Capillary electrophoresis of heparin and other glycosaminoglycans using a polyamine running electrolyte

This study involves the use of polyamines as potential resolving agents for the capillary electrophoresis (CE) of glycosaminoglycans (GAGs), specifically heparin, dermatan sulfate, chondroitin sulfate, over-sulfated chondroitin sulfate (OSCS), and hyaluronan. All of the compounds can be separated from each other with the exception of chondroitin sulfate and hyaluronan. Using optimization software, the final run conditions are found to be 200 mM ethylenediamine and 45.5 mM phosphate as the electrolyte with −14 V applied across a 50 μm ID × 24.5 cm fused silica capillary at 15 °C. The ion migration order, with OSCS as the last instead of the first peak, is in contrast to previous reports using either a high molarity TRIS or lithium phosphate run buffer with narrower bore capillaries. Total analysis time is 12. 5 min and the relative standard deviation of the heparin migration time is about 2.5% (n = 5). The interaction mechanism between selected polyamines and heparin is explored using conductivity measurements in addition to CE experiments to show that an ion-pairing mechanism is likely.

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► Ethylenediamine is likely acting as an ion-pairing agent.
► Oversulfated chondroitin sulfate is last peak instead of first peak.
► There is about a factor of five improved detectability with a 12.5 min analysis time.
► Use of a 50 μm ID capillary is possible.