N-linked glycosylation is one of the most important protein post-translational modifications. Despite the importance of N-glycans, the structural determination of N-glycan isomers remains challenging. Here we develop a mass spectrometry method, logically derived sequence tandem mass spectrometry (LODES/MSⁿ), to determine the structures of N-glycan isomers that cannot be determined using conventional mass spectrometry. In LODES/MSⁿ, the sequences of successive collision-induced dissociation are derived from carbohydrate dissociation mechanisms and apply to N-glycans in an ion trap for structural determination. We validate LODES/MSⁿ using synthesized N-glycans and subsequently applied this method to N-glycans extracted from soybean, ovalbumin, and IgY. Our method does not require permethylation, reduction, and labeling of N-glycans, or the mass spectrum databases of oligosaccharides and N-glycan standards. Moreover, it can be applied to all types of N-glycans (high-mannose, hybrid, and complex), as well as the N-glycans degraded from larger N-glycans by any enzyme or acid hydrolysis.

N-连接糖基化是最重要的蛋白质翻译后修饰之一。尽管 N-聚糖很重要，但 N-聚糖异构体的结构测定仍然具有挑战性。在这里，我们开发了一种质谱法，即逻辑衍生序列串联质谱法 (LODES/MS ⁿ )，以确定使用传统质谱法无法确定的N-聚糖异构体的结构。在 LODES/MS ⁿ中，连续碰撞诱导解离的序列源自碳水化合物解离机制，并应用于离子阱中的N-聚糖以进行结构测定。我们使用合成验证 LODES/MS ⁿN-聚糖，随后将该方法应用于从大豆、卵清蛋白和 IgY 中提取的N-聚糖。我们的方法不需要 N-聚糖的全甲基化、还原和标记，也不需要寡糖和N-聚糖标准品的质谱数据库。此外，它可以应用于所有类型的N-聚糖（高甘露糖、杂化和复合物），以及通过任何酶或酸水解从较大的N-聚糖降解的N-聚糖。