Development of a reversed-phase high-performance liquid chromatography method for the analysis of components from a closed-loop insulin delivery system.

ABSTRACT

A reversed-phase HPLC method has been developed which enables separation of the three components of a closed-loop insulin delivery system, namely concanavalin A methacrylamide (Con A-MA), dextran methacrylate (Dex-MA) and bovine insulin. The analysis of Con A-MA represents a significant challenge due to the formation of multiple conformations on contact with the chromatographic surface and the mobile phase. The extent of conformational change is shown to be dependent on a number of parameters column temperature, mobile phase pH, contact time with the chromatographic surface, salt type and concentration and the organic modifier. By manipulation of these variables, protein denaturation can be minimised and recovery improved.