Ultrasound-pab Extraction Ntawm Phenolic Acids, Flavonols, Thiab Flavan-3-ols Los ntawm Muscadine Grape Skins thiab noob siv Natural Deep Eutectic Solvents Thiab Predictive Modeling Los ntawm Artificial Neural Networking

Thov hu rauoscar.xiao@wecistanche.comyog xav paub ntxiv

AbstractLub hom phiaj ntawm txoj kev tshawb no yog los tshawb xyuas qhov kev ua tau zoo ntawm 9 lub ntuj sib sib zog nqus eutectic solvents (NDES) nrog kev pab ntawm ultrasound rauphenolic acids, flavonol, thiab flavan-3-ols hauv muscadine txiv hmab txiv ntoo (Carlos) tawv thiab cov noob hauv kev sib piv rau 75 feem pua ethanol. Artificial neural networking (ANN) tau siv los ua kom zoo dua NDES cov ntsiab lus dej, lub sij hawm ultrasonication, solid-to-solvent ratio, thiab extraction kub kom ua tiav qhov siab tshaj plaws extraction yields rau ellagic acid, catechin, thiab epicatechin. Ib qho kev tsim tawm tshiab NDES (#1) muaj choline chloride:levulinic acid: ethylene glycol 1: 1: 2 thiab 2 0 feem pua cov dej tau muab rho tawm ntau tshaj plaws ntawm ellagic acid hauv daim tawv nqaij ntawm 22.1 mg / g. Cov txiaj ntsig no yog 1.73-fold ntawm qhov ntawd los ntawm 75 feem pua ethanol. Kev hloov kho NDES (#3) suav nrog choline chloride: proline: malic acid 1: 1: 1 thiab 30 feem pua cov dej rho tawm ntau tshaj plaws ntawm catechin (0.61 mg / g) thiab epicatechin (0.89 mg / g) hauv daim tawv nqaij, thiab 2.77 mg / g thiab 0.37 mg / g hauv cov noob, feem. Qhov kev pom zoo ntawm ellagic acid hauv daim tawv nqaij siv NDES #1 yog 25.3 mg / g (saib) thiab 25.3 mg / g (kwv yees). Qhov zoo tshaj tawm los ntawm (catechin plus epicatechin) hauv cov noob siv NDES #3 yog 9.8 mg / g (saib) thiab 9.6 mg / g (kwv yees). Txoj kev tshawb no tau pom cov txiaj ntsig zoo ntawm kev xaiv NDES rau polyphenols nyob rau hauv cov xwm txheej zoo.

Thov nias ntawm no kom paub ntxiv

Taw qhia

Ntuj sib sib zog nqus eutectic kuab tshuaj (NDES) yog npaj los ntawm kev sib xyaw hydrogen-bond pub nrog hydrogen-bond txais ntawm qhov tsim nyog molar ratio [1]. Lub melting point ntawm ib feem yuav tsum qis dua lub melting point ntawm lwm tus [1]. Tom qab cua sov thiab sib tov, qhov nruab nrab no dhau los ua kua hauv chav sov. Dej yog ntxiv kom stabilize thiab polarize qhov sib tov. Kev tshawb fawb hauv thaj tsam ntawm phytochemical extraction siv NDES tau nthuav dav vim lawv cov txiaj ntsig zoo extractability thiab solubility. Txawm li cas los xij, ntau yam tseem ceeb ua lub luag haujlwm tseem ceeb thaum sib piv NDES rau cov kuab tshuaj organic, suav nrog kev tawm los, nqi, rov qab, thiab toxicity. Kev tshawb fawb yav dhau los tau tshawb xyuas NDES ntawm kev rho tawm cov polyphenols sib txawv los ntawm ntau yam khoom noj khoom haus matrices. Piv txwv li, Bubalo et al. (2016) piv 5 NDES, dej, 70 feem pua methanol (v/v), thiab acidified 70 feem pua methanol (v/v) kom rho tawm anthocyanins, catechin, thiab quercetin -3-O-glucoside los ntawm cov tawv nqaij liab. NDES muaj xws li choline chloride: oxalic acid (1: 1) nrog 25 feem pua ntawm cov dej (v/v) tau pom tias yog cov kuab tshuaj extraction zoo tshaj plaws [2]. Hauv lwm txoj kev tshawb fawb, Pani'c et al. (2019) kuaj 8 NDES thiab acidified 70 feem pua ntawm ethanol thiab pom choline chloride: citric acid (2: 1) nrog 30 feem pua dej (v/v) raws li NDES zoo tshaj plaws los rho tawm anthocyanins los ntawm txiv hmab txiv ntoo pomace [3]. Muscadine txiv hmab txiv ntoo (Vitis rotundifolia) yog ib txwm nyob rau sab qab teb sab hnub tuaj xeev thiab thawj cultivated qus txiv hmab hauv Tebchaws Meskas [4]. Muscadine grapes yog tsim nyob rau hauv 12 lub xeev thiab tag nrho txog 5000 daim av [5]. Muaj 100 ntau yam ntawm muscadine txiv hmab txiv ntoo thiab txhua qhov sib txawv hauv lub cev, kev xav, lossis tshuaj lom neeg yam ntxwv [4]. Ntawm lawv, Carlos yog cog cog muscadine dav vim nws cov qoob loo siab thiab loj hlob zoo sib xws [4]. Carlos muscadine txiv hmab txiv ntoo yog nruab nrab, bronze nyob rau hauv cov xim, thicker nyob rau hauv daim tawv nqaij thiab muaj plaub noob nyob rau nruab nrab [6]. Muscadine grapes muaj ntau npaum li cas ntawmpolyphenolsuas paub tias txo qhov mob [7], inhibit prostate qog loj hlob [8], thiab txhim kho metabolic teb ntawm cov ntshav qab zib [9]. Muscadine grape pomace, ib tug byproduct ntawm muscadine txiv hmab txiv ntoo juicing los yog winemaking, muaj ntawm daim tawv nqaij thiab noob. Kev tshawb fawb yav dhau los siv acetone: dej: acetic acid sib tov (70:29.7:0.3, v/v) kom rho tawm cov phenolic tebchaw los ntawm cov noob, tawv nqaij, thiab pulp ntawm yim cultivars ntawm Florida-loj hlob muscadine txiv hmab txiv ntoo, nrog rau Carlos [10]. Txawm li cas los xij, kev siv cov kuab tshuaj flammable organic thiab lawv cov extraction tsis tshua muaj kev cuam tshuam rau kev siv tswv yim. Feem ntau ntawm muscadine txiv hmab txiv ntoo pomace tseem muab pov tseg ua pov tseg. Artificial neural networking (ANN) yog ib daim ntawv qhia uas tsis yog-linear uas muaj ntau yam kev ua haujlwm yooj yim txuas nrog cov koom haum hnyav. Cov txheej txheem no hu ua "neurons" [11] . Artificial neural networking yog ib txoj hauv kev kawm tshuab los kwv yees lossis kwv yees cov lus teb raws li ntau qhov kev nkag [11]. Kev tshawb fawb ua ntej tau siv cov lus teb saum npoo av (RSM) rau kev tshem tawm kom zoo thiab kev kwv yees. Txawm li cas los xij, qee qhov kev tshawb fawb tau siv ANN rau tib lub hom phiaj. Piv txwv li, Sinha et al. (2013) tau qhia tias ANN muaj kev kwv yees ua tau zoo dua li RSM ntawm kev rho tawm cov xim ntuj los ntawm cov noob ntawm Bixa Orellana (Annatto) [12]. Hauv kev tshawb fawb zoo sib xws, Ciric et al. (2020) tau tshaj tawm tias ANN qauv zoo dua li RSM rau kev kwv yees phenolic compound extractions los ntawm qej [13]. Lub hom phiaj ntawm qhov kev tshawb fawb no yog los tshawb xyuas qhov kev ua tau zoo ntawm 9 NDES rau phenolic acids, flavonols, thiab flavan{17}}ols piv rau 75 feem pua ethanol nrog kev pab ntawm ultrasound. ANN tau siv los kwv yees thiab ua kom zoo tshaj qhov kev rho tawm ntawm phenolic tawm los. Qhov kev xav tau yog tias NDES nrog cov ntsiab lus tshwj xeeb rho tawm ntau dua ntawm phenolic acids, flavonols, thiab flavan-3-ols ntau dua 75 feem pua ethanol, thiab cov txiaj ntsig zoo tshaj plaws tuaj yeem ua tiav los ntawm ANN-raws li kev kwv yees qauv.

