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

Abstract

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.

Enzymatic Mechanism for Arabinan Degradation and Transport in the Thermophilic Bacterium Caldanaerobius polysaccharolyticus

“…Furthermore, very little knowledge was gained on the activity of CpAbf127A and CpAbf127B from the hydrolytic assays with the polymeric substrates or linear, ␣-1,5-linked arabino-oligosaccharides. Therefore, various branched arabino-oligosaccharides, previously isolated from different plant materials (6,7,27), were incubated with the purified proteins. These oligosaccharides did not allow further characterization of CpAbp27A, since the available oligosaccharides contain only arabinofuranose units.…”

“…Arabinan and debranched arabinan were purchased from Megazyme (Bray, Ireland). Branched arabino-oligosaccharides were isolated from various plant materials and characterized by HPAEC-PAD, liquid chromatography-mass spectrometry (LC-MS), and NMR spectroscopy (6,7,27). Other chemicals and sources of materials are described in the relevant paragraphs below.…”

Characterization of Dietary Fiber Polysaccharides from Dehulled Common Buckwheat (Fagopyrum esculentum) Seeds

“…Because methylation analysis yields limited information about specific structural elements (Wefers et al 2014), buckwheat insoluble and soluble fiber were digested with endo ‐arabinanase and endo ‐galactanase to liberate mostly branched oligosaccharides. Information about specific substituents and backbone substitutions of arabinans and galactans are obtained by using this enzymatic‐chromatographic approach (Wefers et al 2015a). The HPAEC‐PAD chromatogram of the endo ‐arabinanase hydrolysate of buckwheat insoluble fiber is shown in Figure 2.…”

“…3). Coelution was reported for the linear, O 3‐branched, and O 2‐branched arabinan tetramers when analyzed by HPAEC‐PAD (Westphal et al 2010b; Wefers et al 2015a). By using LC‐PGC‐MS 2 , it is, however, possible to separate these oligosaccharides.…”

“…Buckwheat insoluble and soluble fiber hydrolysates were analyzed for diagnostic oligosaccharides as described previously (Wefers et al 2015a). Standard compounds for this procedure were isolated from different plant materials as also described by Wefers et al (2015a). Briefly, 10 mg of insoluble fiber or dialyzed soluble fiber (24 h dialysis against demineralized water, molecular weight cutoff of 3,500, water changed three times) was incubated with endo ‐arabinanase (EC 3.2.1.99, from A. niger , 9 U/mg, Megazyme) and endo ‐galactanase (EC 3.2.1.89, from A. niger , 408 U/mg, Megazyme) for 24 h at 40°C (2 U/100 mg of fiber).…”