Physicochemical Properties,Solution Property And Structural Characteristics Of β-glucans And Arabinoxylans From Hull-less Barley And Oat

Cereals are abundant in non-starch polysaccharides（NSPs）,including glucan,arabinoxylan and cellulose.Meanwhile,the functional and nutritional value ofβ-glucan and arabinoxylan are increasingly concerned.In this paper,hull-less barley（Hordeum vulgare Linn.var.nudum Hook.F.）and oat（Avena Nuda）,which are rich in NSPs but undervalued in China,were chosen as the research objects to continuously extractβ-glucan and arabinoxylan.Then,physicochemical properties,solution properties and solid morphologies of purifiedβ-glucans and arabinoxylans in hull-less barley and oat were determined by chemical and instrumental analysis,combined with methylation and nuclear magnetic resonance（NMR）techiques.Main contents and results of the present study are as follows:（1）β-glucan was prepared from hull-less barley,and its physicochemical properties,solution properties and structure features were characterized.Crude polysaccharide（HBBG）was obtained from hull-less barley by hot water extraction,enzymatic hydrolysis,ethanol precipitation,and freeze-drying.Gradient ethanol precipitation was successfully applied to purify the HBBG.The fraction（HBBG-20）was a homogeneousβ-glucan（78.0%）,and detected by high performance gel permeation chromatography（HPGPC）and monosaccharide composition（only composed of 83.2%glucose）.The water absorption capacity（WAC）of HBBG-20 was14.5 g/g at 25℃,while the solubility was 11.7%at 30℃.The results of high performance size-exclusion chromatography connected with multi-angle laser light scattering（HPSEC-MALLS）showed that the weight-average molecular weight（M_w）of HBBG-20 was 3.6×10⁴ g/mol,and it presented random coil conformation in 0.1mol/L Na NO₃ solution.The microstructure of HBBG-20 was mainly composed of linear structures,along with some spherical shapes observed by scanning electronic images（SEM）.Thermogravimetric analysis（TGA）showed that HBBG-20 had the highest mass loss rate（61.1%）at 300～355℃.Structural characteristics investigated by methylation analysis and 1D/2D NMR spectroscopy suggested that HBBG-20 was an unbranched polysaccharide composed ofβ-（1→3）-andβ-（1→4）-linked glucose residues,and the ratio of（1→4）to（1→3）was 2.53.Matrix-assisted laser desorption ionization-time of flight mass spectrometry（MALDI-TOF/MS）was used to analyzed the ratio of 3-O-β-cellobiosyl-D-glucose（DP3）to 3-O-β-cellotriosyl-D-glucose（DP4）（DP3/DP4）in the lichenase hydrolysate of HBBG-20（HBBG-20-L）,and the value was1.47.Thus,the possible molecular structure of the main units for HBBG-20 was proposed as follows:（?）（2）Arabinoxylan was prepared from hull-less barley,and its physicochemical properties,solution properties and structure features were characterized.The crude arabinoxylan（HBAX）was obtained from the water-extracted residues of hull-less barley by saturated barium hydroxide solution extraction,enzymatic hydrolysis and ethanol precipitation,then a highly pure and homogeneous arabinoxylan（HBAX-60）was fractioned by 60%ethanol precipitation.HBAX-60,a neutral polysaccharide,mainly consisted of arabinose（40.7%）and xylose（59.3%）,and the ratio of arabinose to xylose（A/X）was 0.69.The WAC of HBAX-60 was 8.4 g/g at 25℃,and the solubility was 42.1%at 30℃.HBAX-60 presented random coil conformation in 0.1mol/LNa NO₃,while its surface was composed of fibrous shapes,and some aggregated spherical structures intertwined with linear chains.The loss rate of HBAX-60 was the highest at 240～335℃.After being heated to 600℃,HBAX-60 retained 9.8%of its initial mass.Methylation and NMR analysis indicated that a low-branchedβ-（1→4）-xylan existed in HBAX-60,comprising of un-substituted（1,4-linkedβ-Xylp,36.2%）,mono-substituted（β-1,3,4-linked Xylp,5.9%）,and di-substituted（1,2,3,4-linkedβ-Xylp,12.1%）xylose units as the backbone,with four possible branches or individuals included.The possible molecular structure of core unit for HBAX-60 was proposed as follows:（?）where R₁ represeted the terminalα-Araf-(1→orβ-Xylp-（1→might be substituted at O-3 of the→4）-β-Xylp-(1→residue,while R₂ represtedα-Araf-(1→,β-Xylp-(1→,β-Xylp-（1→3）-α-Araf-(1→,α-Araf-（1→3）-α-Araf-(1→.