2. Cov ntaub ntawv thiab cov txheej txheem 2.1. Cov tshuaj thiab reagents Choline chloride, levulinic acid, 1, 2-propanediol, DL-malic acid, oxalic acid, hydrochloric acid, thiab formic acid tau txais los ntawm Acros Organics (Morris Plains, NJ, USA). Lactic acid, ethylene glycol, glycine, HPLC-qib acetonitrile, methanol, thiab ethanol tau yuav los ntawm Fishers Scientific (Waltham, Massachusetts, USA). L-proline thiab betaine hydrochloride tau yuav los ntawm Alfa Aesar (Ward Hill, MA, USA). HPLC-qib cov qauv ntawm ellagic acid, gallic acid, ferulic acid, (ntxiv )-catechin, (− )-epicatechin, myricetin, quercetin, thiab kaempferol tau txais los ntawm Sigma Aldrich (St. Louis, MO, USA).

2.2. Design of NDESNDES #1–2 hauv Table 1 tau tsim los hauv peb txoj kev kawm dhau los [14]. Choline chloride hauv NDES #1–2 raug xaiv los ua tus neeg txais hydrogen, thaum ob tus neeg pab hydrogen sib txawv raug xaiv rau txhua NDES tshiab. Molar ratios ntawm hydrogen pub thiab txais thiab cov ntsiab lus dej tau txiav txim siab hauv kev sim ua ntej. NDES #3–9 hauv Table 1 raug xaiv los ntawm cov ntaub ntawv raws li cov kev tshawb fawb yav dhau los tau xaiv lawv ua NDES zoo hauv kev rho tawm cov polyphenols. Cov ntsiab lus dej hauv NDES #3 tau hloov kho los ntawm cov ntaub ntawv sau tseg. Ib txoj kev cua sov tau siv los npaj NDES [15]. Luv luv, tus neeg txais nyiaj hydrogen tau sib xyaw nrog txhua yam ntawm hydrogen-daim ntawv cog lus pub khoom hauv Erlenmeyer flasks nrog lub nplawm. Qhov sib tov hauv lub khob ntim tau kaw thiab ua kom sov ntawm 50 ◦C rau li 30 min lossis kom txog thaum cov kua ntshiab tsim thiab nyob ruaj khov ntawm chav tsev kub. Cov ntsiab lus dej hauv Table 1 raug xam raws li qhov kawg ntim ntawm NDES sib tov. Cov pH ntawm NDES teev nyob rau hauv Table 1 yog ntsuas siv pH meter (AB15, Accumet, Fisher Scientific, Waltham, MA, USA). 2.3. Cov qauv kev npaj / Ultrasound-pab extraction Frozen muscadine grape (Vitis rotundifolia) tawv thiab noob (cultivar: Carlos) tau muab los ntawm Paulk Vineyards (Wray, Georgia, USA). Tom qab tshem tawm cov hulls, nplooj, los yog petioles, pomace tau muab cais rau hauv cov noob thiab daim tawv nqaij. Cov qauv no tau qhuav siv lub tshuab nqus tsev (Isotemp, Model 285A, Fisher Scientific, Waltham, Massachusetts, USA) ntawm 60 ◦C thiab lub tshuab nqus tsev siab qis dua − 30 in.Hg. Tom ntej no, cov qauv no tau homogenized rau hauv cov hmoov zoo uas siv lub tshuab sib tsoo (A1{{105}}00, RRH Inc., 2800 W, Zhejiang, Suav). Siv qhov sib piv ntawm qhov sib piv ntawm 1:20 (g: mL), 0.50 g ntawm daim tawv nqaij muscadine los yog cov noob tau muab sib xyaw rau hauv 10 mL NDES lossis 75 feem pua ethanol hauv triplicates. Cov qauv no tau muab tso rau hauv lub da dej (60 ◦C) thiab sonicated (VCX 1500, Sonics & Materials Inc., 1500-Watt, 50/60 Hz, Newtown, CT, USA) rau 30 min ntawm 100 feem pua. amplitude rau ob rounds (15 min / round). Tom ntej no, cov qauv no tau centrifuged tam sim ntawd (Sorvall ST 8, Fisher Scientific, Suzhou, Tuam Tshoj) ntawm 3,260 g kom txog thaum tau txais ib qho supernatant ntshiab. Thaum kawg, cov supernatants tau sau thiab khaws cia rau hauv − 20 ◦C lub freezer rau HPLC tsom xam phenolic acids (ellagic acid, gallic acid, ferulic acid), flavononls (myricetin, quercetin, thiab kaempferol), thiab flavan-3-ols. (catechin thiab epicatechin). 2.4. HPLC kev tshuaj xyuas ntawm phenolic acids, flavonols, thiab flavan-3-ols Phenolic acids, flavononls, thiab flavan-3-ols tau txheeb xyuas ntawm HPLC system (Agilent Technologies 1200, Waldbronn, Lub teb chaws Yelemees) raws li txoj kev tau piav qhia hauv Sandhu thiab Gu (2013) [16]. Lub HPLC system muaj ib lub twj tso kua mis binary, ib qho autosampler, ib lub thawv thermostatted, ib lub diode array detector, thiab lub fluorescence detector. Grape daim tawv nqaij los yog noob rho tawm tau hydrolyzed ua ntej kev soj ntsuam ntawm phenolic acids thiab flavonols. Lub hydrolysis tau ua los ntawm kev sib xyaw 1 ml ntawm cov extract nrog 4 mL ntawm hydrolysis kua (1.2 M HCI uas muaj 50 feem pua methanol) thiab muab tso rau hauv dej da dej (Precision, Model 2837, 400 W, 50/60 Hz, Thermo Scientific, Marietta , OH, USA) ntawm 90 ◦C rau 80 min. Tom ntej no, cov qauv tau txias mus rau 25 ◦C tom qab sonication rau 5 min. Lub hydrolysis ntawm cov extract tsis xav tau rau kev tsom xam ntawm catechin thiab epicatechin. Cov hydrolyzed thiab un-hydrolyzed extracts tau lim los ntawm 0.45 μm polytetrafluoroethylene (PTFE) membrane ua ntej HPLC tshuaj ntsuam xyuas. Rau kev tshuaj xyuas ellagic acid, gallic acid, ferulic acid, myricetin, quercetin, kaempferol, catechin, thiab epicatechin, 10 µL raug txhaj rau hauv kab SB-C18 (4.6 × 250 mm, 5 µm, Zorbax, Agilent, Santa Clara, CA, CA, USA). Cov theem mobile yog (A) 0.5 feem pua formic acid thiab (B) 100 feem pua acetonitrile. Tus nqi ntws yog 1 mL / min nrog 25 min hloov gradient raws li hauv qab no: 0-5 min, 10-30 feem pua B; 5–10 feeb, 30–40 feem pua B; 10–20 feeb, 40–50 feem pua B; 20–25 feeb, 50–10 feem pua B; ua raws li 5 feeb ntawm kev sib npaug. Qhov ntsuas kub ntawm kem tau teem rau ntawm 30 ◦C. Kev kuaj pom wavelength yog 260 nm rau ellagic acid, gallic acid, thiab ferulic acid thiab 360 nm rau myricetin, quercetin, thiab kaempferol ntawm photodiode array detector. Lub excitation thiab emission rau catechin thiab epicatechin yog 230 nm, 321 nm, raws li, siv lub fluorescence detector. Polyphenol tebchaw raug suav hais tias siv cov qauv nkhaus ntawm ellagic acid, gallic acid, ferulic acid, myricetin, quercetin, kaempferol, catechin, thiab epicatechin. Txhua tus qauv nkhaus muaj 7 ntsiab lus thiab R2> 0.99. 