| Black soybeans are known to be rich in functional phytochemicals,especially the phenolics including phenolic acids,flavanols,isoflavonoids and anthocyanidins which have effective bioactivities.They have anti-oxidant,anti-inflammatory,neuroprotective and anti-tumor activities.Epidemiological and intervention studies indicate that regular consumption of black beans and relevant products is negatively correlated with the occurrence of a variety of chronic diseases such as diabetes and senile dementia.In China,Black beans were used as traditional Chinese medicine which is also used for the prevention and treatment of cardiovascular diseases,such as hyperlipidemia,atherosclerosis,and estrogen metabolism related diseases such as breast cancer,prostate cancer and osteoporosis,etc.Therefore black beans play an important role in maintaining human health and prevention of chronic diseases.There is few study about the phytochemical composition rich in black beans stability,and their distribution and chemical antioxidant activity in vitro of the research.As for the study of phenolic compounds combining in food matrix and their in vitro antioxidant activity,as well as the research of soluble-type and bound-type phenolics composition and chemical oxidation activity affected by indigestion,it still leaves a gap waiting to be filled.To furtherly study the nutritive value,we divided phenolics of black soybeans into extractable phenolics and bound phenolics,and accurately qualify and quantify them by UPLC-ESI-QTOF-MS/MS and HPLC-ESI-QqQ-MS/MS.Using 96-well plates rapid detection method for determining their and total phenolic content and antioxidant activity affected by in vitro digestion process.The result is presented below:1.A total of 124 compounds in soluble were qualified in phenolics extract of malonyl glucoside);21 soysaponins: deacetylated Soyasaponin Aa,soyasaponin Aa,deoxy Soyasaponin Aa,soyasaponin Ae,soyasaponin Ba,soyasaponin Bb,soyasaponin αg,soyasaponin β,soyasaponinⅡ,soyasaponin Ab,soyasaponin A3,soyasaponin Bb’,soyasaponin Be,soyasaponin βa,soyasaponin Bd,soyasaponin IV,soyasaponin Ah,soyasaponin Ag,soyasaponin Bd and soyasaponin Bb isomer;8 amino acids: aspartic acid,glutamate-tyrosine,tryptophan,glutamate-phenylalanine,L-glutamic acid,phenylalanine,arginine,tyrosine;2 other compounds triisovalerin and uridine.black soybean using UPLC-ESI-QTOF-MS including 6 saccharides: glucose,arabinose,gentiobiose,gentianose,tetrasccharide and hexaose;15 organic acids: gluconic/galactonic acid,galactaric acid/glucaric acid,threonic acid,glucoascorbic acid,malic acid,2,5-diketogluconic acid,xylonic acid,malonic acid,guloascorbic acid,glutaconic acid,2-methyl-cis-aconitic acid,succinic acid,quinic acid,malyngic acid and tartaric acid;6 phenolic acids: 4-hydroxyphenylglycolic acid hexoside,gallic acid,protocatechuic acid glucosyl pentoside,4-hydroxyphenylglycolic acid,protocatechuic acid and chlorogenic acid;15 isoflavonoids: daidzin,glycitin,wistin,daidzein-4’-O-glucoside,genistin 6’’-O-acetate,prunetrin,daidzein,glycitin 6’’-O-acetate,daidzin-6’’-O-acetate,genistin,genistein-5-O-glucoside,genistein,pratensein,glycitein and 3’,4’,7-trihydroxyisoflavone;18 flavonol: quercetin-7-O-hexoside,kaempferol-7-O-B-D-glucoside,vitexin,kaempferol galacoside,myrecetin rutinoside,rutin,3-O-methylmyricetin hexoside,hyperoside,dihydroxyflavone,trihydroxyflavone,isorhamnetin,dihydroxyflavone,quercetin,apigenin,myricetin hexoside,quercetin-3-xyloside 7-glucoside,quercetin 3-lathyroside and quercetin di-pentoside;8 flavanone: naringenin-8-C-β-D-glucoside,taxifolin hexoside,naringenin-7-O-glucoside,naringenin,taxifolin,naringenin-4’-O-glucoside,hemipholin and naringenin-7-O-galacoside;13 flavanol:(epi)catechin-(epi)catechin and its isomers,(epi)catechin-(epi)catechin-(epi)catechin and its isomers,catechin 3-rhamnoside,(epi)catechin-(epi)catechin,epigallocatechin,catechin,procyandin C,epicatechin,procyanidin A2 and cinnamtanin A;12 anthocyanidin: