Catrin E. Tyl

Isolation and characterization of feruloylated arabinoxylan oligosaccharides from the perennial cereal grain intermediate wheat grass (Thinopyrum intermedium).

In comparison to the annual grain crops dominating current agricultural production, perennial grain species require fewer chemical and energy inputs and improve soil health and erosion control. The possibility for producing sustainable grain harvests from marginal land areas is motivating research initiatives to integrate perennial grains into commercial cropping and food processing systems. In this study, the feruloylated arabinoxylans from intermediate wheat grass (Thinopyrum intermedium, IWG), a promising perennial grain candidate in agronomic screening studies, were investigated. Insoluble fiber isolated from IWG whole grain flour was subjected to either mildly acidic (50 mM TFA, 100 °C, 2 h) or enzymatic (Driselase) hydrolysis. The liberated feruloylated arabinoxylan oligosaccharides were concentrated with Amberlite XAD-2, separated with gel chromatography (Sephadex LH-20, water), and purified with reversed-phase HPLC (C18, water-MeOH gradient). Thirteen feruloylated oligosaccharides were isolated (including eight structures described for the first time) and identified by LC-ESI-MS and NMR. Linkage-type analysis via methylation analysis, as well as the monosaccharide and phenolic acid profiles of the IWG insoluble fiber were also determined. IWG feruloylated arabinoxylans have a relatively simple structure with only short feruloylated side chains, a lower backbone substitution rate than annual rye and wheat varieties, and a moderate phenolic acid content.

Antioxidant activity-guided fractionation of blue wheat (UC66049 Triticum aestivum L.)

Antioxidant activity-guided fractionation based on three in vitro antioxidant assays (Folin-Ciocalteu, TEAC, and leucomethylene blue assays) was used to identify major antioxidants in blue wheat (UC66049 Triticum aestivum L.). After consecutive extractions with solvents of various polarities and multiple chromatographic fractionations, several potent antioxidants were identified by NMR spectroscopy and mass spectrometry. Anthocyanins (delphinidin-3-glucoside, delphinidin-3-rutinoside, cyanidin-3-glucoside, and cyanidin-3-rutinoside), tryptophan, and a novel phenolic trisaccharide (β-D-glucopyranosyl-(1→6)-β-D-glucopyranosyl-(1→6)-(4-hydroxy-3-methoxyphenyl)-β-D-glucopyranoside) were the most active water-extractable constituents. However, anthocyanins were found to be major contributors to the overall blue wheat antioxidant activity only when the extraction steps were performed under acidic conditions. Alkylresorcinols were among the most active antioxidants extractable with 80% ethanol in the TEAC assay. However, this may be due to a color change instead of a bleaching of the ABTS radical. Ferulic acid was found to be the major antioxidant in alkaline cell-wall hydrolysates. The contents of the most active antioxidants were determined.

Characterization of diferuloylated pectic polysaccharides from quinoa (Chenopodium quinoa WILLD.)

In plants belonging to the order of Caryophyllales, pectic neutral side chains can be substituted with ferulic acid. The ability of ferulic acid to form intra- and/or intermolecular polysaccharide cross-links by dimerization was shown by the isolation and characterization of diferulic acid oligosaccharides from monocotyledonous plants. In this study, two diferulic acid oligosaccharides were isolated from the enzymatic hydrolyzate of seeds of the dicotyledonous pseudocereal quinoa by gel permeation chromatography and preparative HPLC and unambiguously identified by LC-MS(2) and 1D/2D NMR spectroscopy. The isolated oligosaccharides are comprised of 5-5- and 8-O-4-diferulic acid linked to the O2-position of the nonreducing residue of two (1→5)-linked arabinobioses. To get insight into the structure and the degree of phenolic acid substitution of the diferuloylated polysaccharides, polymeric sugar composition, glycosidic linkages, and polysaccharide-bound monomeric phenolic acids and diferulic acids were analyzed. This study demonstrates that diferulic acids are involved into intramolecular and/or intermolecular cross-linking of arabinan chains and may have a major impact on cell wall architecture of quinoa and other dicotyledonous plants of the order of Caryophyllales.

Novel arabinan and galactan oligosaccharides from dicotyledonous plants

Arabinans and galactans are neutral pectic side chains and an important part of the cell walls of dicotyledonous plants. To get a detailed insight into their fine structure, various oligosaccharides were isolated from quinoa, potato galactan, and sugar beet pulp after enzymatic treatment. LC-MS2 and one- and two-dimensional NMR spectroscopy were used for unambiguous structural characterization. It was demonstrated that arabinans contain β-(1→3)-linked arabinobiose as a side chain in quinoa seeds, while potato galactan was comprised of β-(1→4)-linked galactopyranoses which are interspersed with α-(1→4)-linked arabinopyranoses. Additionally, an oligosaccharide with two adjacent arabinofuranose units O2-substituted with two ferulic acid monomers was characterized. The isolated oligosaccharides gave further insight into the structures of pectic side chains and may have an impact on plant physiology and dietary fiber fermentation.

