Elastin tsim cov Elastic Protein ntawm Epidermal Connective Tissues nrog Collagen

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

Abstract:Ib tug tshiab xanthone glycoside,135,6-tetrahydroxyxanthone-C-4-D-glucopyranoside raug cais tawm los ntawm methanol extract ntawm Mangifera Indica nplooj (Anacardiaceae) loj hlob hauv Egypt. Cov qauv tau qhia meej los ntawm 1D thiab 2D-NMR spectroscopic cov ntaub ntawv. Lub physicochemical zog ntawm cov compound xws li lipophilicity, solubility, thiab formulation txiav txim siab tau kwv yees nyob rau hauv silico ADMET txheej txheem siv SwissADME server. Cov txheej txheem no tau muab Lipinski txoj cai ntawm tsib, xws li GIT nqus, faib, metabolism, thiab daim tawv nqaij permeation. Cov kev ua hauv vitro inhibitory tiv thaiv kev laus-mediated enzymes xws li collagenase, elastase, hyaluronidase, thiab tyrosinase raug soj ntsuam. Cov khoom sib xyaw tau nthuav tawm cov txiaj ntsig zoo tshaj plaws los tiv thaiv collagenase, tiv thaiv elastase, tiv thaiv hyaluronidase, thiab tiv thaiv tyrosinase cuam tshuam nrog IC50 qhov tseem ceeb ntawm 1.06,419.10,1.65, thiab 0.48 ug / mL, feem, piv rau kev tswj tau zoo. Lub compound qhia tau hais tias muaj kev cia siab tias cov dej solubility thiab tsim nyog ntawm daim tawv nqaij nkag mus rau hauv lub tswv yim hais txog qhov tsim nyog ntawm cov khoom xyaw rau cov tshuaj pleev ib ce raws li kev tiv thaiv kev laus rau kev npaj tshuaj pleev ib ce.

Ntsiab lus:xanthone; Mangifera indica; kev laus; ceollagenase; elastase; tyrosinase; hyaluronidase; hauv silico ADMET

1. Taw qhia

Cov txheej txheem kev laus yog cov txheej txheem biochemical nyuaj cuam tshuam nrog oxidative kev nyuaj siab uas tau tsav los ntawm endogenous oxygen thiab nitrogen dawb radicals generated thaum lub neej expectancy, uas tej zaum yuav ua rau muaj kev loj hlob ntawm cov hnub nyoog-kev tshwm sim [1]. Cov tawv nqaij laus pathologies xws li wrinkles, hyperpigmentation, laus me ntsis, wrinkles, melasma, freckles, lentigo, ephelides, nevus, browning, thiab melanoma yog ua los ntawm activation ntawm reactive oxygen hom (ROS) [2]. Dawb radicals los yog ROS tuaj yeem ua rau muaj kev hloov pauv hauv cov qauv ntawm cov tawv nqaij ntawm tes thiab ua rau cov cell membranes puas tsuaj los ntawm kev txhawb nqa cov oxidation ntawm lipids thiab cov proteins, ua rau DNA puas thiab cell tuag [3,4]. Ntxiv mus, ROS ua lub luag haujlwm tseem ceeb hauv cov txheej txheem kev laus ntawm daim tawv nqaij los ntawm kev puas tsuaj rau cov tawv nqaij loj xws li collagen thiab elastin los ntawm kev ua kom cov collagenase thiab elastase enzymes. Tsis tas li ntawd, cov dawb radicals ua rau hyaluronic acid degradation los ntawm kev ua kom hyaluronidases, uas ua rau tsis zoo hydration ntawm daim tawv nqaij [5,6]. Kev cia siab ntawm cov txheej txheem dynamic no yog suav tias yog qhov teeb meem tseem ceeb rau thaj chaw dermo-sau tshuaj pleev ib ce thiab kev tshawb fawb kev ua haujlwm tseem ceeb tau raug siv los nrhiav cov qauv tiv thaiv tshiab[7]. Antioxidants yog lub ntuj tiv thaiv enzymes tam sim no nyob rau hauv daim tawv nqaij thiab tiv thaiv cov dawb radicals ntau heev uas tsim nyob rau hauv lub cev. Txawm li cas los xij, oxidative tshuav nyiaj li cas yuav cuam tshuam los ntawm ntau yam xws li kev noj zaub mov thiab huab cua phem uas ua rau oxidative kev nyuaj siab. Nyob rau hauv no oxidative kev nyuaj siab, lub load ntawm dawb radicals nyob rau hauv lub cev yog ho siab tshaj tej yam ntuj tso antioxidants. Yog li ntawd, nws yog ib qho tseem ceeb heev uas yuav tsum tau siv cov tshuaj tiv thaiv kab mob sab nraud los ntawm qhov chaw xws li kev noj zaub mov, tshuaj pleev ib ce, los yog tshuaj los tiv thaiv cov dawb radicals thiab tiv thaiv kev laus ntawm daim tawv nqaij [5,8]. Cov kev tshawb fawb niaj hnub no tau qhia tias ob peb lub cev metabolites, feem ntau yog flavonoids thiab polyphenolics pab txhawb rau tag nrho cov tshuaj tiv thaiv antioxidant ntawm ntau cov nroj tsuag [9]. Qhov no tau ua rau muaj kev txaus siab rau kev tshawb nrhiav cov tshuaj antioxidant ntawm ntau cov nroj tsuag [10]. Cov txiv hmab txiv ntoo tau tshaj tawm rau lawv cov nplua nuj nyob hauv cov phenolics thiab flavonoids nrog rau cov vitamins [11].

