AnnoSM: An Automated Annotation Tool for Determining the Substituent Modes on the Parent Skeleton Based on a Characteristic MS/MS Fragment Ion Library

Mass spectrometry (MS) is a powerful technology for the
structural
elucidation of known or unknown small molecules. However, the accuracy
of MS-based structure annotation is still limited due to the presence
of numerous isomers in complex matrices. There are still challenges
in automatically interpreting the fine structure of molecules, such
as the types and positions of substituents (substituent modes, SMs)
in the structure. In this study, we employed flavones, flavonols,
and isoflavones as examples to develop an automated annotation method
for identifying the SMs on the parent molecular skeleton based on
a characteristic MS/MS fragment ion library. Importantly, user-friendly
software AnnoSM was built for the convenience of researchers with
limited computational backgrounds. It achieved 76.87% top-1 accuracy
on the 148 authentic standards. Among them, 22 sets of flavonoid isomers
were successfully differentiated. Moreover, the developed method was
successfully applied to complex matrices. One such example is the
extract of Ginkgo biloba L. (EGB),
in which 331 possible flavonoids with SM candidates were annotated.
Among them, 23 flavonoids were verified by authentic standards. The
correct SMs of 13 flavonoids were ranked first on the candidate list.
In the future, this software can also be extrapolated to other classes
of compounds.

质谱（Mass spectrometry, MS）是一种功能强大的分析技术，可用于解析已知或未知小分子的结构。然而，由于复杂基质中存在大量同分异构体，基于质谱的结构注释准确性仍存在局限。当前在自动解析分子精细结构方面仍面临诸多挑战，例如分子结构中取代基的类型与位置（取代模式，substituent modes, SMs）。本研究以黄酮类、黄酮醇类及异黄酮类化合物为研究对象，基于特征性串联质谱（MS/MS）碎片离子库，开发了一种可自动识别母分子骨架上取代模式的注释方法。尤为重要的是，为方便计算背景有限的研究人员使用，我们开发了一款易用的软件AnnoSM。在148种标准品的测试中，该方法取得了76.87%的Top-1准确率，成功区分了22组黄酮类同分异构体。此外，所开发的方法已成功应用于复杂基质样本，其中一例为银杏叶提取物（Ginkgo biloba L., EGB），从中注释得到331种带有潜在取代模式候选物的黄酮类化合物，其中23种经标准品验证，13种黄酮类化合物的正确取代模式在候选列表中位列首位。未来，该软件还可推广应用至其他类别化合物。