Journal of Shandong University (Health Sciences) ›› 2021, Vol. 59 ›› Issue (3): 41-47.doi: 10.6040/j.issn.1671-7554.0.2020.1448

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Identification of heparin and heparan sulfate by HILIC-MS/MS

LIU Dongke1, WANG Fengshan1,2   

  1. 1. Key Laboratory of Chemical Biology(Ministry of Education), Institute of Biochemical and Biotechnological Drug, School of Pharmaceutical Sciences, Cheeloo College of Medicine, Shandong University, Jinan 250012, Shandong, China;
    2. Laboratory of Carbohydrate Chemistry and Glycobiology of Shandong Province, National Glycoengineering Research Center, Shandong University, Jinan 250012, Shandong, China
  • Published:2021-04-06

Abstract: Objective To establish a hydrophilic interaction chromatography-mass spectrometry(HILIC-MS/MS)method for simultaneous determination of 8 disaccharides in heparin and heparan sulfate after digestion by heparinase Ⅰ, Ⅱ and Ⅲ, and to provide method and data references for the identification of heparin and heparan sulfate. Methods A PC HILIC column(4.6 mm×250 mm, 5 μm)was used and gradiently eluted with 10 mmol/L ammonium acetate aqueous solution(A)and 10 mmol/L ammonium-90% acetonitrile(B)at the flow rate of 0.9 mL/min, column temperature of 40 ℃ and injection volume of 10 μL. The contents of the 8 disaccharides of heparin and heparan sulfate produced by enzymatic digestion were determined by electrospray ionization source and multi reaction monitoring mode and were compared with each other. Results The linear relationships of the 8 components were good in the range of measured molar concentrations(r2>0.996 0), and the average recovery were 90.34%-99.62%, RSD were 4.7%-6.7%. The results showed that the disaccharides composition and proportion of heparin and heparan sulfate were significantly different. In heparin, ΔUA2S-GlcNS6S(TriS)accounted for about 69.8% on average, while in heparan sulfate, ΔUA-GlcNAc(0S)accounted for about 51.0%. Conclusion The method established in this paper is simple, fast, accurate and reliable with high sensitivity, strong specificity, good stability, and can be used as one of the determination and identification methods for heparin and heparan sulfate.

Key words: Heparin, Heparan sulfate, Hydrophilic interaction chromatography-mass spectrometry, Disaccharides, Identification

CLC Number: 

  • R917
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