title: Hydrophilic interaction chromatography-multiple reaction monitoring mass spectrometry method for basic building block analysis of low molecular weight heparins prepared through nitrous acid depolymerization.
abstract: Low molecular weight heparins (LMWHs) are important anticoagulant drugs that are prepared through depolymerization of unfractionated heparin. Based on the types of processing reactions and the structures of the products, LMWHs can be divided into different classifications. Enoxaparin is prepared by benzyl esterification and alkaline depolymerization, while dalteparin and nadroparin are prepared through nitrous acid depolymerization followed by borohydride reduction. Compositional analysis of their basic building blocks is an effective way to provide structural information on heparin and LMWHs. However, most current compositional analysis methods have been limited to heparin and enoxaparin. A sensitive and comprehensive approach is needed for detailed investigation of the structure of LMWHs prepared through nitrous acid depolymerization, especially their characteristic saturated non-reducing end (NRE) and 2,5-anhydro-d-mannitol reducing end (RE). A maltose modified hydrophilic interaction column offers improved separation of complicated mixtures of acidic disaccharides and oligosaccharides. A total of 36 basic building blocks were unambiguously identified by high-resolution tandem mass spectrometry (MS). Multiple reaction monitoring (MRM) MS/MS quantification was developed and validated in the analysis of dalteparin and nadroparin samples. Each group of building blocks revealed different aspects of the properties of LMWHs, such as functional motifs required for anticoagulant activity, the structure of heparin starting materials, cleavage sites in the depolymerization reaction, and undesired structural modifications resulting from side reactions.
authors.affiliation: National Glycoengineering Research Center, Shandong Provincial Key Laboratory of Carbohydrate Chemistry and Glycobiology, State Key Laboratory of Microbial Technology, Shandong University, Jinan, Shandong 250100, China; Department of Chemistry and Chemical Biology, Department of Chemical and Biological Engineering, Department of Biology, Department of Biomedical Engineering, Center for Biotechnology and Interdisciplinary Studies, Rensselaer Polytechnic Institute, Troy, NY 12180, USA.
authors.forename: Xiaojun
authors.initials: X
authors.lastname: Sun
authors.affiliation: Key Laboratory of Separation Science for Analytical Chemistry, Dalian Institute of Chemical Physics, Chinese Academy of Sciences, Dalian, Liaoning 116023, China. Electronic address: guozhimou@dicp.an.cn.
authors.forename: Zhimou
authors.initials: Z
authors.lastname: Guo
authors.affiliation: National Glycoengineering Research Center, Shandong Provincial Key Laboratory of Carbohydrate Chemistry and Glycobiology, State Key Laboratory of Microbial Technology, Shandong University, Jinan, Shandong 250100, China.
authors.forename: Mengqi
authors.initials: M
authors.lastname: Yu
authors.affiliation: Jinan Center for Food and Drug Control, Jinan, Shandong 250102, China.
authors.forename: Chao
authors.initials: C
authors.lastname: Lin
authors.affiliation: National Glycoengineering Research Center, Shandong Provincial Key Laboratory of Carbohydrate Chemistry and Glycobiology, State Key Laboratory of Microbial Technology, Shandong University, Jinan, Shandong 250100, China.
authors.forename: Anran
authors.initials: A
authors.lastname: Sheng
authors.affiliation: Department of Virology, School of Public Health, Shandong University, Jinan, Shandong 250100, China.
authors.forename: Zhiyu
authors.initials: Z
authors.lastname: Wang
authors.affiliation: Department of Chemistry and Chemical Biology, Department of Chemical and Biological Engineering, Department of Biology, Department of Biomedical Engineering, Center for Biotechnology and Interdisciplinary Studies, Rensselaer Polytechnic Institute, Troy, NY 12180, USA.
authors.forename: Robert J
authors.initials: RJ
authors.lastname: Linhardt
authors.affiliation: National Glycoengineering Research Center, Shandong Provincial Key Laboratory of Carbohydrate Chemistry and Glycobiology, State Key Laboratory of Microbial Technology, Shandong University, Jinan, Shandong 250100, China. Electronic address: lianlichi@sdu.edu.cn.
authors.forename: Lianli
authors.initials: L
authors.lastname: Chi
mesh_terms: D002853:Chromatography, Liquid; D006493:Heparin / Q000737:chemistry* / Q000378:metabolism; D019763:Heparin Lyase / Q000378:metabolism; D006495:Heparin, Low-Molecular-Weight / Q000737:chemistry* / Q000302:isolation & purification; D057927:Hydrophobic and Hydrophilic Interactions; D017762:Nadroparin / Q000032:analysis; D009608:Nitrous Acid / Q000737:chemistry*; D053719:Tandem Mass Spectrometry
chemical_list: D006495:Heparin, Low-Molecular-Weight; D017762:Nadroparin; D006493:Heparin; D019763:Heparin Lyase; D009608:Nitrous Acid
keywords: Basic building blocks; Hydrophilic interaction chromatography; LC-MRM-MS/MS; Low molecular weight heparin