TY - JOUR
T1 - Sequential Enrichment of Sulfated Glycans by Strong Anion-Exchange Chromatography Prior to Mass Spectrometric Measurements
AU - Lei, Ming
AU - Novotny, Milos V.
AU - Mechref, Yehia
N1 - Funding Information:
The authors acknowledge primary support for this work by grant no. GM24349 from the National Institute of General Medical Sciences , U.S. Department of Health and Human Services. Additional support was provided by NIH/NCRR for the National Center for Glycomics and Glycoproteomics (NCGG) at Indiana University (grant no. RR018942 ).
PY - 2010/3
Y1 - 2010/3
N2 - Structural characterization of sulfated glycans through mass spectrometry (MS) has been often limited by their low abundance in biological materials and inefficient ionization in the positive-ion mode. Here, we describe a microscale method for sequentially enriching sulfated glycans according to their degree of sulfation. This method is based on modifying the binding ability of strong anion-exchange material through the use of different sodium acetate concentrations, thus enabling fairly selective binding and a subsequent elution of different glycans according to their degree of sulfation. Before this enrichment, the negative charge on the sialic acid, which is commonly associated with such glycans, was eliminated through permethylation that is used to enhance the positive-ion mode matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-MS) signal for all glycans. This enrichment approach minimizes competitive ionization between sulfated and neutral glycans, as well as that between sulfated species with a different degree of sulfation. The described method was initially optimized using sulfated oligosaccharide standards, while its potential has been verified for the sulfated N-glycans originated from the bovine thyroid-stimulating hormone (bTSH), a glycoprotein possessing mono- and disulfated N-glycans. This enhancement of the MALDI-MS signal facilitates analysis of some otherwise undetected components.
AB - Structural characterization of sulfated glycans through mass spectrometry (MS) has been often limited by their low abundance in biological materials and inefficient ionization in the positive-ion mode. Here, we describe a microscale method for sequentially enriching sulfated glycans according to their degree of sulfation. This method is based on modifying the binding ability of strong anion-exchange material through the use of different sodium acetate concentrations, thus enabling fairly selective binding and a subsequent elution of different glycans according to their degree of sulfation. Before this enrichment, the negative charge on the sialic acid, which is commonly associated with such glycans, was eliminated through permethylation that is used to enhance the positive-ion mode matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-MS) signal for all glycans. This enrichment approach minimizes competitive ionization between sulfated and neutral glycans, as well as that between sulfated species with a different degree of sulfation. The described method was initially optimized using sulfated oligosaccharide standards, while its potential has been verified for the sulfated N-glycans originated from the bovine thyroid-stimulating hormone (bTSH), a glycoprotein possessing mono- and disulfated N-glycans. This enhancement of the MALDI-MS signal facilitates analysis of some otherwise undetected components.
UR - http://www.scopus.com/inward/record.url?scp=76749097658&partnerID=8YFLogxK
U2 - 10.1016/j.jasms.2009.09.017
DO - 10.1016/j.jasms.2009.09.017
M3 - Article
C2 - 20022260
AN - SCOPUS:76749097658
VL - 21
SP - 348
EP - 357
JO - Journal of the American Society for Mass Spectrometry
JF - Journal of the American Society for Mass Spectrometry
SN - 1044-0305
IS - 3
ER -