2.5. Customized tsim rau Artificial neural networking Plaub qhov kev ywj pheej extraction kuj muaj plaub theem: dej cov ntsiab lus (15-60 feem pua), ultrasonication lub sij hawm (5-35 min), solid-to-solvent ratio (1:5-1:20), thiab extraction kub (30–60 ◦C) (Table S1) tau siv los ua kom zoo tshaj qhov kev rho tawm tawm ntawm phenolic acids, flavononls, thiab flavan-3-ols. Tsis zoo li cov qauv tsim classic xws li cov lus teb tsim qauv, ANN-raws li tsim tsis tas yuav rov ua dua thiab nyiam cov ntaub ntawv sib txawv. Hauv peb txoj kev tshawb fawb dhau los [14], ANN yog ib txoj hauv kev txhim khu kev qha ntau dua rau kev kwv yees cov txiaj ntsig rho tawm dua li RSM. Yog li, ANN tau raug xaiv nyob rau hauv txoj kev tshawb no los kwv yees qhov kev rho tawm ntawm ellagic acid, catechin, thiab epicatechin. Kev tsim kho nrog 40 khiav (Table S2) tau tsim los ntawm JMP Pro (Version 14.2, SAS Institute Inc., Cary, NC, USA) los muab cov ntaub ntawv tshwj xeeb rau ANN kev kwv yees ua qauv. Randomization ntawm 40 khiav tau siv los tshem tawm kev tsis ncaj ncees. Qhov sib npaug tseem ceeb ntawm ANN yog qhia raws li hauv qab no: lub=∑jj=1 wh jpg plus bhk, k=1toK (1) qhov twg h yog tus naj npawb ntawm cov neurons hauv txheej zais, j thiab k yog tus naj npawb ntawm input variables thiab zais cov neurons, ntsig txog, p yog qhov sib txawv input, bh yog qhov tsis ncaj ncees ntawm cov txheej txheem zais, thiab wh yog qhov hnyav hauv txheej zais. Extraction yields ntawm ellagic acid, catechin, thiab epicatechin nyob rau hauv kev sib raug zoo rau plaub lub ywj siab sib txawv tau soj ntsuam siv ANN los ntawm kev cob qhia cov ntaub ntawv ua ntej thiab ces xaiv qhov zoo tshaj plaws ua kom hom thiab ib tug xov tooj ntawm neurons uas ua rau ib tug txaus haum ntawm cov ntaub ntawv. Txhawm rau ntsuas qhov ua tau zoo ntawm cov qauv kev twv ua ntej, peb qhov tseem ceeb tau soj ntsuam: R-square, lub hauv paus square ntawm qhov nruab nrab ntawm qhov kev kwv yees yuam kev (RASE) ( kab zauv (2)), thiab qhov nruab nrab tsis meej (AAE). RASE yog RASE=̅̅̅̅̅̅̅̅̅̅̅̅̅ SSE/n √ (2) Qhov twg SSE pub nyiaj rau square thiab suav qhov kev twv ua yuam kev (qhov sib txawv ntawm qhov tseeb cov lus teb thiab cov lus teb kwv yees) thiab n rau ntau qhov kev soj ntsuam. R-square ze rau 1 nrog RASE thiab AAE ze rau xoom txhais tau hais tias ntau dua haum ntawm cov ntaub ntawv rau hauv tus qauv. 2.6. Statistics Extraction yields ntawm phenolic acids, flavonols, thiab flavan{142}}ols tau muab piv nrog ib-txoj kev ANOVA ua raws li Tub Kawm Ntawv's t-test ntawm p Tsawg dua los yog sib npaug rau 0.05 siv JMP Pro (Version 14.2, SAS Institute Inc., Cary, NC, USA). Txhua nde thiab 75 feem pua ethanol tau muab piv nrog Dunnett qhov kev xeem ntawm p Tsawg dua lossis sib npaug rau 0.05. Lub hauv paus ntsiab lus tsom xam (PCA) tau ua tiav ntawm JMP Pro (Version 14.2, SAS Institute Inc., Cary, NC, USA) rau cov phenolic tebchaw muab rho tawm los ntawm muscadine txiv hmab txiv ntoo tawv thiab noob. 3. Cov txiaj ntsig thiab Kev Sib Tham 3.1. Polyphenols muab rho tawm los ntawm NDES los ntawm muscadine txiv hmab txiv ntoo tawv Nine NDES thiab 75 feem pua ethanol tau siv rau kev rho tawm ntawm polyphenols los ntawm muscadine txiv hmab txiv ntoo tawv. Daim duab 2 qhia txog kev rho tawm tawm ntawm ellagic acid, gallic acid, ferulic acid, myricetin, quercetin, kaempferol, catechin, thiab epicatechin. Ellagic acid yog ntau tshaj extractable polyphenol nyob rau hauv txiv hmab txiv ntoo ntawm daim tawv nqaij, ua raws li los ntawm gallic acid thiab ferulic acid, feem. Qhov kev tshawb pom no tau ua raws li cov kev tshawb fawb yav dhau los [17,18]. NDES # 1, # 8, # 7, # 3, # 2 thiab # 9 tau rho tawm ntau dua ntawm ellagic acid hauv daim tawv nqaij ntau dua 75 feem pua ethanol. Qhov siab tshaj plaws extraction tawm los ntawm ellagic acid tau ua tiav los ntawm NDES #1 ua raws NDES #8 ntawm 22.1 ± 2.2 mg / g thiab 21.3 ± 2.5 mg / g, raws li (Table 2). Txawm li cas los xij, tsis muaj qhov sib txawv tseem ceeb ntawm NDES #1 thiab NDES #8 raws li Tub Ntxhais Kawm t-test. Interestingly, NDES #1 tau pom tias yog qhov tsawg kawg nkaus NDES kom rho tawm cov anthocyanins los ntawm

