Profiling of cool-season forage arabinoxylans via a validated HPAEC-PAD method

Abstract

Cool-season pasture grasses contain arabinoxylans (AX) as their major cell wall hemicellulosic polysaccharide. AX structural differences may influence enzymatic degradability, but this relationship has not been fully explored in the AX from the vegetative tissues of cool-season forages, primarily because only limited AX structural characterization has been performed in pasture grasses. Structural profiling of forage AX is a necessary foundation for future work assessing enzymatic degradability and may also be useful for assessing forage quality and suitability for ruminant feed. The main objective of this study was to optimize and validate a high-performance anion-exchange chromatography with pulsed amperometric detection (HPAEC-PAD) method for the simultaneous quantification of 10 endoxylanase-released xylooligosaccharides (XOS) and arabinoxylan oligosaccharides (AXOS) in cool-season forage cell wall material. The following analytical parameters were determined or optimized: chromatographic separation and retention time (RT), internal standard suitability, working concentration range (CR), limit of detection (LOD), limit of quantification (LOQ), relative response factor (RRF), and quadratic calibration curves. The developed method was used to profile the AX structure of four cool-season grasses commonly grown in pastures (timothy, Phleum pratense L.; perennial ryegrass, Lolium perenne L.; tall fescue, Schedonorus arundinaceus (Schreb.) Dumort.; and Kentucky bluegrass, Poa pratensis L.). In addition, the cell wall monosaccharide and ester-linked hydroxycinnamic acid contents were determined for each grass. The developed method revealed unique structural aspects of the AX structure of these forage grass samples that complemented the results of the cell wall monosaccharide analysis. For example, xylotriose, representing an unsubstituted portion of the AX polysaccharide backbone, was the most abundantly-released oligosaccharide in all the species. Perennial rye samples tended to have greater amounts of released oligosaccharides compared to the other species. This method is ideally suited to monitor structural changes of AX in forages as a result of plant breeding, pasture management, and fermentation of plant material.

Fructan Catabolism by Rumen Microbiota of Cattle and Sheep

“…Species differences in short-chain fructan catabolism in the present study were also reflected in patterns of appearance and disappearance of bifurcose and 1-nystose, but not in free sugar concentrations. A more frequent sampling interval and employing techniques to improve resolution of glucose, fructose, and sucrose peaks such as the method recently developed by Joyce et al [44] could aid in characterizing fluctuations in these mono and disaccharides during catabolism of grass fructans. It should be noted that in bovine fermentations in the current study, the greatest degradation of both long-and short-chain fructans occurred during the 2-4 h period, with long-chain fructans non-detectable and short-chain fructans almost completely utilized by 4 h. It is therefore possible that the capacity of the bovine rumen microbiota to catabolize grass fructans exceeded the fructans provided by the orchardgrass used as the fermentative substrate in this experiment.…”

Exploration and Improvement of Acid Hydrolysis Conditions for Inulin-Type Fructans Monosaccharide Composition Analysis: Monosaccharide Recovery and By-Product Identification

“…Key variables such as acid hydrolysis duration, temperature, and acid concentration are meticulously adjusted to ensure the optimal recovery, precision, and reproducibility of monosaccharide data [8]. Tailoring hydrolysis conditions to suit the dissociation of polysaccharides with diverse structural characteristics is crucial to achieving accurate results [9,10]. For instance, distinct ketoses (e.g., D-fructose) and aldoses (e.g., D-glucose) may yield identical alditol products upon reduction, owing to the enolization and dehydration processes converting D-fructose to alditol under harsh hydrolytic conditions.…”