HPAEC-PED

Borromei, C., Careri, M., Cavazza, A., Corradini, C., Elviri, L., Mangia, A. and Merusi, C. (2009) Evaluation of fructooligosaccharides and inulins as potentially health benefiting food ingredients by HPAEC-PED and MALDI-TOF MS, Int. J. Anal Chem., doi 10.1155/2009/530639. 

Once released, sialic acids may be analyzed in its underivatized form by HPAEC-PED or derivatized with DMB (l,2-diamino-4,5-methylenedioxybenzene) ° or OPD(0-phenylene diamine) followed by reversed-phase chromatography with fluorescence detection. If detection and quantification of O-acetylated forms are required HPAEC is not recommended as deacetylation will occur in alkaline conditions. In our experience, the DMB method is highly sensitive, robust, and the chromatographic conditions are amenable to online MS analysis. 

GC, for their direct detection of carbohydrates, with high sensitivity and specificity and without the need for sample derivatization. Specifically, the technique of high-performance anion-exchange chromatography with pulsed electrochemical detection (HPAEC-PED) is fast becoming the technique of choice for this. HPLC techniques with R1 detection are greatly limited in their ability to analyze complex carbohydrates in foods. 

HPAEC analyses were carried out to determine the oligomeric products released from various pectic substrates after depolymerization by the PL isoenzymes. Action pattern analyses for the concerted action of PL isoenzymes utilized 68% esterified pectin as substrate. One-ml reaction mixtures in a buffer system as detailed in section 2.2. comprising 0.5% (w/v) substrate and 5 U of enzyme were incubated for 30 s to 18 h, and then thermoinactivated. Samples of 750 pi were applied to a Carbopac PA-1 (Dionex) column before the carbohydrates were eluted over a period of 70 min using a gradient of 0.2 M KOH, 0.05 M K-acetate to 0.2 M KOH, 0.7 M K-acetate. Detection employed a Pulsed Electrochemical Detector (PED, Dionex) in the integrated amperometry mode according to the manufacturer s recommendations. 

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