EFFECTS OF PUSHING AGENTS ON THE SEPARATION AND DETECTION OF DEBRANCHED AMYLOPECTIN BY HIGH-PERFORMANCE ANION-EXCHANGE CHROMATOGRAPHY WITH PULSED AMPEROMETRIC DETECTION

The effects of acetate and nitrate as pushing agents on the high-perfo rmance anion-exchange chromatography with pulsed amperometric detector (HPAEC-PAD) for the separation and detection of debranched amylopecti n were evaluated. Our experimental data indicated that nitrate could d ifferentiate branched from linear isomers, whereas acetate could not. Therefore, the nitrate system had better separating power than the ace tate system provided that the chromatograms under both gradient system s were completed within 100 min. Nitrate was more compatible with the detector than acetate; thus, a higher detector response resulted. Unde r our experimental conditions, with a signal-to-noise ratio of two, th e number of DP (degree of polymerization) at various concentrations th at was resolved by the nitrate system (62 at 0.5 mg/mL, 65 at 1.0 mg/m L, and 66 at 1.5 mg/mL) was larger than those resolved by the acetate system (56 at 0.5 mg/mL, 58 at 1.0 mg/mL, and 58 at 1.5 mg/mL). The ch romatograms obtained from the nitrate gradient system were more reprod ucible than those from the acetate gradient system in terms of the con sistencies of retention time and the peak area of the chromatographic peak. To various concentrations of the component with the same DP, the detector responses were more consistent with nitrate as a pushing age nt compared with acetate. Neither pushing agent, however, resulted in a quantitative response for homologous debranched amylopectin.