The use of high-performance size exclusion chromatography (HPSEC) as a molecular weight screening technique for polygalacturonic acid for use in pharmaceutical applications

Abstract

Polygalacturonic acid is a linear carbohydrate polymer of monomeric galacturonic acid. It is commercially available as apple and citrus pectins comprised of a mixture of partially methoxylated and/or amidated polygalacturonic acids with molecular weights ranging from 25 000 to >100 000 Da. Pectin can be chemically or enzymatically hydrolyzed to yield polygalacturonic acid fractions of diverse average molecular weight ranges and polydispersities for a variety of uses. Pectin and polygalacturonic acid are used extensively as gelling agents and stabilizers by the food industry, and have applications as therapeutic, and diagnostic pharmaceutical agents such as the magnetic resonance imaging agent LumenHance®. A simple high-performance size exclusion chromatography (HPSEC) method, employing commonly available non-specialized HPLC instrumentation, is described for use as a rapid molecular weight screening technique to determine the average molecular weight range and polydispersity of polygalacturonic acid intended for use in pharmaceutical formulations. A TosoHaas G3000PWXL HPLC column, 50 mM phosphate buffer (pH≈6.9) mobile phase, and refractive index detection were used. A molecular weight calibration curve was linear for polysaccharide standards of 180–100 000 Da with a coefficient of correlation of 0.999. The method was employed to screen commercially available polygalacturonic acid raw materials for average molecular weight data ( M n, M w, and M p) and polydispersity ( M w/ M n).