| Highland barley is the most characteristic crop in Qinghai-Tibet Plateau,it is key poverty alleviation industries and the guarantee of staple food crop and health for people.People exist blindness and following to highland barley because of its nutrition and function of colored barley is rarely studied,Which limits the development of highland barley health industry.Colored highland barley is a precious germplasm resource,An interest in Tibetan highland barley has been steadily increasing because of their abundant phenolic antioxidants.Research has reported composition and content of polyphenols and their activities.However,there have been no reports regarding research on the difference on nutritions、phenolic compounds and their biological activities in colored highland barley,separation and purification of special polyphenols from the highland barley did not report.The special value of highland barley couldn’t highlight,and affected consumption of colored highland barley and polyphenol products development.Therefore,it is necessary to study the nutrients and polyphenols of colored highland barley to promote its development and utilization.In this study,we analyzed the difference on nutrient and composition of phenolic compounds,and selected excellent varieties from highland barley;The extraction and purification method of highland barley polyphenols has been established to evaluate its antioxidant activity and prevention and treatment of metabolic syndrome,and to identify special phenolics of highland barley.The main results are as follows:（1）The content of nutrients in different colored highland barley was significantly different,the protein content of black highland barley was the highest,the total starch and fat content of blue highland barley is the highest,the amylose content of purple highland barley is higher,the fiber content of yellow highland barley is higher,and the content ofβ-glucan of dark highland barley is higher;The average value of essential amino acids in colored highland barley was close to WHO/FAO recommended value.Lysine was the first limiting amino acid,lysine content was higher in deeper highland barley.Amino acid evaluation showed that the proportion of essential amino acids in blue highland barley protein was the highest,and its amino acid composition was the most consistent with the amino acid model recommended by WHO/FAO,and the nutritional value of black highland barley protein was the highest.52 highland barley varieties were clustered into three categories by cluster analysis.（2）80%acetone extraction of highland barley free phenolics exhibited the highest free phenolics content,varieties and the strongest antioxidant activity.Acidic method was more efficiently extraction method than alkaline,it could release more phenolic substances,which was an effective method for hydrolyzing bound phenols;The result showed that chlorogenic acid,benzoic acid,catechin,quercetin and rutin were the main free polyphenols from 80%acetone extraction of highland barley,Gallic acid,p-coumaric acid,syringic acid,benzoic acid,3,4-dimethoxybenzoic acid,hesperidin and syringic acid were the main bound phenols in acid hydrolysis of highland barley.（3）Colored highland barley grains were rich in phenolic compounds and had a strong antioxidant activity.In addition,there were significant differences among the different color;The majority phenolic compounds of colored highland barley were present in the bound form,their composition and content of free and bound phenolics were difference.The free phenolics mainly consist of protocatechuic acid,chlorogenic acid and vanillic acid,while the bound phenolics mainly consist of p-coumaric acid,benzoic acid and veratric acid and 3,4-dimethoxybenzoic acid.The most abundant phenolic acids in the free phenolic extracts were benzoic acid,phloroglucinol,chlorogenic acid and chlorogenic acid,and in the bound phenolic extracts were gallic acid,p-coumaric acid,benzoic acid and benzoic acid of purple,black,blue and yellow barley,respectively.Catechins was the main flavonoids in free phenolic extracts of colored highland barley.Hesperidin is the main flavonoid in the bound phenolic extracts of purple,black and blue highland barley.Naringin is the main bound flavonoid in yellow highland barley;The correlation analysis results showed that protocatechuic acid,chlorogenic acid,catechin and benzoic acid in the free phenolic extracts were the main contributors to the DPPH·and ABTS·+radical scavenging and FRAP capacity,while 2,4-dihydroxybenzoic acid,benzoic acid,chlorogenic acid,syringic acid,3,4-dimethoxybenzoic acid and kaempferol in the bound phenolic extracts are the main contributors to the free radical scavenging capacity.The results of comprehensive scores from highland barley of purple,blue,black and yellow groups were Yunqing 2,Beiqing 2,947 and Kunlun 15 highland barley.（4）The phenolic substances of milling fractions of highland barley mainly existed in bound forms,The outer bran was the main part rich in phenolic substances;There were significant differences in the milling fractions and content of different forms phenolic acids and flavonoids.Protocatechuic acid was dominant free phenolic acid of the highland barley coarse bran and fine powder,phloroglucinol was the dominant free phenolic acid in highland barley fine bran,and catechin was the dominant free flavonoid in barley coarse,fine bran and powder;The syringic acid was the dominant bound phenolic acid in the coarse,fine bran and fine powder,and the hesperidin is the dominant bound flavone in the coarse bran,bran and fine powder;The coarse bran and fine bran of colored highland barley had strong scavenged DPPH,ABTS·+and FRAP ability,and the distribution of antioxidants had difference in milling fractions of colored highland barley.（5）To select DA201-C,NKA-9,HPD826 and NKA-9 were suitable resin for purification of free phenolic compounds from 947,Yunqing 2,Beiqing 2 and Kunlun 15,respectively,suitable resin for purification of bound phenolic compounds were AB-8、AB-8、DA201-C and X-5;The optimum conditions for adsorption and desorption of free and bound phenolics were established by the dynamic adsorption and desorption process optimization;LC/MS analysis showed that the purified phenolics could significantly increase the content of71.43%and 66.67%monomer phenolics of free and bound phenols in bran,the antioxidant capacity of the purified free and bound polyphenol performed was better than crude sample.（6）Free and bound phenolics from colored highland barley showed inhibitory effects on the proliferation of Hep G2 and MCF-7 cells,and the inhibitory effect on human Hep G2 cells was stronger than MCF-7 cells.Among them,the inhibitory effect of bound phenolic was stronger than free,dark highland barley was stronger than yellow highland barley,and Yunqing 2（purple）was the strongest;Highland barley polyphenols could exert the antioxidant capacity of cells by increasing the activities of GSH-Px,SOD and CAT in cells and reducing the activity of LDH and MDA.The bound phenolic was stronger than free.There are differences among highland barley varieties.（7）In vitro antioxidant activity was used to trace,and purified the phenolic substances in bound phenolics were isolated by polyamide chromatography column and semi-preparative liquid chromatograph.The infrared spectrum analysis showed that the purified material conta ined phenol-OH and methyl（-CH3,-CH2,-CH）,benzene ring structure and carbon-oxygen b ond（C-O-C,C-O）functional groups;2’-hydroxy-3,7,8,4’,5’-tetramethoxyflavone,methyl-2,3-isopropylidene-α-L-rhamnopranoside,sinapoyl malate,kaempferol-glucose-rhamnose-rhamnose and methyl dihydromyricetin were isolated and identified from highland barley by UHPLC-MS/MS for the first time.In addition,gallic acid and syringic acid were also detected.（8）The high and low dose groups of highland barley whole grain powder and polyphenols could reduce the spleen and body weight of high fat mice,significantly reduce blood glucose,glucose tolerance,ACU and Ins levels,TC,TG,HDL and LDL of blood lipid,MDA activity of serum and liver,and enhance the activity level of SOD,CAT,GSH-Px,T-AOC.Black and purple highland barley had better ability of reducing blood sugar,blood lipid and improving antioxidant activity of serum and liver than other highland barley.Among them,whole grains and polyphenols of black highland barley 947 had the strongest effect,which indicates that dark highland barley had stronger effect in preventing and treating metabolic syndrome in vivo.Polyphenols was the main functional factor in highland barley to regulate blood sugar and blood lipid. |
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