cranberry ntoo [14] Qhov no tau qhia tias NDES #1 tuaj yeem xaiv rho tawm ellagic acid lossis ellagitannins los ntawm cov zaub mov matrix uas kuj muaj anthocyanidins. Cov kev xaiv zoo li no tuaj yeem raug ntaus nqi los ntawm qhov sib txawv ntawm kev sib cuam tshuam ntawm molecular ntawm NDES thiab cov chav kawm phenolic tshwj xeeb. Daim duab S1 ( vaj huam sib luag A) qhia HPLC chromatogram ntawm gallic acid, ellagic acid, thiab ferulic acid muab rho tawm ntawm daim tawv nqaij txiv hmab txiv ntoo los ntawm NDES #1 thiab kuaj pom ntawm 260 nm. Lub 75 feem pua ethanol rho tawm 12.7 ± 1.2 mg ntawm ellagic acid ib gram ntawm daim tawv nqaij grape. Qhov qis tshaj tawm tawm ntawm ellagic acid tau pom nyob rau hauv NDES #4 ntawm 7.44 ± 0.6 mg / g. Lub extraction tawm los ntawm gallic acid los ntawm NDES # 9, # 8, # 1, # 4, # 7, thiab # 3 tau sib piv thiab ntau dua 75 feem pua ethanol. Qhov siab tshaj plaws ntawm gallic acid tau muab rho tawm los ntawm NDES #9 ntawm 10.4 ± 0.5 mg/g, qhov qis tshaj ntawm 5.55 ± {{40}} .1 mg/g tau muab rho tawm los ntawm NDES #5. Qhov siab tshaj plaws ntawm ferulic acid tau muab rho tawm los ntawm NDES #1 ntawm 6.32 ± 0.7 mg/g thiab qhov qis tshaj plaws tau muab rho tawm los ntawm NDES #5 ntawm 3.11 ± 0.{{5{{52} }}}} mg/g. Tsis tas li ntawd, tsis muaj qhov sib txawv tseem ceeb ntawm NDES #1 thiab 75 feem pua ethanol hauv extracting ferulic acid (Table 2). Qhov siab tshaj plaws ntawm catechin thiab epicatechin tau muab rho tawm los ntawm NDES #3 ntawm 0.61 ± 0.1 mg/g thiab 0.89 ± 0.1 mg/ g, raws li (Table 2). Lub caij no, NDES # 3 thiab # 6 tau rho tawm ntau ntau ntawm epicatechin ntau dua li 75 feem pua ethanol. Daim duab S2 ( vaj huam sib luag A) qhia HPLC chromatogram ntawm catechin thiab epicatechin rho tawm los ntawm NDES #3 los ntawm daim tawv nqaij txiv hmab txiv ntoo. Txawm li cas los xij, catechin tsis tau kuaj pom hauv 75 feem pua ethanol extract. Qhov qis tshaj ntawm catechin (0.02 mg/g) thiab epicatechin ({{90}}.14 mg/g) tau muab rho tawm los ntawm NDES #2 thiab NDES # 5 ,ua. Myricetin yog cov flavonol ntau tshaj plaws thiab kaempferol yog qhov tsawg tshaj plaws. Dunnett qhov kev sim tau qhia tias NDEs thiab 75 feem pua ethanol tau muab piv rau kev rho tawm myricetin, quercetin, thiab kaempferol (Table 2). Qhov siab tshaj plaws myricetin tau muab rho tawm los ntawm NDES #1 (1.84 mg / g), tom qab ntawd 75 feem pua ethanol (1.73 mg / g), thiab tom qab ntawd NDES #8 (1.67 mg / g). Qhov siab tshaj plaws quercetin tau muab rho tawm los ntawm 75 feem pua ethanol (0.41 mg/g), NDES #1 (0.40 mg/g), thiab NDES #8 ({ {143}}.38 mg/g). Hauv qhov sib piv, qhov qis tshaj ntawm myricetin thiab quercetin tau muab rho tawm los ntawm NDES #5 ntawm 0.87 mg/g thiab 0.27 mg/g, raws li. Qhov kev tshawb pom no qhia ntxiv txog qhov muaj peev xwm tsis muaj zog tag nrho ntawm NDES #5 kom rho tawm cov polyphenols los ntawm daim tawv nqaij txiv hmab txiv ntoo. Qhov siab tshaj plaws kaempferol tau muab rho tawm los ntawm 75 feem pua ethanol ({152}}.05 mg/g), thiab qhov qis tshaj yog rho tawm los ntawm NDES #5 thiab NDES #6 (0.03 mg/g). Daim duab S1 ( vaj huam sib luag B) qhia HPLC chromatogram ntawm myricetin, quercetin, thiab kaempferol muab rho tawm ntawm daim tawv nqaij los ntawm NDES #1 kuaj pom ntawm 360 nm. Qhov siab tshaj plaws ntawm phenolic acids, flavonols, thiab flavan{103}} ols yog 40.7 mg / g muab rho tawm nrog NDES #1 ua raws li 39.8 mg / g muab rho tawm nrog NDES #8, qhov qis tshaj yog 18.4 mg / g rho tawm los ntawm NDES #5 (Table 2). Cov pH ntawm NDES nyob nruab nrab ntawm 0.3 thiab 3.3 (Table 1). Rsquared correlation ntawm pH ntawm NDEs thiab phenolic acids, flavonols, thiab flavan-3-ols yields tau teev nyob rau hauv Table 2. Qhov tsis muaj kev sib raug zoo ntawm pH thiab extraction yields qhia pH tsis cuam tshuam rau extraction efficiency. 3.2. Polyphenols muab rho tawm los ntawm NDES los ntawm muscadine txiv hmab txiv ntoo Lub zuag qhia tag nrho extraction yields phenolic acids, flavonols, thiab flavan- 3-ols los ntawm grape noob tau pom tsawg dua cov ntawm daim tawv nqaij (Table 3). Qhov ntau tshaj extractable polyphenols nyob rau hauv cov noob yog catechin thiab epicatechin, whereas kaempferol tsis tau kuaj. Cov noob matrix uas muaj roj (13 feem pua , w / w qhuav puag) yog qhov ua tau piav qhia ntawm qhov qis qis ntawm cov phenolic tebchaw los ntawm cov txiv hmab txiv ntoo [19]. Qhov siab tshaj plaws ntawm catechin tau muab rho tawm los ntawm NDES #3 ntawm 2.77 mg / g (Table 3). Cov txiaj ntsig no tau nce siab dua li tag nrho lwm cov NDES thiab 75 feem pua ethanol. Daim duab S2 ( vaj huam sib luag B) qhia HPLC chromatogram ntawm catechin thiab epicatechin muab rho tawm los ntawm NDES #3 los ntawm cov txiv hmab txiv ntoo. Tus nqi qis tshaj ntawm catechin tau muab rho tawm los ntawm NDES #5 ntawm 0.30 mg / g. Tag nrho NDES tab sis NDES # 1, # 2, thiab # 9 tau rho tawm ntau cov epicatechin ntau dua li 75 feem pua ethanol (Table 3). Qhov siab tshaj plaws epicatechin concentrations tau muab rho tawm los ntawm NDES #4 (0.71 mg / g) thiab NDES #5 (0.68 mg / g), qhov qis tshaj tau muab rho tawm los ntawm 75 feem pua ethanol (0.11 mg / g). Gallic acid yog cov extractable phenolic acid ntau tshaj nyob rau hauv cov txiv hmab txiv ntoo, ua raws li ferulic acid thiab ellagic acid, feem. Qhov siab tshaj plaws ntawm gallic acid tau muab rho tawm los ntawm NDES #4 ntawm 0.45 mg / g, ua raws NDES #9 thiab NDES #8. Cov NDEs no tau rho tawm cov kua qaub ntau ntau dua li 75 feem pua ethanol. Tus nqi qis tshaj ntawm gallic acid (0.2 mg / g) tau muab rho tawm los ntawm NDES #3. Qhov siab tshaj plaws extraction