Neutral Pectin Side Chains of Amaranth (Amaranthus hypochondriacus) Contain Long, Partially Branched Arabinans and Short Galactans, Both with Terminal Arabinopyranoses

Amaranth is a pseudocereal of high nutritional value, including a high dietary fiber content. Amaranth dietary fiber was suggested to contain large amounts of neutral rhamnogalacturonan I side chains. In this study, endo-arabinanase and endo-galactanase were used to liberate arabinan and galactan oligosaccharides from amaranth fiber. The liberated oligosaccharides were identified by high-performance anion exchange chromatography with pulsed amperometric detection (HPAEC-PAD) and HPLC-MS(n) using standard compounds, which were isolated from amaranth, sugar beet, potato, and red clover sprouts and characterized by one- and two-dimensional NMR spectroscopy. It was demonstrated that insoluble amaranth arabinans have linear and branched areas, with the O-3 position being the dominant branching point. Minor amounts of branches at position O-2 and double substitution were also found. Amaranth arabinans were also demonstrated to contain terminal α-(1→5)-linked l-arabinopyranose units. In addition, it was evidenced that galactans from amaranth seeds are composed of β-(1→4)-linked d-galactopyranose units, which can also be terminated with l-arabinopyranose units. In direct comparison to structural elucidation of amaranth fiber by using methylation analysis, the advantage of the enzymatic approach over methylation analysis was demonstrated.

Studying non-covalent enzyme carbohydrate interactions by STD NMR

Saturation transfer difference NMR spectroscopy is used to study non-covalent interactions between four different glycostructure transforming enzymes and selected substrates and products. Resulting binding patterns represent a molecular basis of specific binding between ligands and biocatalysts. Substrate and product binding to Aspergillus fumigatus glycosidase and to Candida tenuis xylose reductase are determined under binding-only conditions. Measurement of STD effects in substrates and products over the course of enzymatic conversion provides additional information about ligand binding during reaction. Influences of co-substrates and co-enzymes in substrate binding are determined for Schizophyllum commune trehalose phosphorylase and C. tenuis xylose reductase, respectively. Differences between ligand binding to wild type enzyme and a corresponding mutant enzyme are shown for Corynebacterium callunae starch phosphorylase and its His-334-->Gly mutant. The resulting binding patterns are discussed with respect to the possibility that ligands do not only bind in the productive mode.

Pectic arabinans in quinoa seeds (Chenopodium quinoa Willd.) are acylated with p-coumaric acid

A modified digestion method was used to hydrolyze large amounts of quinoa meal to study polysaccharide-bound phenolic acids. Besides the well-characterized oligosaccharide O-(2-O-trans-feruloyl-α-l-arabinofuranosyl)-(1→5)-l-arabinofuranose, O-(2-O-trans-coumaroyl-α-l-arabinofuranosyl)-(1→5)-l-arabinofuranose was isolated from the hydrolysate and identified by LC-MS, GC-MS, and NMR experiments. This study demonstrates for the first time that p-coumaric acid can be linked to pectic side chains of dicotyledonous plants.

Activity-Guided Fractionation to Identify Blue Wheat (UC66049 Triticum aestivum L.) Constituents Capable of Inhibiting In Vitro Starch Digestion

ABSTRACT The inhibition or delay of starch digestion by dietary compounds could help manage postprandial blood glucose levels. The objective of this study was to identify constituents from whole grain blue wheat capable of decreasing α-amylase-catalyzed starch digestion. An activity-guided fractionation approach based on liquid chromatography was used to identify solvent- and alkaline-extractable blue wheat constituents reducing α-amylase-mediated starch digestion in vitro. Fatty acids, potentially released from cell wall polymers by alkaline hydrolysis, inhibited the digestion of amylose, probably through the formation of amylose-lipid complexes. However, the degradation of amylopectin was not affected by fatty acids. In addition, 1-(3,5-dihydroxyphenyl)heneicosan-2-one, a 5-(2′-oxoalkyl)resorcinol, was found to reduce starch digestion. However, because the digestion of both amylopectin and amylose was reduced, the inhibition mechanism was different from that of fatty acids. Further research is needed to...

Effect of Processing on Antioxidant Activity, Total Phenols, and Total Flavonoids of Pigmented Heirloom Beans

While extensive research has been performed on the composition and cooking quality of commodity beans, relatively little is known about pigmented heirloom varieties and the effects of processing on their antioxidant capacity. The aim of this study was to evaluate the effect of soaking and cooking on antioxidants in four heirloom bean varieties compared to Pinto. Water absorption kinetics, soaking and cooking time, DPPH radical scavenging activity, and total phenolic and total flavonoid content were determined in raw, soaked, and cooked samples. Heirlooms required less time to hydrate compared to Pinto, whereas cooking times were similar. The effect of soaking on antioxidant capacity and flavonoids was minimal compared to cooking, which led to losses of up to 57%. Each pigmented heirloom bean had specific characteristics, and three of them had equal or higher amounts of antioxidants or antioxidant activity than Pinto at every processing step. Among heirlooms, Koronis Purple and Jacob’s Cattle had the highest antioxidant activity and Jacob’s Cattle and Tiger’s Eye the highest amount of flavonoids, even after cooking.

Compositional evaluation of perennial wheatgrass (Thinopyrum intermedium) breeding populations

A shift to perennial grain cultivation offers environmental benefits such as reduced soil erosion and nitrogen leaching. Recently, tremendous agronomic accomplishments have been made in domesticating perennial intermediate wheatgrass (Thinopyrum intermedium, IWG). Thirteen IWG breeding populations were evaluated for characteristics relevant to food use, including chemical composition, activity of enzymes responsible for rancidity, content of antioxidants (carotenoids and hydroxycinnamic acids) and in vitro antioxidant activity based on the 2,2-diphenyl-1-picrylhydrazyl and Folin-Ciocalteu assays. IWG populations had higher insoluble dietary fibre, protein, fat and ash, but less starch than wheat. IWG populations were deficient in high-molecular weight glutenins yet contained appreciable amounts of gliadins. Some populations exhibited significantly lower lipase activity than wheat, and all had lower lipoxygenase activities. Most IWG populations contained higher levels of antioxidants than wheat. This work provided key phenotypic information for developing IWG lines that combine high fibre, protein and antioxidant contents with minimal rancidity development.