Chromatographic analysis and sequencing approach of heparin oligosaccharides using cetyltrimethylammonium dynamically coated stationary phases

Heparin Dodecasaccharide Containing Two Antithrombin-binding Pentasaccharides

“…The solution was then desalted on Sephadex G-10. The structure was determined using a controlled digestion with heparin lyase I (heparinase I, EC 4.2.2.7) followed by HPLC analysis according to the procedure described previously (13). In the reduced dodecasaccharide, only one site was cleaved by heparinase I, and a mixture of the tetrasaccharide ⌬U-A NAc,6S -GlcUA-A NS,3S,6S and the octasaccharide ⌬U 2S -A NS,6S -I-A NAc,6S -GlcUA-A NS,3S,6S -I 2S -A NS,6S-ol was predominantly obtained as shown in Fig.…”

“…AT-binding oligosaccharides are usually obtained after partial chemical or enzymatic depolymerization of heparin and purification using affinity chromatography on an immobilized Sepharose-AT column (12). In other studies, the AT-binding oligosaccharides were isolated from LMWHs, such as enoxaparin, using a combination of orthogonal separation techniques, including gel permeation chromatography, HPLC, and AT affinity chromatography (13).…”

Pharmacology of Heparin and Related Drugs

“…The use of cetyltrimethylammonium-coated columns has been noted to improve the performance of strong anion exchange chromatography in the separation of heparin-derived oligosaccharides (Mourier and Viskov, 2004). The resulting column was used in a sequencing strategy without the use of MS, merely specific fragmentation and chromatography; minor disaccharides were resolved and identified by NMR.…”

The Interaction of Heparin Tetrasaccharides with Chemokine CCL5 Is Modulated by Sulfation Pattern and pH

“…Anion exchange chromatography on a C 18 stationary phase coated with cetyltrimethylammonium ions was performed by adapting the guidelines presented by Mourier and Viskov (48). A preparative 250 ϫ 21.2-mm, 5-m Prep C 18 column (Phenomenex, Torrance, CA) was coated with cetyltrimethylammonium using 1 mM cetyltrimethylammonium bromide dissolved in 32% methanol at a flow rate of 2.5 ml/min at room temperature overnight.…”