cov txiaj ntsig ntawm ellagic acid tau txais los ntawm NDES #9 (0.26 mg / g) ua raws li NDES #6 (0.17 mg / g), uas tau ntau dua 75 feem pua ethanol. Ib yam li ntawd, NDEs #3 tau rho tawm tus nqi qis tshaj ellagic acid ntawm 0.05 mg/g. Tsis tas li ntawd, NDES # 6, # 7, thiab # 3 tau rho tawm ntau cov ferulic acid ntau dua li 75 feem pua ethanol. Qhov qis tshaj ferulic acid extraction tawm los yog 0.5 mg/g los ntawm NDES #5. Tsis tas li ntawd, ferulic acid tsis tau kuaj pom hauv NDES #9 extract. Qhov no yuav yog vim qhov solubility ntawm ferulic acid qis dua hauv NDES #9 dua li lwm cov NDEs. Qhov siab tshaj plaws myricetin extract tau txais los ntawm 75 feem pua ethanol thiab NDES #7 ntawm 0.18 mg / g, uas siab dua txhua NDEs. Qhov siab tshaj plaws quercetin extraction tawm los yog NDES #6 ({27}}.14 mg / g) thiab NDES #3 (0.13 mg / g) thiab ob qho tib si NDES zoo dua 75 feem pua ethanol. Ib yam li ntawd, pH ntawm NDES tsis cuam tshuam rau kev rho tawm tawm raws li qhia los ntawm kev sib raug qis (R-squared) ntawm pH ntawm NDEs thiab cov txiaj ntsig ntawm phenolic acids, flavonols, thiab flavan-3-ols teev nyob rau hauv Table 3. 3.3 . Kev tsom xam cov ntsiab lus tseem ceeb (PCA) tau ua los koom nrog cov txiaj ntsig ntawm kev rho tawm ntawm ntau yam phenolic tebchaw hauv daim tawv nqaij thiab cov noob nrog NDEs thiab 75 feem pua ethanol (Fig. 1). PCA tau ua tiav ntawm kev sib txheeb matrix txhawm rau txheeb xyuas qhov muaj peev xwm xaiv tau ntawm qee qhov NDEs rau kev rho tawm cov phenolic tebchaw tshwj xeeb lossis pab pawg. Txog 85 feem pua ntawm qhov sib txawv ntawm cov ntaub ntawv ntawm daim tawv nqaij tau piav qhia los ntawm cov ntsiab lus tseem ceeb 1 thiab 2. Cov phiaj xwm thauj khoom (Daim duab 1B) qhia txog kev sib raug zoo ntawm phenolic acids (ellagic acid, gallic acid, ferulic acid) thiab flavonols (myricetin, quercetin, thiab kaempferol). Txhawm rau rho tawm cov pab pawg no, cov kuab tshuaj zoo tshaj plaws yog NDES #1, #8, #7, thiab 75 feem pua ethanol raws li qhia hauv cov qhab nia (Fig. 1A). Lub caij no, catechin thiab epicatechin tshwm sim cais tawm ntawm lwm pawg phenolic. Raws li pom nyob rau hauv daim duab 1A, NDES #3 tau xaiv los rho catechin thiab epicatechin los ntawm cov tawv txiv hmab txiv ntoo. Qhov no yog qhov kev soj ntsuam nthuav dav vim tias NDES # 3 yog qhov tsawg kawg nkaus NDES kom rho tawm cov proanthocyanidins, uas yog oligomers thiab polymers ntawm catechin thiab epicatechin [14]. Qhov no tau qhia tias NDES #3 tuaj yeem xaiv rau proanthocyanidins ntawm qhov me me molecular. Kev sib koom ua ke ntawm phenolic tebchaw rau ntawm qhov chaw thauj khoom ntawm daim tawv nqaij (Daim duab 1B) txawv ntawm cov noob (Daim duab 1D) tsis hais qhov qis yields ntawm cov tebchaw hauv cov noob txiv hmab txiv ntoo. Thawj thiab thib ob lub hauv paus tseem ceeb tau piav qhia txog 73 feem pua ntawm qhov sib txawv ntawm cov ntaub ntawv noob. Quercetin, myricetin, thiab ferulic acid tau muab rho tawm tau zoo dua los ntawm NDES #6, #7, thiab 75 feem pua ethanol, raws li qhia hauv cov qhab nia (Fig. 1C). Ellagic acid thiab gallic acid tau muab rho tawm zoo dua los ntawm NDES #9. Ib zaug ntxiv, catechin tau muab rho tawm nrog qhov ua tau zoo tshaj plaws los ntawm NDES # 3 uas zoo ib yam li tau pom nrog cov tawv txiv hmab txiv ntoo. Epicatechin tau muab rho tawm nrog kev ua haujlwm siab dua los ntawm NDES #5, #4, thiab NDES #8.

