20200520 Khoo KH paper photo 1

Permethylation and Microfractionation of Sulfated Glycans for MS Analysis

Permethylation and Microfractionation of Sulfated Glycans for MS Analysis

BIO-PROTOCOL. 2020 May 20; 10(10):e3617.
doi: 10.21769/BioProtoc.3617


Yu, Shin-Yi; Snovida, Sergei; Khoo, Kay-Hooi


Sulfated glycans are barely detectable in routine mass spectrometry (MS)-based glycomic analysis due to ion suppression by the significantly more abundant neutral glycans in the positive ion mode, and sialylated non-sulfated glycans in the negative ion mode, respectively. Nevertheless, the negative charge imparted by sulfate can be advantageous for selective detection in the negative ion mode if the sialic acids can first be neutralized. This is most conveniently achieved by a concerted sample preparation workflow in which permethylation is followed by solid phase fractionation to isolate the sulfated glycans prior to MS analysis. Importantly, we demonstrated that conventional NaOH/DMSO slurry permethylation method can retain the sulfates. Instead of extracting permethylated glycans into chloroform for sample clean-up, reverse phase C18 cartridge coupled with self-packed amine-tip or mixed mode weak anion exchange cartridge can be utilized to obtain in good yield the non-sulfated, mono-sulfated, and multiply sulfated permethylated glycans in separate fractions for sulfoglycomic analysis.