cyanidin,delphinidin,petunidin,pelargonidin,peonidin and their indican derivatives,Pelargonidin-3-O-(6”-malonyl glucoside),Apigeninidin-3-O-(6”-2.A total of 82 phenolic compounds were qualified in bound phenolics extract by using UPLC-ESI-QTOF-MS including 23 phenolic acids: trimethyl citrate,dimethyl itaconate,dimethyl citrate,nonanedioic acid,dimethyl butylene acid,aconitic acid,2,2-bis(hydroxymethyl)mal-onic acid,piscidic acid,glucoascorbic acid,malic acid,2,5-diketogluconic acid,guloascorbic acid,2-methyl-cis-aconitic acid and quinic acid;4-hydroxyphenylglycolic acid,4-methoxyphthalic acid,dihydroxybenzoic acid,3-O-methyl gallic acid,sinapic acid,p-hydroxybenzoic acid,ferulic acid,vanillic acid,acetoxy methoxybenzoic acid,trihydroxybenzoic acid,dihydroxybenzoic acid hexoside,dihydroxybenzoic acid hexoside,woodorien,dihydroxybenzoic acid,dihydroxybenzoic acid hexoside,hydroxy benzenedicarboxylic acid,isoferulic acid,2-hydroxybenzoic acid,gallic acid,methoxysalicylic acid;8 isoflavonoids: Daidzin,Glycitin,Daidzein,Daidzin-6’’-O-acetate,Genistin,Genistein-5-O-glucoside,Genistein and Glycitein;10 flavonol: 3’,4’-DiMethoxy-α-naphthoflavone,Rhamnetin,Myricetin,Hexahydroxyflavone,Kaempferol 7-O-B-D-glucoside,Vitexin,Rutin,Hyperoside,Apigenin,Isorhamnetin and Quercetin;6 flavanone: Hovenitin I,4’ or 5’-O-Methyl-(epi)catechin rhamnoside,Ampelopsin,Prunin,Naringenin and Taxifolin;5 flavanes: Gallocatechin hexoside,Epigallocatechin,Catechin,Epicatechin and Procyanidin A2;4 anthocyanidin: cyanidin,delphinidin,petunidin,pelargonidin;3.Soluble and bound phenolic content and antioxidant activity were determined by using 96-well plates rapid detection method.The TPC of soluble phenolics extract are 38.04 and 10.60 mg GAE/g DW,respectively.Besides,the results of DPPH of coat and cotyledon of blackbean is 119.73 and 21.93 μmol TE/g DW,and those of FRAP and ABTS are 251.84 and 19.94 μmol FE/g DW,165.86 and 8.43 μmol TE/g DW,respectively.This result showed that the content of soluble phenolics in black beans coat is much higher than that of cotyledon,and it is similar as to antioxidant activities.The TPC of bound phenolics in coats are 9.33 GAE/g DW(A-BPE),1.63 GAE/g DW(A-B-BPE),8.84 GAE/g DW(B-BPE)and 3.01 GAE/g DW(B-A-BPE),in cotyledon are 3.67 mg GAE/g DW(A-BPE),1.23 mg GAE/g DW(A-B-BPE),4.19 mg GAE/g DW(B-BPE)and 1.54 mg GAE/g DW(B-A-BPE).The antioxidant activities of coat bound phenolics extracted by different hydrolysis method are 6.76-23.53 TE /g DW(DPPH),6.92-27.16 TE /g DW(ABTS)and 47.53-133.14 μmol FE/g DW(FRAP),those of cotyledon phenolics extract are 0.42-8.79 μmol TE /g DW(DPPH),0.66-5.35 μmol TE(FE)/g DW(ABTS)and 1.82-7.86 μmol FE/g DW(FRAP).The positive colleration between the TPC and antioxidant acitivity means that both bound and soluble phenolics are the major antioxidants in black bean.4.The TPC and antioxidant activity of bound phenolics treated by enzymes and in-vitro digestion were determined by 96-well plates rapid detection method,the TPC of coat bound phenolics are 1.07 mg GAE/g DW(β-glucanase),0.45 mg GAE/g DW(xylanase),10.97 mg GAE/g DW(cellulase),16.52 mg GAE/g DW(hemicellulase),the TPC of cotyledon bound phenolics are 0.51 mg GAE/g DW(β-glucanase),0.54 mg GAE/g DW(xylanase),2.42 mg GAE/g DW(cellulase),1.04 mg GAE/g DW(hemicellulase);the DPPH,ABTS and FRAP results of coat bound phenolics are 2.08-5.78,11.16-165.90 和 4.96-77.19 TE(FE)/g DW,and those of cotyledon bound phenolics are 0-0.52,9.07-36.51 and 3.29-13.56 TE(FE)/g DW.Besides,the TPC of coat extracts in different digestion stage ranged from 6.22 to 13.39 mg GAE/g DW,those of cotyledon extracts ranged from 5.36 to 8.65 mg GAE/g DW.The DPPH,ABTS and FRAP results of coat extracts in different digestion stage ranged from 8.80 to 22.35,25.97 to 60.64 and 24.69 to 117.35 TE(FE)/g DW,respectively.Those of cotyledon extracts ranged from 0 to 2.37,3.18 to 6.90 and 2.71 to 7.83 TE(FE)/g DW. |
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