Figure 5.

Analysis of 3-O-sulfated HS using a LC-MS/MS method. The HS polysaccharides from biological sources are mixed with known amounts of ¹³C-labeled 8-mer calibrants. The mixture is subjected to the digestion with heparin lyases. The 3-O-sulfated oligosaccharides, presented as Tetra-1, Tetra-2, and Di3S are liberated along with non-3-O-sulfated disaccharides. The products are then analyzed by LC-MS/MS. Comparing the ratio of peak area ¹³C-labeled Tetra-1 and unlabeled Tetra, one can determine the amount of Tetra-1 from the HS polysaccharides. Red dots denote the location of the ¹³C-atoms. HS, heparan sulfate; LC-MS/MS, liquid chromatography coupled with tandem mass spectrometry.