Phenolic Compound Profile,Antioxidant And Antiproliferative Activities Of Highland Barley

Epidemiological studies have shown that regular consumption of fruits,vegetables as well as whole grains is associated with reduced risks of chronic diseases such as cardiovascular disease,type?diabetes and cancers,the health benefits of these foods are mainly attributed to their phenolic compounds.In whole grains,phenolic compounds exist as free and bound forms,the bound phenolics may provide protective effect to colon.Highland barley was planted in Qinghai-Tibet Plateau of China,it has survival ability under intensive geo-climatic conditions,including poor soil fertility,drought,hypoxia,hypothermia and high irradiance,this may be relate to the components of highland barley.In the present study,the phenolic compound profile,antioxidant,and antiproliferative activities of four highland barley varieties were investigated.(1)Free and bound extracts were extracted from four varieties(BQ,Dulihuang;CHQK,Changheiqingke;ZQ25,Zangqing 25 and ZQ320,Zangqing 320)of highland barley.Total phenolics,total flavonoids and total anthocyanins of these extracts were determined by Folin –Ciocalteu colorimetric,sodium borohydride / chloranil – based assay and spectrophotometric pH differential protocol,respectively.Phenolic compound profile was analysed by HPLC.Total phenolics ranged from 333.9 ± 17.1 to 460.8 ± 5.7 mg GAE/100 g DW,total flavonoids were between 145.5 ± 10.0 and 247.4 ± 20.3 mg CE/100 g DW.The contribution of bound phenolics and bound flavonoids to the corresponding total contents were 38.8 – 49.7% and 26.9 – 44.0%,respectively.Protocatechuic acid,chlorogenic acid,(+)-catechin,caffeic acid,p-coumaric acid and Ferulic acid were identified and quantified in these extracts.Ferulic acid was the dominant phenolic compound,and the contents ranged from 48.14 ± 0.28 to 67.53 ± 0.30 mg/100 g DW.Total phenolics and phenolic compound profile of bran/germ and endosperm fractions of highland barley were also determined.In BQ,CHQK,ZQ25 and ZQ320,the contribution of total phenolics of bran/germ fraction to the whole grains were 72.0,76.8,80.9 and 74.1%,respectively.Ferulic acid and p-coumaric acid presented in bran/germ and endosperm fractions,the contributions of ferulic acid content of bran/germ fraction to the whole grains were 71.7 – 82.5%.These results suggested that highland barley exhibits abundant phenolic compounds,which existed as free and bound forms,and the most phenolics presented in bran/germ fraction.(2)Antioxidant activity of free and bound extracts of highland barley was determined by ORAC,PSC and CAA assays.Total ORAC values(free and bound extracts)of four highland barley varieties ranged from 35.54 ± 2.15 to 44.93 ± 0.85 ?mol TE/g DW,total PSC values of four highland barley varieties ranged from 22.55 ± 1.40 to 30.15 ± 2.41 ?mol Vc E/g DW,the proportion of bound ORAC values and bound PSC values to the corresponding total values were 49.21 – 64.41% and 34.87 – 48.71%,respectively.To evaluate cellular uptake of antioxidant compounds,CAA values both in no PBS wash and PBS wash protocol were determined.In no PBS wash protocol,total CAA values(free and bound extracts)of four highland barley varieties ranged from 185.5 ± 7.9 to 439.1 ± 47.1 ?mol QE/100 g DW;in PBS wash protocol,total CAA values were between 33.85 ± 1.81 and 62.37 ± 7.37 ?mol QE/100 g DW.These results indicated that highland barley extracts show antioxidant activities in chemical and cellular based methods.(3)A multistep enzymatic extraction method was compared with a conventional chemical extraction process to evaluate the phenolic content and antioxidant activity of highland barley(ZQ25,CHQK).The enzymatic digestion yielded a higher retrieval of(+)-catechin compared to the conventional chemical extraction procedure(p < 0.05),while,ferulic acid,p-coumaric acid,caffeic acid and chlorogenic acid contents in enzymatic digested extracts were lower than the chemical extracts.The multi-enzymatic digestion extracts exhibited significantly higher(p < 0.05)antioxidant activities(ORAC and CAA assays)when compared to chemical extracts.These results suggested that highland barley subjected to multi-enzymatic digestion exhibits a higher phenolic content and antioxidant activity than the chemical extraction.(4)Antiproliferative activities of highland barley extracts were investigated,all extracts showed potent antiproliferative activities toward HepG2(human liver cancer cells),MDA-MB-231(human breast cancer cells)and Caco-2(human colon cancer cells).The potential mechanism of CHQK free extracts(CHQK-F)in regulating cell proliferation in Caco-2 human colon cancer cells was investigated,the results showed that CHQK-F suppressed TRAF2 expression,and up-regulated expressions of p-p38,p-p53 and p21,then down-regulated expressions of Cyclin D1,CDK4,PCNA and p-Rb;additionally,CHQK-F activated procaspase 8.These suggested that CHQK-F may induce Caco-2 G1/S cell cycle arrest through p38/MAPK signal transduction pathway,and induce Caco-2 cell apoptosis through extrinsic(death receptor)pathway.