Dataset Information

Bottom-up low molecular weight heparin analysis using liquid chromatography-Fourier transform mass spectrometry for extensive characterization.

ABSTRACT: Low molecular weight heparins (LMWHs) are heterogeneous, polydisperse, and highly negatively charged mixtures of glycosaminoglycan chains prescribed as anticoagulants. The detailed characterization of LMWH is important for the drug quality assurance and for new drug research and development. In this study, online hydrophilic interaction chromatography (HILIC) Fourier transform mass spectrometry (FTMS) was applied to analyze the oligosaccharide fragments of LMWHs generated by heparin lyase II digestion. More than 40 oligosaccharide fragments of LMWH were quantified and used to compare LMWHs prepared by three different manufacturers. The quantified fragment structures included unsaturated disaccharides/oligosaccharides arising from the prominent repeating units of these LMWHs, 3-O-sulfo containing tetrasaccharides arising from their antithrombin III binding sites, 1,6-anhydro ring-containing oligosaccharides formed during their manufacture, saturated uronic acid oligosaccharides coming from some chain nonreducing ends, and oxidized linkage region oligosaccharides coming from some chain reducing ends. This bottom-up approach provides rich detailed structural analysis and quantitative information with high accuracy and reproducibility. When combined with the top-down approach, HILIC LC-FTMS based analysis should be suitable for the advanced quality control and quality assurance in LMWH production.

SUBMITTER: Li G

PROVIDER: S-EPMC4082394 | biostudies-other | 2014 Jul

REPOSITORIES: biostudies-other

ACCESS DATA

Publications

Bottom-up low molecular weight heparin analysis using liquid chromatography-Fourier transform mass spectrometry for extensive characterization.

Li Guoyun G Steppich Julia J Wang Zhenyu Z Sun Yi Y Xue Changhu C Linhardt Robert J RJ Li Lingyun L

Analytical chemistry 20140617 13

Low molecular weight heparins (LMWHs) are heterogeneous, polydisperse, and highly negatively charged mixtures of glycosaminoglycan chains prescribed as anticoagulants. The detailed characterization of LMWH is important for the drug quality assurance and for new drug research and development. In this study, online hydrophilic interaction chromatography (HILIC) Fourier transform mass spectrometry (FTMS) was applied to analyze the oligosaccharide fragments of LMWHs generated by heparin lyase II dig ...[more]

PMID: 24905078

Similar Datasets

Project description:Calmodulin (CaM) is a highly conserved, ubiquitous, calcium-binding protein; it binds to and regulates many different protein targets, thereby functioning as a calcium sensor and signal transducer. CaM contains 9 methionine (Met), 1 histidine (His), 17 aspartic acid (Asp), and 23 glutamine acid (Glu) residues, all of which can potentially react with platinum compounds; thus, one-third of the CaM sequence is a possible binding target of platinum anticancer drugs, which represents a major challenge for identification of specific platinum modification sites. Here, top-down electron capture dissociation (ECD) was used to elucidate the transition metal-platinum(II) modification sites. By using a combination of top-down and bottom-up mass spectrometric (MS) approaches, 10 specific binding sites for mononuclear complexes, cisplatin and [Pt(dien)Cl]Cl, and dinuclear complex [{cis-PtCl(2)(NH(3))}(2)(?-NH(2)(CH(2))(4)NH(2))] on CaM were identified. High resolution MS of cisplatin-modified CaM revealed that cisplatin mainly targets Met residues in solution at low molar ratios of cisplatin-CaM (2:1), by cross-linking Met residues. At a high molar ratio of cisplatin:CaM (8:1), up to 10 platinum(II) bind to Met, Asp, and Glu residues. [{cis-PtCl(2)(NH(3))}(2)(?-NH(2)(CH(2))(4)NH(2))] forms mononuclear adducts with CaM. The alkanediamine linker between the two platinum centers dissociates due to a trans-labilization effect. [Pt(dien)Cl]Cl forms {Pt(dien)}(2+) adducts with CaM, and the preferential binding sites were identified as Met51, Met71, Met72, His107, Met109, Met124, Met144, Met145, Glu45 or Glu47, and Asp122 or Glu123. The binding of these complexes to CaM, particularly when binding involves loss of all four original ligands, is largely irreversible which could result in their failure to reach the target DNA or be responsible for unwanted side-effects during chemotherapy. Additionally, the cross-linking of cisplatin to CaM might lead to the loss of the biological function of CaM or CaM-Ca(2+) due to limiting the flexibility of the CaM or CaM-Ca(2+) complex to recognize target proteins or blocking the binding region of target proteins to CaM.

Dataset Information

Bottom-up low molecular weight heparin analysis using liquid chromatography-Fourier transform mass spectrometry for extensive characterization.

Publications

Bottom-up low molecular weight heparin analysis using liquid chromatography-Fourier transform mass spectrometry for extensive characterization.

Similar Datasets

OmicsDI is part of the ELIXIR infrastructure

Tweets