Radzikow, 05-870 Blonie, Poland High molar mass water-extractable arabinoxylans in wheat grain: Variation in the level, molar mass and relationship with grain extract viscosity Malgorzata R. Cyran1*, Krzysztofa K. Snochowska1 and Tadeusz Śmiałowski2 1Department of Plant Biochemistry and Physiology, 2Department of Seed Science and Technology, Institute of Plant Breeding and Acclimatization - National Research Institute (IHAR-PIB), Radzikow, 05-870 Blonie, Poland INTRODUCTION Cell wall arabinoxylans (AX) are the major dietary fiber (DF) constituents in wheat grain, exerting positive health effects. The water-extractable counterparts with unique viscous potential are implicated in lowering risk factors of coronary heart disease and diabetes [1]. During the past decade, an intensive breeding for bread wheat with high viscous potential was focused on increased grain water extract viscosity (WEV) and water-extractable (WE) AX content. However, molar mass distribution of these polysaccharides is a key determinant of their viscosity-generating ability that has never been used in wheat breeding. AIM The aim of this study was to develop a routine, small-scale method for evaluation of high-molar mass (HMM) WE-AX level and weight average molar mass (Mw) to examine the extent of their variations in breeding wheat population and relationship with grain WEV. The HMM-AX of wheat grains that eluted as a first peak in the HPSEC profile of WE-DF polysaccharides (Fig. 1) were completely digested by β-xylanase. The second peak in HPSEC profile represented highly branched AX associated with β-glucans, which were digested by β-xylanase combined with α-arabinofuranosidases (AXH-m and AXH-d3) acting on mono- and di-substituted branches of AX, respectively. They were characterized by ten times lower Mw. The HPSEC profiles of WE-DF polysaccharides (Fig. 2) revealed variation in Mw (coefficient of variation, CV=10%) and in a relative proportion of HMM-AX (CV=12%) in wheat breeding materials (Fig. 3 and Fig. 4, Table 1). EXPERIMENTAL Seven winter wheat (Triticum aestivum L.) cultivars and fifty one breeding lines, grown at different Plant Breeding Stations in Poland in 2017, were analyzed. The WE-DF fractions were isolated from whole-meals by the AOAC 985.29 DF method modified appropriately to perform direct separation of high-, intermediate- and low-molar mass subunits on three Shodex OHpak columns (SB-807, SB-804 and SB-806M) with a guard column (SB-807G) as well as to analyze their macromolecular parameters using a HPSEC system (Omnisec Resolve/Reveal, Malvern) equipped with four detectors (RI, RALS/LALS, DV and UV). The DF polysaccharides isolated from wheat whole-meals were treated with specific enzymes to point out the presence of AX and other DF polysaccharides in their HPSEC profiles. The sugar analysis of DF fractions was carried out by gas chromatography of alditol acetates [2]. The viscosity of whole-meal water extract (1:3, w/v) was determined in a Brookfield LVDV-II+ cone/plate rheometer at a constant shear rate [3]. RESULTS The WE-AX content in wheat whole-meal samples investigated varied from 0.63 to 1.13% dm (Table 1). The HMM-AX made of 34 – 48% of the entire WE-AX population. The grain WEV level ranged from 1.44 to 3.58 mPaˑs. Assuming a minimal contribution of intermediate- and low-molar mass polysaccharides to grain WEV, the reduced value of Mw (MwRed) found for HMM-AX, i.e. multiplied by their relative proportion, reflects the Mw of the entire polysaccharide fraction present in the extract (72–143 kDa) (Fig. 5, Table 1). This parameter showed the strongest correlation with grain WEV (r2 = 0.81, Fig. 7), being much stronger than those found for WE-AX content (r2 = 0.64, Fig. 6) and for HMM-AX content (r2 = 0.61), estimated on the basis of sugar and HPSEC analyses. CONCLUSIONS Recently developed HPSEC method allowed insight into proportion and weight-average molar mass of arabinoxylans with the highest molar masses, which control water extract viscosity of wheat grain, and thus, its health effects. The reduced weight-average molar mass of high molar mass arabinoxylans is a new selection index for breeding of wheat cultivars with improved viscous potential. ACKNOWLEDGEMENTS This study was a part of the IHAR-PIB project (3-2-00-0-09, Task No 2.9) financed by the Ministry of Agriculture and Rural Development of Poland. The authors thank breeders from Polish Plant Breeding Companies for providing wheat samples. REFERENCES [1] Jenkins, D.J.A. et al. (2004) Clin. Nutr. Suppl. 1(2), 39–49. [2] Cyran, M.R. & Dynkowska, W.M. (2014) Food Chem., 145, 356–364. [3] Cyran, M.R. & Ceglinska, A. (2011) J. Sci. Food Agric., 91, 469–479.