Thov nias ntawm no kom paub ntxiv

Elastin tsim cov protein elastic ntawm cov ntaub so ntswg sib txuas nrog collagen thiab sawv cev rau lub luag haujlwm tseem ceeb hauv kev tiv thaiv ntawm daim tawv nqaij wrinkles thiab firmness [12]. Thaum lub sij hawm cell aging, matrix metalloproteinases xws li collagenase, elastase, thiab hyaluronidase yog upregulated [13]. Kev tawm tsam ntawm cov enzymes no yog ib txoj hauv kev kho mob zoo tshaj plaws los tswj kev ua kom tsis zoo ntawm daim tawv nqaij thaum laus [14]. Cov nroj tsuag-derived phenolic tebchaw tau piav raws li muaj zog whitening mediators vim lawv cov tyrosinase inhibitory zog [15] thiab qee qhov (ntawm lawv tuaj yeem ua lub luag haujlwm hauv matrix remodeling ntawm elastase inhibitory nyhuv [16].

Mangifera indica L. yog ib qho tseem ceeb tshaj plaws edible nroj tsuag uas belongs rau tsev neeg Anacardiaceae faib nyob rau hauv ntau lub teb chaws, tshwj xeeb tshaj yog nyob rau hauv cheeb tsam sov [17]. Mango txiv hmab txiv ntoo tau nthuav tawm lub luag haujlwm tseem ceeb hauv kev ua liaj ua teb, zaub mov, tshuaj, thiab nutraceutical teb rau ntau tshaj 4000 xyoo [18]. Ntxiv mus, txiv nkhaus taw txiv hmab txiv ntoo sawv nyob rau hauv 2nd txoj hauj lwm ntawm cov industrial cov qoob loo, tom qab balana, nyob rau hauv cov nqe lus ntawm ntau lawm thiab kev pab them nqi [19]. Ntau qhov chaw ntawm Mangifera indica sawv cev rau cov khoom muaj txiaj ntsig ntawm cov phytoconstituents xws li flavonoids, xanthonoids, phenolic acids, thiab triterpenoids uas muaj peev xwm ua tau raws li kev ua haujlwm molecules [18]. Tsis tas li ntawd, txiv hmab txiv ntoo muab tib neeg lub cev nrog cov khoom tseem ceeb xws li carbohydrates, proteins, rog, minerals, vitamins, tseem ceeb amino acids, carotenoids, noj fibers, thiab phenolics [20]. Kev lig kev cai, cov kab lis kev cai sib txawv tau tshaj tawm tias cov nplooj infusion tau siv rau ntau yam mob xws li raws plab, ntshav dysentery, anemia, hawb pob, mob ntsws, kub siab, insomnia, rheumatism, plab zom mov, kab mob siab, kab mob tetanus, nchuav menyuam, thiab ntshav [21]. Tsis tas li ntawd, hlawv nplooj 'fumes tau nqus rau hiccups thiab caj pas [18]. Cov tawv ntoo tau siv los ua diuretic thiab hauv kev hnav khaub ncaws antirheumatic. Cov noob yog paub zoo rau kev kho mob khaub thuas, mob hawb pob, raws plab, thiab los ntshav. Antioxidant kev ua haujlwm, siab tag nrho phenolic (TPC), tag nrho cov ntsiab lus flavonoid (TFC), 2, 2-diphenyl-1-picrylhydrazyl (DPPH) scavenging kev ua, thiab linoleic inhibition muaj peev xwm ntawm Mangifera indica cog qhov chaw tau raug tshaj tawm [ 22] ib.