3.4. Extraction optimization ntawm phenolic acids thiab flavonols los ntawm muscadine txiv hmab txiv ntoo tawv thiab ANN kev twv ua qauv qauv Choline chloride: levulinic acid: ethylene glycol 1: 1: 2 (NDES # 1) pom qhov siab tshaj plaws extraction tawm los rau ellagic acid, thiab yog li ntawd tau xaiv rau ntxiv optimization thiab kwv yees. Kev cuam tshuam ntawm plaub yam, suav nrog cov ntsiab lus dej, lub sijhawm ultrasonication, qhov sib piv ntawm cov khoom sib npaug rau cov kuab tshuaj, thiab qhov kub thiab txias tau raug ntsuas rau kev rho tawm ntawm phenolic acids thiab flavonols. Tsis tas li ntawd, plaub theem rau txhua qhov kev rho tawm tau siv rau hauv tag nrho ntawm 40 randomized khiav. Kev sim extraction tawm los ntawm ellagic acid, gallic acid, ferulic acid, myricetin, thiab quercetin, nrog rau cov sum ntawm cov tsib no, yog qhia nyob rau hauv Table 4. Zuag qhia tag nrho, qhov ntau yam ntawm extraction tawm los txawv ntawm qhov qis tshaj thiab siab tshaj yog kuj loj. rau phenolic acids. Piv txwv li, qhov qis tshaj tawm rau ellagic acid yog 9. (Nkauj #17). Ntxiv mus, cov txiaj ntsig qis tshaj plaws yog 20.7 mg / g, thiab qhov siab tshaj plaws yog 71.5 mg / g. Qhov no qhia txog qhov cuam tshuam tseem ceeb ntawm cov qib sib txawv ntawm txhua qhov kev rho tawm ntawm kev rho tawm tawm los. Khiav #15 rho tawm qhov siab tshaj plaws ntawm ellagic acid. Qhov kev rho tawm ntawm kev khiav #15 yog 45 mL /10{{130}} mL dej ntsiab lus, 25 min ntawm ultrasonication, 1:10 (g: mL) solid-to-solvent ratio , thiab rho tawm kub ntawm 60 ◦C. Daim duab S3 qhia txog HPLC chromatogram ntawm optimized phenolic acids muab rho tawm ntawm daim tawv nqaij txiv hmab txiv ntoo los ntawm NDES #1 (khiav # 15 hauv Table 4). Qhov siab tshaj plaws gallic acid (18.7 mg / g) tau ua tiav siv cov txheej txheem rho tawm hauv kev khiav # 24 thiab qis tshaj yog 6.63 mg / g siv khiav # 40. Rau ferulic acid, khiav #22 rho tawm qhov siab tshaj plaws ntawm 19.2 mg / g, thaum tsis muaj ferulic acid tau kuaj pom hauv # 14, # 17, # 29, thiab # 34. Khiav #22 tau muab rho tawm nrog 60 mL / 100 mL dej cov ntsiab lus, 5 min ntawm ultrasonication, 1: 5 rau cov khoom-rau-kua sib piv, thiab rho tawm kub ntawm 60 ◦C. Khiav # 2 rho tawm qhov siab tshaj plaws myricetin (10.1 mg / g) thiab quercetin (1.87 mg / g). Lub extraction tej yam kev mob ntawm khiav #2 yog 60 mL / 100 mL dej ntsiab lus, 35 min ntawm ultrasonication, 1:20 rau solid-to solvent ratio, thiab extraction kub ntawm 60 ◦C. Qhov qis tshaj kuv cov txiaj ntsig (3.79 mg / g) tau muab rho tawm los ntawm kev khiav # 40. Cov phiaj xwm contour hauv daim duab 2 qhia txog cov txiaj ntsig ntawm kev rho tawm tsis tau (X1, X2, X3, thiab X4) ntawm qhov kwv yees tawm los ntawm ellagic acid muab rho tawm los ntawm NDES #1 los ntawm daim tawv nqaij. Kev kwv yees tawm los ntawm ellagic acid hauv Table 4 tau siv los tsim cov count plots. Txhua lub vaj huam sib luag qhia txog qhov cuam tshuam ntawm 2 qhov kev rho tawm tsis tau. Cov kab contour tau sau nrog cov txiaj ntsig ntawm ellagic acid (mg / g). Qhov zoo tshaj plaws kwv yees dej cov ntsiab lus yog kwv yees li 35–45 mL / 100 mL NDES, raws li qhia hauv daim