Separation, Detection, and Qua.gification of Galacturonic Acid Oligomers with a Degree of Polymerization Greater than 50

Abstract

A high performance anion exchange chromatography (HPAEC)–evaporative light scattering detector (ELSD)‐method was developed to detect, separate, and qua.gify galacturonic acid (GA) oligomers. Following digestion of polygalacturonic acid (PGA) with a monocomponent endo‐polygalacturonase (EPG), more than 70 GA oligomer peaks could be resolved using a convex/linear ammonium formate gradient. Linear calibration curves were produced for 0.015–1.0% mono‐, di‐, and tri‐GA. The mass response for mono‐GA differed from those for di‐ and tri‐GA, as evidenced from the slope of the calibration curve regression lines (1.611 ± 0.0201 for mono‐GA vs. 1.3068 ± 0.0291 and 1.3004 ± 0.0262 for di‐, and tri‐GA, respectively). The degree of polymerization (DP) appeared to affect mass response as the trend line for log‐transformed peak areas of DP 3, 4, 6, and 8 oligomers had a slope of −0.0304 ± 0.0032 (r ² = 0.98). Buffer concentration also affected mass response. ANOVA of peak areas from isocratic elution of trimer and hexamer with 50 mM to 0.8 M ammonium formate indicated mass response was dependent on buffer concentration for each oligomer (P < 0.005), although Duncan's Multiple Range Test described concentration ranges within which mass response was not affected (P < 0.05).

^#K. Grohmann is retired.

Keywords:

• pectin
• oligogalacturonic acid
• ELSD
• evaporative light scattering detection
• endo‐polygalacturonase
• EPG

Acknowledgments

The authors would like to acknowledge expert technical support provided by Steven W. Kauffman, statistical advice provided Dr. Anne Plotto, and the kind gift of purified GA oligomers by Dr. Arland Hotchkiss.

Notes

^#K. Grohmann is retired.