Cistanche tuaj yeem tiv thaiv kev laus

Txoj kev tshawb no tau tsim los tshawb xyuas cov khoom hauv vitro inhibitory ntawm xanthone tshiab tau los ntawm 70 feem pua ​​​​methanolic extract ntawm Mangifera indica L.leaves tawm tsam collagenase, elastase, tyrosinase, thiab hyaluronidase. Tsis tas li ntawd, pharmacokinetic zog xws li nqus, faib, metabolism, thiab toxicity (hauv silico ADMET kev kwv yees) tau siv SwissADME thiab Lipinski txoj cai ntawm tsib los ntsuas cov tshuaj solubility, permeability, thiab formulation tsim nyog.

2.Qhov tshwm sim thiab kev sib tham

2.1. Structure Elucidation ntawm Isolated Compound

Cov compound tau txais los ua cov muaju daj (18.29mg). Kev tshuaj ntsuam chromatographic tau qhia txog qhov daj-txiv kab ntxwv-daj ntawm TLC nyob rau hauv lub teeb UV ntev (365 nm) hauv DCM: MeOH (7: 3) thiab BAW hnyav tshuab. Thaum qhov chaw tau kho nrog 1 feem pua ​​​​methanolic ferric chloride tov, nws tig ntsuab tsaus thiab muab daj nrog tshuaj tsuag ammonia. Lub lH-NMR, APT, thiab HMBC spectral cov ntaub ntawv ntawm lub compound yog muab tso ua ke nyob rau hauv Table 1. lH-NMR spectral tsom xam (DMSO-dg, chav tsev kub) (Figures Sl thiab S2) qhia lub xub ntiag ntawm ob downfield aromatic doublet signals ntawm oH 7.57 thiab 6.79 nrog qhov sib txuas tas li ntawm 8.24 Hz qhia tias muaj ob lub ortho-coupled protons. Tsis tas li ntawd, thaj tsam aromatic ntawm H-NMR spectrum tau pom tias muaj lub teeb liab singlet ntawm Su 5.96, qhia tias muaj ib qho proton hauv lub nplhaib uas muaj ntxhiab. Lub anomeric proton ntawm C-glucosyl chav tsev tau kuaj pom ntawm Su 4.60 ppm nrog qhov sib txuas tas li ntawm 7 Hz muab rau -anomer. Cov qauv no ntawm cov ntxhiab tsw protons hauv H-NMR spectrum ntawm cov khoom sib cais zoo ib yam li cov qauv xanthone. Tsis tas li ntawd, lub xub ntiag ntawm -anomeric proton ntawm ou 4.60 nrog cov tshuaj lom neeg hloov pauv ntawm cov piam thaj ntawm 3.21 txog 4.60 ntawm lH-NMR (qab zib thaj tsam) qhia tias muaj C-glycosidic txuas ntawm qab zib thiab xanthone. Raws li pom nyob rau hauv daim duab S3, APT spectrum ntawm lub compound qhia lub xub ntiag ntawm kaum cuaj carbon signals, sawv cev rau lub xanthone thiab cov qauv C-glucoside.cistanche tubulosa tshuaj xyuasQhov kev sim APT tau txheeb xyuas peb CH pawg (C-8},131.95; C-7,115.10;C-2, 95.35 ppm) hauv xanthone moiety thiab anomeric C-1' C-glucose ntawm SH75.13 ppm. Peb tau ua ob qhov kev sim NMR (HMBC; Daim duab S4) kom paub meej tias qhov sib txuas glycosidic ntawm C-glucose ntawm C-4 ntawm xanthone. Muaj ntau 2,3JCH log-ntau sib txheeb cov cim qhia tau hais txog xanthone qaum nrog pom hla-peaks los ntawm H-1'(6) 4.60) rau C-4(6c 104.13 ppm) thiab C-3 (6c 158.95).Citrus bioflavonoidsRau cov xanthone moiety, 2,3JCH qhov sib txuas ntev kuj tau kuaj pom los ntawm H-2(8) rau C-4(6c 104.13 ppm), C-3 (8c 158.95), thiab C-4b(6c 107.45).Tsis tas li ntawd, muaj ib qho 2-37-ntev kev sib raug zoo los ntawm H-8(8)7.57) rau carbonyl pawg C-9( 6c 195.14), C-8a(6c 131.22), thiab C-6 (6c161.87).Rau tib lub nplhaib, aJcH teeb liab los ntawm H-7 (6μ 6.79) rau C{ {39}}a(6c 131.22) tau pom. Cov kev sib raug zoo no tau piav qhia nyob rau hauv cov qauv ntawm cov khoom sib xyaw hauv daim duab 1. HSQC ntawm qhov sib xyaw pom 1JcH kev sib raug zoo ntawm txhua tus proton thiab nws cov pa roj carbon monoxide raws li qhia hauv daim duab S5. Qhov kev sim H,H COZY (Daim duab S6) tau qhia txog kev sib raug zoo ntawm H-7 (δu 6.79) thiab H-8 (8H7.57). Tsis tas li ntawd, qhov sib xyaw ua ke tau nthuav tawm cov pseudomolecular ion ncov [MH]- ntawm m / z 421, qhia txog cov mis molecular ntawm CgHngOrin hauv hom tsis zoo ntawm ESI-MS tsom xam (Daim duab S7). Cov ntaub ntawv los ntawm 1H-NMR, APT, IH, H COSY, HMBC, thiab HSQC coj peb mus postulate tias cov compound yuav yog 1,3,5,6-tetrahydroxyxanthone-C-4- -D-glucopyranoside. Qhov no tau lees paub los ntawm kev sib piv nrog NMR cov ntaub ntawv qhia ntawm isomangiferin hauv cov ntaub ntawv [23,24].

