Extraction Of Polyphenols,and Isolation And Identification Of Antioxidant Constituents From Special South China Vegetables(Artemisia Selengensis Turcz And Ipomoea Batatas L.leaves)

Artemisia selengnesis Turcz?AST?and Sweet potato?Ipomoea batatas L.?leaves?SPL?are both special vegetables in South China.They characterize as low utilization and high bioactivity,but the optimun extraction method and clear profiles of antioxidants are all not sure.Therefore,the extraction method of antioxidant constituents in special vegetables of South China were screened,and the antioxidant profiles were characterized by taking AST and SPL as symbols.This research aimed to provide technique reference and theoretical basis for the screening,isolation and identification,bioactivity research as well as high-valuation of AST,SPL and other special vegetables in South China though investigating the antioxidant potent and comprehensive antioxidant profiles.The major results are listed as follows:?1?The 50%ethanol extract?FEE?of AST showed the highest total phenolics content?17.03 mg GAE/g DM?,total flavonoids content?11.11 mg RT/g DM?,along with the strongest DPPH·scavenging ability and reducing power.But the water extract contained the richest total sugar?102.43 mg Glu/g DM?,as well as the best HO·scavenging and Fe²⁺chelating ability.Totally,27 compounds were identified or tentatively identified from FEE by using HPLC-QTOF-MS²,including 11 organic acids,1 amino acid and 15 fatty acids.Notably,26 compounds of them were identified in AST for the first time,and the 15 fatty acids were firstly identified in Artemisia genus.Quinic acid,3-caffeoylquinic acid?3-CQA?,5-CQA,3,4-dicaffeoylquinic acid?diCQA?and 3,5-diCQA were identified as the dominant phenolic acids in AST extract.Meanwhile,the MS/MS fragmentation behaviors of 6compounds including 9,12,13-trihydroxyoctadecenoic acid,et al.were firstly depicted.Above results indicated that 50%ethanol can be the suitable solvent for extracting polyphenols from AST,and AST can be used as a potential natural antioxidants source for developing functional food additives and health-promoting products.?2?Effect of dynamic high pressure micro-fluidization?DHPM?extraction,microwave extraction,ultrasound extraction,high temperature and maceration on the extraction of polyphenols from AST was compared.Microwave extract exhibited the highest total phenolics?40.30?g GAE/mg Extract?and total flavonoids content?37.75?g RE/mg Extract?,as well as the best in vitro AAs.Phenolic acids contributed most to the antioxiand activities of AST extracts,and ten phenolic acids were identified and semi-quantified.The phenolic acid profiles of five extracts were similar,but the content of each compound were quite different.3-CQA,3,5-diCQA and1,5-diCQA were the most abundant phenolic acids in AST.Microwave extract had the highest content in diCQA isomers and 5-CQA,however,high temperature extract showed the lowest content in diCQA isomers and 3-CQA,but the highest 1-CQA and5-CQA content,which suggested that high temperature could promote the degradation of diCQAs,and 4-and 5-caffeoyl was more liable to decompose compared to 1-and 3-caffeoyl.Thus,microwave is the suitable extracting method of AST polyphenols.?3?Ethyl acetate and n-butanol were the suitable solvent for enriching flavonoids and phenolic acids from the extract ofAST leaves.While,the n-butanol?nBuF?showed the best DPPH·scavenging ability(IC₅₀=543.0 mg BHA/g Fr.),ABTS⁺·scavenging ability(IC₅₀=158.2 mg Trolox/g Fr.)and reducing power(IC_0.3=401.6 mg BHA/g Fr.),following by ethyl acetate?EAF?and chloroform fraction.HPLC-QTOF-MS² was carried out to characterize the antioxidant profile of AST leaves.Fifty-seven compounds were detected from EAF and nBuF by HPLC-QTOF-MS² including 11 organic acids,16 flavonoids,16 hydroxycinnamic acids,6hydroxylbenzoic acids,5 amino acids and 3 other compounds,40 of them were reported in AST for the first time.Totally,21 and 38 compounds were identified or tentatively identified in n BuF and EAF,respectively,only quercetin-3-O-galactoside and quercetin-3-O-glucoside were dected in both fractions.Flavonoids were the major compounds in EAF,while phenolic acids and organic acids were the dominant constituents in nBuF.Meanwhile,the MS/MS fragmentation behaviors of five