Isoamyl nitrite depolymerized heparin as a universal calibrator for heparins and low molecular weight heparins

Because the subcutaneous absorption of heparin depends on its molecular weight, high performance size exclusion chromatography methods, among others, have been developed for its determination. Because heparin consists of polymer chains of varying lengths, a large number of calibrators, 19 in our work, are required for these molecular weight analyses. Because the preparation of that many calibrants is both time-consuming and expensive, other methods requiring fewer calibrants are being sought. In pursuit of these aims, a heparinase-degraded heparin has been accepted as the first international reference preparation low molecular weight heparin for molecular weight calibration. We have previously proposed another calibrant, the Heparin Mass Calibrator. In this article, we introduce another calibrant, called the Heparin Molecular Mass Calibrant. It offers the advantage of a much wider availability because it is a commercial product. In addition to having a superior refractive index chromatography run profile, when used in combination with two Narrow Range Calibrators with molecular weights of 34.4 and 18.4 kDa, this new calibrant covers both the higher and the lower molecular weight ranges. Besides, ultraviolet detection is not needed for molecular weight calculations. Therefore, it is proposed that its feasibility as a universal reference calibrator for both heparins and low molecular weight heparins be considered.