According to Nagel et al. (2014), stated that carbohydrates analysis is a significant aspect in the food industry since they provide the primary source of energy. Carbohydrates analysis is as well accompanied by nutritional benefits such as the composition of dietary fibers and other unobtainable starches that resist assimilation. Specifically, in the food industry, carbohydrates are used as coadjuvants so as to get physicochemical and chosen sensorial features for some products. Nagel et al. (2014), adds by stating they are mainly taken as determinants of value and authenticity control.
Numerous analytic methods have been projected for carbohydrates analysis. Among them is the High-performance anion-exchange chromatography (HPAEC) coupled with pulsed amperometric detection (PED). Rothenhöfer et al. (2015), claims that it has the powerful technique in carbohydrates separation since it separates all groups of alditols, amino sugars, and both oligo and polysaccharides. HPAEC-PAD is recently being used to several unchanging research applications and monitoring. Additionally, this method has had major influences on the analysis of oligo and polysaccharides. However, Rothenhöfer et al. (2015) suggest that the compatibility of gradient elution with electrochemical detection tied with the greater performance of anion-exchange stationary stages lets compounds of simple sugars, oligo- and polysaccharides to be separated with appropriate resolution in a single phase.
Nagel et al. (2014), argues that HPAEC technique is not necessarily linked with the analysis of neutral carbohydrates. Conversely, several carbohydrates contain weak acids with pKa values between 12 and 14, and subsequently, at elevated pH values, their hydroxyl groups are partly or entirely changed into oxyanions, allowing this group of mixtures to be selectively discharged as anions by high-performance anion-exchange chromatography in a single phase. The favorable alkaline environment allows carbohydrate to be separated by quaternary-ammonium-bonded pellicular anion-exchange columns, whereby the hierarchy of accumulation is associated with reducing pKa value.
Anion-exchange chromatography on large pH-resistant polymeric-based strong anion-exchange columns particularly is structured for carbohydrate analysis since they allow selective discharge of carbohydrates (Nagel et al., 2014). Monosaccharides exhibit several theoretically ionisable hydroxyl classes having, taking glucose as a reference, and the following order of acidity: 1-OH > 2-OH ≥ 6-OH > 3-OH > 4-O. Aldoses indicates higher retention than their reduced alditol forms. Research carried out by (Rothenhöfer et al., 2015), on the retention behavior of 93 alditols and carbohydrates on HPAEC has validated that capacity elements for monosaccharaides are lowest for the alditols and are higher and nearly related for analogous aldoses and ketoses. There is a common inclination, witnessed to be the strongest with alditols, for capacity elements to upsurge with increasing numbers of carbon atoms. The best conventional columns used in HPAEC-PED, particularly structured for carbohydrate anion-exchange chromatography are produced by Dionex (Rothenhöfer et al., 2015).
CarboPac PA1 and CarboPac PA10 columns are designed to separate monosaccharide that has a greater selectivity for mono- and disaccharide separations. These sugars are separated using isocratically technique using sodium hydroxide solutions. However, as (), demonstrates, substances such as glucose and maltose which are closely related in structure and mode of reaction will differ only in the axial-equatorial configuration of their hydroxyl groups. To achieve greater eluent performance, underlying conditions must be provided which is comparable to pKa values of sugar molecules namely pH 12.
HPAEC-PAD is a highly selective technique and precise for carbohydrates because (1) pulsed amperometric detects only those substances that are attached to functional groups that are oxidizable at the detection voltage employed and (2) neutral sample constituents in the matrix elute in. Therefore when they are oxidized, they do not inhibit the analysis of the carbohydrate elements of interest.
Nagel, A., Sirisakulwat, S., Carle, R., & Neidhart, S. (2014). An acetate–hydroxide gradient for the quantitation of the neutral sugar and uronic acid profile of pectins by HPAEC-PAD without postcolumn pH adjustment. Journal of agricultural and food chemistry, 62(9), 2037-2048.
Rothenhöfer, M., Grundmann, M., Bernhardt, G., Matysik, F. M., & Buschauer, A. (2015). High performance anion exchange chromatography with pulsed amperometric detection (HPAEC-PAD) for the sensitive determination of hyaluronan oligosaccharides. Journal of Chromatography B, 988, 106-115.