（3）β-glucan was prepared from oat and its physicochemical properties,solution properties and structure features were characterized.Crude polysaccharide（OBG）was extracted by hot water,enzymatic purification,ethanol precipitation,and freeze-drying.Gradient ethanol precipitation was chosen to purify OBG,and the sub-fraction OBG-20（20%ethanol precipitate）was chosen as an ideal research target due to its high yield（60.1%）and good homogeneity.The WAC of OBG-20 was 10.3 g/g at room temperature,while the solubility was 9.9%at 30℃.OBG-20 mainly exhibited random coil conformation in 0.1 mol/L Na NO₃ solution,and linear and fibrous structure with a small amount of spherical structure in its solid morphology by SEM.The loss rate of OBG-20 was the highest at 300～360℃,and the final mass retained 13.2%at 600℃.Methylation analysis and 1D/2D NMR spectroscopy suggested that OBG-20 was a linear polysaccharide composed of mixedβ-（1→3）-andβ-（1→4）-linked glucose residues,and the ratio of（1→4）/（1→3）was 2.02.MALDI-TOF/MS was applied to analyze the DP3/DP4 ratio in the lichenase hydrolysate of OBG-20（OBG-20-L）,and the value of DP3/DP4 was 1.63.Thus,the possible core unit for OBG-20 was proposed as follows:（4）Arabinoxylan was prepared from oat,and its physicochemical properties,solution properties and structure features were characterized.The alkali-extracted arabinoxylan in oat（OAX）was prepared by saturated barium hydroxide solution extraction,enzymatic hydrolysis and ethanol precipitation.A homogeneous arabinoxylan（OAX-L-80）was purified from OAX by lichenase hydrolysis and 80%ethanol precipitation.OAX-L-80 mainly consisted of neutral polysaccharides（88.2%）,and the content of arabinoxylan was 86.0%.The WAC was 6.7 g/g（25℃）and the solubility was 61.5%at 30℃.In 0.1 mol/L Na NO₃ solution,OAX-L-80 mainly showed random coil conformation,while the solid morphology presented irregular spherical,with some ribbon-like and small diameter fibrous structures.The loss rate of OAX-L-80 was 64.8%in the range of 242℃to 330℃,and the total mass loss was 95.6%at600℃.Methylation and NMR analysis of OAX-L-80 indicated that the presence of a low-branchedβ-（1→4）-xylan,composed of un-substituted（1,4-linkedβ-Xylp,27.2%）,mono-substituted（1,2,4-linked and 1,3,4-linkedβ-Xylp,30.4%）,and di-substituted（1,2,3,4-linkedβ-Xylp,6.2%）xylose units as backbone,with four possible branches or individuals included,mainly in the form ofα-Araf-(1→（28.4%）.The possible molecular structure of core unit for OAX-L-80 was proposed as follows:（?）where R₁ represeted the terminalα-Araf-(1→orβ-Xylp-(1→,while R₂ represtedα-Araf-(1→,β-Xylp-(1→,β-Xylp-（1→5）-α-Araf-(1→,α-Araf-（1→5）-α-Araf-(1→.In conclusion,the chemical compositions,solution properties and structural characteristics of NSPs（β-glucan and arabinoxylan）extracted from hull-less barley and oat in China were identified systematically and comprehensively.The main structure of barley and oatβ-glucans were mixed linear（1→3）（1→4）-β-glucans,without adjacentβ-（1→3）linkages.Arabinoxylans were mainly composed of a linear backbone ofβ-（1→4）-xylan with low branching degree,which may be substituted at O-2 and/or O-3positions withα-arabinofuranosyl units.They mainly presented random coil conformation in 0.1 mol/L Na NO₃ solution.This work will provide a structutal foundation for the functional activity and pros development of NSPs in hull-less barley and oat.