compounds including eupatilin,et al,were herein depicted firstly.Thus,AST leaves might be a potential low-cost resource of natural antioxidants in the field of food processing,health care products abd cosmetics.?4?Taking orange-fleshed and cream-fleshed sweet potato leaves?OFL and CFL?as materials,the potential of DHPM-assisted extraction?DHPMAE?on extracting polyphenols from plants was evaluated by comparing with traditional extraction methods.DHPMAE showed the best extraction efficiency on total phenolics and total flavonoids,it also exhibited the strongest DPPH·and ABTS⁺·scavenging ability,chelating ability and reducing power.In addition,DHPM post-treatment would not lead to the degradation of polyphenols,but microwave and ultrasoud post-treatment did.Therefore,DHPMAE is the best method for extracting polyphenols from SPL,and can be used as a new extraction technique in extracting natural products.Twenty-two compounds were detected in the DHPMAE extracts of OFL and CFL by comparing the difference of OFL and CFL in chemical constituents.Ten compounds of them,p-hydroxybenzoic acid,isopropylmalic acid and fatty acid,et al.,were detected in SPL for the first time.Dihydrokaempferol-glucoside,succinic acid and13-keto-octadeca-9,11-dienoic acid were only found in OFL,while,Glu-Leu/Ile hexose and octadecatrienoic acid dipentose were just detected in CFL.Beside,this research depicted the MS/MS fragmentation behavior of 3 compounds including13-keto-octadeca-9,11-dienoic acid,et al..?5?After successive fractionation of DHPMAE extract by chloroform,ethyl acetate,and n-butanol,the antioxidant constituents were greatly enriched in the ethyl acetate fraction?EAF?.The EAF showed the highest total phenolic?569.38 mg GAE/g Extract?and total flavonoid?43.15 mg RE/g Extract?content,which were much higher than that in nBuFwith the corresponding values at 96.62 mg GAE/g Fr.and 23.79 mg RT/g Fr.,respectively.EAF also yielded the highest in vitro AAs with the IC_{50 DPPH·},Ab_{0.3 reducing power} and IC_{50 ABTS+·}of 1306.5,1840.1 and 702.74 mg Trolox/g Fr.,respectively.Rapid identification of the dominant antioxidants in SPL was established with HPLC-QTOF-MS²,and 37 compounds were identified,including 20 phenolic acids,12 flavonoids,3 organic acids,1 nucleoside and 1 ester.Among them,caffeoyldexopentitol,6''-?4-carboxy-3-hydroxy-3-methylbutanoyl?hyperin and butyl-dicaffeoylquinate,et al.,20 compounds were firstly detected in SPL.CQAs derivatives and flavonoids were the most abundant compounds.In addition,the MS² fragmentation behaviorf of 6''-?4-Carboxy-3-hydroxy-3-methylbutanoyl?hyperin,3,7-dimethylquercetin and Kaempferide-3-O-sulate was demstrated herein for the first time.The results could lay a theoretic foundation for the bioactivity researches of SPL,and provide technique support and reference for the rapid identification of bioactive consitutents in other plants extracts.?6?Bioactivities-guided isolation and identification of DPPH·scavengers and?-glucosidase inhibitors from SPL was performed.By using a series of phytochemistrical methods,9 compounds were isolated and identified from the subfraction?EA-1?with the best DPPH·scavenging ability,including 3,4,5-triCQA?4??3,5-diCQA?6?and 7,3'-dimethylquercetin?13?,et al.Seven compounds were isolated and identified from the subfraction?EA-4?with the strongest?-glucosidase inhibitory,including N-trans-?p-coumaroyl?tyramine?1??trans-N-feruloyltyramine?2?,N-cis-feruloyl-tyramine?3?,et al..Compounds 1-3,8,10,11,13 and 15 were identified in SPL for the first time,compounds 1,2,3 and 10 were firstly found in genus Ipomoea.Compounds 1⁴,10¹2,5 and 7 showed higher?-glucosidase inhibitory than positive control Acarbose,suggesting them to be the dominant?-glucosidase inhibitors in SPL.Compounds 4⁷,9,10,12 and 14 showed higher DPPH·scavenging ability than positive control Vc,indicating them to be the dominat antioxidants in SPL.In addition,higher caffeoylation of quinic acid and lower methoxylation of flavonoids would result in stronger antioxidant activity.Methylation of phenethyl cinnamides and cis structure could decrease their?-glucosidase inhibitory.Aforementioned results could help to explain the antioxidant activities and anti-daibetes of SPL extracts,an lay a foundation for it further application in function food.