulfonate (APTS)-labeled glycans was based on the combination of ion-pair assisted extraction (IPAE) with hydrophilic interaction chromatography-solid phase extraction (HILIC-SPE). Compared to commonly used sample preparation methods, the IPAE/HILIC-SPE method undergoes minimal nonspecific loss and undesirable degradation ofN-glycans during the purification step. Furthermore, our method required only 10 min, and the entire workflow, including glycan release, labeling, and concentration processes was completed within 4 h. Although the present system should be improved to enable analysis of more complex mixtures, ME-based separation of APTS-labeledN-glycans offers a fully automated operation including conditioning, sample loading, separation, and can be analyzed with a sample-to-sample throughput of 120 s in parallel processes. The present workflow is easy to implement, does not require expensive reagents and instruments and may be useful for glycoscientists across disciplines.