Unravelling Structural Information from Complex Mixtures Utilizing Correlation Spectroscopy Applied to HSQC Spectra

The first use of <i>statistical</i> correlation spectroscopy to extract chemical information from 2D-HSQC spectra, termed HSQC correlation spectroscopy (<b>HSQCcos</b>), is reported. <b>HSQCcos</b> is illustrated using heparin, a heterogeneous polysaccharide, whose diverse composition causes signals in HSQC spectra to disperse. <b>HSQCcos</b> has been used to probe the chain modifications that cause this effect and reveals hitherto unreported structural details. An interesting finding was that the signal for position 2 of trisulfated glucosamine [N-, 3-O-, and 6-O-sulfated] (<b>A</b>*) is bifurcated, owing to the presence of <b>A</b>* residues in both the “normal” antithrombin binding site and also at the nonreducing end of the molecule, which is reported in intact heparin for the first time. The method was also applied to investigating the environment around other rare sequences/disaccharides, suggesting that the disaccharide; 2-O-sulfated iduronic acid linked to 6-O-sulfated N-glucosamine, which contains a free amine at position 2, is adjacent to the heparin linkage region. <b>HSQCcos</b> can extract chemically related signals from information-rich spectra obtained from complex mixtures such as heparin