Title: HPAEC Coupled to an Evaporative Light Scattering Detector for Separation, Detection and Quantification of Galacturonic Acid Oligomers Authors
|Williams, Martin A. - MASSEY UNIV, NEW ZEALAND|
|Gorhmann, Karel - RETIRED USDA|
Submitted to: International Ion Chromatography Symposium
Publication Type: Abstract Only
Publication Acceptance Date: July 20, 2006
Publication Date: N/A
Technical Abstract: A model homogalacturonan (HG); composed of galacturonic acid (94-97 %) and galactose (3-6 %), having a 94 % degree of esterification (DE) was used to produce a demethylated HG series by reacting it with the salt-independent pectin methylesterase (PME) from citrus fruit at pH 4.5 and 7.5. HGs with a predicted DM (degree of methylation) of 90, 80, 70, and 60 % were produced. Structural properties of the demethylated HG and PME mode of action were probed by performing a limited digest with an endo-polygalacturonase (EPG) from Aspergillus aculeatus. Liberated demethylated blocks (DB) of galacturonic acid (GA) were separated and quantified by high performance anion exchange chromatography (HPAEC) coupled to an evaporative light scattering detector (ELSD). With demethylation at pH 7.5 the longest DB size increased dramatically from 19 at 80 % DE to 47 at 70 % DE while the average DB size increased from 6 to 11 and the average number of DBs per molecule decreased from 6 to 4. A similar change was observed in the pH 4.5 demethylated HG, only at the transition from 70 % to 60 % DE. The presence of a continuum of DB sizes at both pHs suggests a single chain mode of action for the salt-independent PME.