Abstract
Glycosaminoglycan samples are usually polydisperse, consisting of molecules with differing length and differing sequence. Methods for measuring the molecular weight of heparin have been developed to assure the quality and consistency of heparin products for medicinal use, and these methods can be applied in other laboratory contexts. In the method described here, high-performance gel permeation chromatography is calibrated using appropriate heparin molecular weight markers or a single broad standard calibrant, and used to characterize the molecular weight distribution of polydisperse samples or the peak molecular weight of monodisperse, or approximately monodisperse, heparin fractions. The same technology can be adapted for use with other glycosaminoglycans.
This is a preview of subscription content, log in via an institution to check access.
Access this chapter
Tax calculation will be finalised at checkout
Purchases are for personal use only
References
Al Dieri R, Wagenvoord R, van Dedem GW, Beguin S, Hemker HC (2003) The inhibition of blood coagulation by heparins of different molecular weight is caused by a common functional motif—the C-domain. J Thromb Haemost 1:907–914
Kailemia MJ, Li L, Xu Y, Liu J, Linhardt RJ, Amster IJ (2013) Structurally informative tandem mass spectrometry of highly sulfated natural and chemoenzymatically synthesized heparin and heparan sulfate glycosaminoglycans. Mol Cell Proteomics 12:979–990
Volpi N, Maccari F, Suwan J, Linhardt RJ (2012) Electrophoresis for the analysis of heparin purity and quality. Electrophoresis 33:1531–1537
Sommers CD, Ye H, Kolinski RE, Nasr M, Buhse LF, Al-Hakim A, Keire DA (2011) Characterization of currently marketed heparin products: analysis of molecular weight and heparinase-I digest patterns. Anal Bioanal Chem 401:2445–2454
Mulloy B, Gee C, Wheeler SF, Wait R, Gray E, Barrowcliffe TW (1997) Molecular weight measurements of low molecular weight heparins by gel permeation chromatography. Thromb Haemost 77:668–674
Mulloy B, Heath A, Shriver Z, Jameison F, Al-Hakim A, Morris TS, Szajek A (2014) Development of a compendial method for the chromatographic determination of molecular weight distributions for unfractionated heparin. Anal Bioanal Chem 460:4815–4823
Beirne J, Truchan H, Rao L (2011) Development and qualification of a size exclusion chromatography coupled with multiangle light scattering method for molecular weight determination of unfractionated heparin. Anal Bioanal Chem 399:717–725
Bertini S, Bisio A, Torri G, Bensi D, Terbojevich M (2005) Molecular weight determination of heparin and dermatan sulfate by size exclusion chromatography with a triple detector array. Biomacromolecules 6:168–173
Mulloy B (2002) Gel permeation chromatography of heparin. In: Volpi N (ed) Analytical techniques to evaluate the structure and function of natural polysaccharides, glycosaminoglycans. Research Signpost, Trivandrum
Knobloch JE, Shaklee PN (1997) Absolute molecular weight distribution of low-molecular-weight heparins by size-exclusion chromatography with multiangle laser light scattering detection. Anal Biochem 245:231–241
Huckerby TN, Sanderson PN, Nieduszynski IA (1986) N.M.R. studies of oligosaccharides obtained by degradation of bovine lung heparin with nitrous acid. Carbohydr Res 154:15–27
Khan S, Gor J, Mulloy B, Perkins SJ (2010) Semi-rigid solution structures of heparin by constrained X-ray scattering modelling: new insight into heparin-protein complexes. J Mol Biol 395:504–521
Khan S, Fung KW, Rodriguez E, Patel R, Gor J, Mulloy B, Perkins SJ (2013) The solution structure of heparan sulfate differs from that of heparin: implications for function. J Biol Chem 288:27737–27751
Murphy KJ, Merry CL, Lyon M, Thompson JE, Roberts IS, Gallagher JT (2004) A new model for the domain structure of heparan sulfate based on the novel specificity of K5 lyase. J Biol Chem 279:27239–27245
Hasan J, Shnyder SD, Clamp AR, McGown AT, Bicknell R, Presta M, Bibby M, Double J, Craig S, Leeming D, Stevenson K, Gallagher JT, Jayson GC (2005) Heparin octasaccharides inhibit angiogenesis in vivo. Clin Cancer Res 11:8172–8179
Lauder RM, Huckerby TN, Nieduszynski IA, Sadler IH (2011) Characterisation of oligosaccharides from the chondroitin/dermatan sulphates: (1)H and (13)C NMR studies of oligosaccharides generated by nitrous acid depolymerisation. Carbohydr Res 346:2222–2227
Toida T, Sato K, Sakamoto N, Sakai S, Hosoyama S, Linhardt RJ (2009) Solvolytic depolymerization of chondroitin and dermatan sulfates. Carbohydr Res 344:888–893
Pomin VH, Park Y, Huang R, Heiss C, Sharp JS, Azadi P, Prestegard JH (2012) Exploiting enzyme specificities in digestions of chondroitin sulfates A and C: production of well-defined hexasaccharides. Glycobiology 22:826–838
Volpi N, Bolognani L (1993) Glycosaminoglycans and proteins: different behaviours in high-performance size-exclusion chromatography. J Chromatogr 630:390–396
Author information
Authors and Affiliations
Corresponding author
Editor information
Editors and Affiliations
Rights and permissions
Copyright information
© 2015 Springer Science+Business Media New York
About this protocol
Cite this protocol
Mulloy, B., Hogwood, J. (2015). Chromatographic Molecular Weight Measurements for Heparin, Its Fragments and Fractions, and Other Glycosaminoglycans. In: Balagurunathan, K., Nakato, H., Desai, U. (eds) Glycosaminoglycans. Methods in Molecular Biology, vol 1229. Humana Press, New York, NY. https://doi.org/10.1007/978-1-4939-1714-3_11
Download citation
DOI: https://doi.org/10.1007/978-1-4939-1714-3_11
Published:
Publisher Name: Humana Press, New York, NY
Print ISBN: 978-1-4939-1713-6
Online ISBN: 978-1-4939-1714-3
eBook Packages: Springer Protocols