duab 2B thiab 2C. Ntev ultrasonication lub sij hawm nce yield ntawm ellagic acid (Fig. 2D thiab 2E), qhia lub luag hauj lwm tseem ceeb ntawm sonication nyob rau hauv NDES extraction. Thaum lub sij hawm rho tawm, sib tov cov tawv nqaij los yog cov noob nrog NDES qhia cov khoom thiab cov pa roj, uas ntxiv cov chaw acoustic cavitation rau ultrasounds los tsim cov npuas me me hauv NDES. Lub imploding ntawm cov npuas no ua rau huab cua kub, siab sib txawv, siab shear quab yuam, macro-turbulences, thiab micro-mixing, uas zoo agitated NDES kom ceev huab hwm coj diffusion thiab hloov. Thaum cavitation npuas imploded rau saum npoo ntawm txiv hmab txiv ntoo los yog daim tawv nqaij hais, cov tshwm sim micro-jets thiab inter-particle sib tsoo ua rau nto tev, yaig, particle tawg, sonoporation, thiab cell cuam tshuam [20]. Tag nrho cov kev cuam tshuam ntawm kev ua haujlwm ntawm ultrasound-induced cavitation ua rau muaj kev nkag mus ntawm NDEs mus rau hauv lub cell sab hauv kom intercellular phenolics los ntawm cov zaub mov matrix tau pauv mus rau hauv cov kuab tshuaj. Qhov zoo tshaj plaws khoom-rau-kib piv yog 1:10, raws li qhia los ntawm Fig. 2B, 2D, thiab 2F. Thaum kawg, kev rho tawm ntau dua qhov kub txog li 60 ◦C zoo li muaj txiaj ntsig zoo ntawm ellagic acid extractability raws li qhia hauv daim duab 2C, 2E, thiab 2F. Qhov no qhia txog kev sib raug zoo ntawm kev rho tawm qhov kub thiab qhov tawm los ntawm ellagic acid muab rho tawm los ntawm daim tawv nqaij. Ellagic acid extraction yields (Table 4) tau soj ntsuam rau kev kwv yees ua qauv siv kev sib txuas lus neural. Cov ntaub ntawv sim tau muab faib ua ib qho kev cob qhia thiab cov txheej txheem validation. Yog vim li cas thiaj suav nrog kev siv tau los ntawm cov software txheeb cais yog txhawm rau txhawm rau overfitting. Txhawm rau kwv yees ellagic acid yield (Y), tib yam plaub yam kev ywj pheej rho tawm (X1, X2, X3, thiab X4), 1-2 cov khaubncaws sab nraud povtseg nrog ntau tus naj npawb ntawm cov neurons, thiab peb qhov kev ua haujlwm tau raug soj ntsuam. Cov haujlwm ua kom muaj zog yog hyperbolic tangent, linear, thiab gaussian. Tom qab ntawd, cov ntaub ntawv tau raug cob qhia kom txog rau thaum tus nqi R-squared siab rau ob qho tib si kev cob qhia thiab kev siv tau. Cov ntaub ntawv twv ua ntej thiab tus qauv tau tsim. Cov qauv ANN zoo tshaj plaws tau xaiv los ntawm kev txheeb xyuas plaub qhov kev nkag (X1, X2, X3, thiab X4) nrog ib txheej zais uas siv Gaussian muaj nuj nqi nrog kaum neurons (Daim duab S5). R-squared ntawm kev cob qhia thiab kev siv tau teeb tsa yog 0.99, qhov RASE thiab AAE ntawm tus qauv yog 0.062 thiab 0.044, feem. Qhov R-squared ntawm ANN validation ntawm ellagic acid nyob rau hauv txoj kev tshawb no (0.99) yog siab tshaj ANN validation ntawm procyanidins (0.95) thiab anthocyanins (0.91) nyob rau hauv ib tug yav dhau los txoj kev tshawb no [14]. Txawm li cas los xij, qhov kev nce hauv R2 no tuaj yeem raug ntaus nqi rau cov qauv tsim kom zoo dua ntawm cov ntaub ntawv hauv txoj kev tshawb no, uas tuaj yeem yog vim qhov kev sim me me. Cov qauv kev kwv yees ANN rau kev rho tawm ntawm ellagic acid siv NDES #1 tau qhia raws li qhov sib npaug 3-13:

