Purification of N- and O-glycans and their derivatives from biological samples by the absorbent cotton hydrophilic chromatographic column.
Purification of N- and O-glycans and their derivatives from biological samples by the absorbent cotton hydrophilic chromatographic column.
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DOI：
10.1016/j.chroma.2020.461001
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发表时间：
2020-02
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影响因子：
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通讯作者：
Jianli Han;Qinghui Chen;W. Jin;Meiyi Zou;Yu Lu;Yuxia Liu;Chengjian Wang;Zhongfu Wang;Linjuan Huang
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文献类型：
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作者：
Jianli Han;Qinghui Chen;W. Jin;Meiyi Zou;Yu Lu;Yuxia Liu;Chengjian Wang;Zhongfu Wang;Linjuan Huang
质谱（MS）是对多糖进行定性和定量分析最常用的工具之一。然而，由于生物样品的复杂性和聚糖的低电离效率，这些需要在MS分析之前进行纯化和衍生化。现有的净化策略需要多种方法的结合，操作起来很麻烦。在这里，我们提出了一种新的方法来纯化糖蛋白N/ o聚糖及其衍生物，使用手包装吸水棉亲水性相互作用色谱柱（HILIC）。通过纯化N/ o聚糖及鸡白蛋白、猪胃粘蛋白等标准糖蛋白衍生物，验证了该方法的可靠性和适用性。采用稳定同位素标记法比较不同纯化方法下多糖的回收率。吸收棉HILIC也成功地应用于人血清和胎牛血清糖蛋白n -聚糖的分析。结果表明，多糖的结合能力高（9 mg/g - 1麦芽糖己糖/吸水棉），回收率高（平均回收率为91.7%）。与传统纯化方法相比，该纯化方法具有简单、高效、经济、通用性强、适用范围广等优点，适用于生物样品中多糖及其衍生物在质谱分析前的预处理。
Mass spectrum (MS) is one of the most commonly used tools for qualitative and quantitative analysis of glycans. However, due to the complexity of biological samples and the low ionization efficiency of glycans, these need to be purified and derivatized prior to MS analysis. Existing purification strategies require a combination of multiple methods and are cumbersome to operate. Here, we propose a new method for the purification of glycoprotein N/O-glycans and their derivatives using a hand-packed absorbent cotton hydrophilic interaction chromatography column (HILIC). The method's reliability and applicability were verified by purifying N/O-glycans and the derivatives of standard glycoproteins, such as chicken albumin and porcine stomach mucin. Stable isotope labelling was used to compare the glycans’ recovery following different purification methods. Absorbent cotton HILIC was also successfully applied for the analysis of human serum and fetal bovine serum glycoprotein N-glycans. Finally, testing revealed high binding capacity (9 mg/g−1maltohexaose/absorbent cotton) and good recovery (average recovery was 91.7%) of glycans. Compared with traditional procedures, the proposed purification method offers considerable advantages, such as simplicity, high efficiency, economy, universality, and broad applicability for the pretreatment of glycans and their derivatives in biological samples prior to MS analysis.