Improve immunity

Cistanche rau kev txhim kho kev tiv thaiv

Xaus

Cov kev tshawb pom tam sim no tau nthuav tawm cov pov thawj ntxiv ntawm kev ua tau zoo ntawm NDES lub peev xwm los rho cov polyphenols los ntawm kev lag luam khoom noj khoom haus los ntawm cov khoom lag luam. Cov txiaj ntsig tau txhawb nqa qhov kev xav ntawm qhov zoo tshaj plaws ultrasound-pab extraction ntawm NDES tshaj 75 feem pua ethanol. NDES tau zoo tshem tawm peb phenolic acids, ob flavononls, thiab peb flavan-3-ols los ntawm cov tawv nqaij thiab cov noob. NDES #1 yog qhov ua tau zoo tshaj plaws NDES kom rho tawm ellagic acid, thaum NDES #3 yog qhov tshwj xeeb xaiv rau kev rho tawm catechin thiab epicatechin. Ib qho tsis zoo ntawm NDES yog lawv cov viscosity siab, uas nthuav tawm cov teeb meem thaum tuav thiab rov qab los. Hauv txoj kev tshawb fawb tam sim no, kev sib txuas ntawm cov neural sib txuas, tsis hais txog nws qhov kev txwv qhov tshwm sim, tau ua kom pom qhov ua tau zoo rau kev kwv yees ua qauv. NDEs yog cov xov xwm muaj zog kom rov qab tau phytochemicals los ntawm cov khoom noj. Qee NDEs kuj nthuav tawm cov kuab tshuaj lom tsawg dua los kawm cov phytochemicals hauv cov hlwb nyob [21,22]. Thaum kawg, ntuj sib sib zog nqus eutectic cov kuab tshuaj yog qhov zoo tshaj plaws extraction media rau cov kuab tshuaj organic.

improve-immunity

Cov ntaub ntawv

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[2] M. Cvjetko Bubalo, N. ´ Curko, M. Tomaˇsevi´c, K. Kovaˇcevi´c Ghani's, I. Radojˇci´c Redovnikovi´c, Ntsuab extraction ntawm grape tawv nqaij phenolics los ntawm kev siv sib sib zog nqus eutectic solvents, Khoom noj khoom haus Chem. 200 (2016) 159–166, https://doi.org/10.1016/j. khoom.2016.01.040.
[3] M. Panic, V. Gunjev'c, G. Cravotto, I. Radojˇci´c Redovnikovi´c, Ua kom cov thev naus laus zis rau kev rho tawm cov txiv hmab txiv ntoo-pomace anthocyanins siv cov kuab tshuaj sib sib zog nqus eutectic hauv li-rau-ib nrab-liter batch Extraction ntawm grape-pomace anthocyanins siv NADES, Khoom noj khoom haus Chem. 300 (2019) 125185, https://doi.org/ 10.1016/j.foodchem.2019.125185.
[4] M. Hoffmann, et al., Muscadine Grape Production Guide for the Southeast, North Carolina State University, NC State Extension Publications, 2020 https://content. ces.ncsu.edu/muscadine-grape-production-guide (mus txog 18 Lub Ib Hlis 2021). [5] Cline, B. thiab C. Fisk, Txheej txheem cej luam ntawm muscadine txiv hmab txiv ntoo acreage, cultivars thiab ntau lawm cheeb tsam nyob rau hauv sab hnub tuaj US. Muscadine Grape Rhiav rau Cooperative Extension Agents, nyob rau yav qab teb cheeb tsam me txiv hmab txiv ntoo Consortium. 2006: Southern Region Small Fruit Consortium.
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[7] P. Greenspan, thiab al., Antiinflammatory zog ntawm muscadine txiv hmab (Vitis rotundifolia), J. Agric. Khoom noj khoom haus. Chem. 53 (22) (2005) 8481–8484, https://doi.org/ 10.1021/jf058015.
[8] DN Ignacio, KD Mason, EC Hackett-Morton, C. Albanese, L. Ringer, WD Wagner, PC Wang, MA Carducci, SK Kachhap, CJ Paller, J. Mendonca, L. Li Ying Chan, Bo Lin, DK Hartle, JE Green, CA Brown, TS Hudson, Muscadine grape skin extract inhibits prostate cancer cells by inducing cell-cycle capture and txo migration through heat shock protein 40, Heliyon 5 (1) (2019) e01128, https://doi .org/10.1016/j.heliyon.2019.e01128.