2.2.In Silico Pharmacokinetics Prediction ntawm Isolated Compound

SwissADME online neeg rau zaub mov tau siv los tshuaj xyuas cov peev xwm ntawm cov tshuaj sib cais los ntawm kev kwv yees Lipinski txoj cai ntawm tsib (RO5) rau kev siv tshuaj zoo li [25]. Lipinski tau tshaj tawm tias 90 feem pua ​​​​ntawm cov tshuaj siv qhov ncauj uas tau mus txog theem II kev kho mob muaj MWT Tsawg dua lossis sib npaug li 500, logp Tsawg dua lossis sib npaug li 5, H-cov nyiaj pub dawb tsawg dua lossis sib npaug li 5, thiab H-daim ntawv cog lus lees txais tsawg dua. los yog sib npaug rau 10[26].SwissADME kwv yees ib qho ntxiv rau physicochemical tsis cuam tshuam nrog cov tshuaj zoo li lipophilicity [27], qhov loj, polarity [28], solubility [29,30], yooj, thiab saturation [31,32] raws li tau piav qhia hauv daim duab 2 thiab Table 2.cistanche UKLub SWISSADME zaj duab xis ntawm cov tshuaj zoo li ntawm cov khoom sib cais tau pom tias feem ntau ntawm cov khoom siv physicochemical ntawm lub compound yog nyob rau hauv qhov xav tau ntau yam [33] tshwj tsis yog rau cov naj npawb ntawm H-bond txais, H-bond pub, thiab polarity ntawm lub compound txij thaum PSA tus nqi yog 201.28 A2 (qhov xav tau ntau yam (nruab nrab ntawm 20 thiab 130 A2)[28]. Qhov siab polarity no yuav raug ntaus nqi vim muaj cov piam thaj moiety. Cov khoom sib cais tau pom tau tias muaj kev cia siab rau kev kwv yees topological aqueous solubility log S(Ali)[29 Qhov no yuav pab txhawb kev tsim kho yav tom ntej ntawm cov tshuaj no, kev kwv yees pharmacokinetics tau pom tias qhov sib xyaw ua ke tau kwv yees GIT nqus qis uas tsis muaj permeation rau BBB raws li tus qauv Dania boiled qe [34] tau txais los ntawm SwissADME online server[33].cistanche wrkungTsis tas li ntawd, qhov sib xyaw ua ke tsis pom muaj qhov cuam tshuam rau tsib isoforms ntawm cytochrome P450 (1A2,2C19,2C9,2D6, thiab 3A4) [35], qhia tias muaj kev cuam tshuam tsawg ntawm kev siv tshuaj yeeb-tshuaj. Kev kwv yees ntawm daim tawv nqaij permeation yog xam los ntawm ntau hom kab rov ua dua tshiab uas cuam tshuam cov molecular loj thiab lipophilicity nrog daim tawv nqaij permeability [36]. Qhov ntau qhov tsis zoo ntawm lub cav Kp (nrog Kp hauv cm / s), qhov tsawg ntawm daim tawv nqaij permeant yog cov molecule [33,36].cistanche tubulosa extractQhov kwv yees ntawm daim tawv nqaij permeation cav Kp hauv cm/s yog -9.14 cm/s.

2.3.Kev soj ntsuam ntawm Kev Tiv Thaiv Kev Laus Ntawm Cov tawv nqaij

2.3.1.Kev txiav txim siab ntawm Anti-Collagenase thiab Anti-Elastase Activities

Collagen thiab elastin yog cov txheej txheem tseem ceeb ntawm cov kab mob epidermis uas txhawb nqa lub elasticity, capillary tone, thiab lub zog ntawm shkin nrog hyaluronic acid [39]. Thaum lub sij hawm kev laus, oxidative kev nyuaj siab, thiab ntau heev raug UV teeb ua rau lub activation ntawm hydrolyzing enzymes xws li elastase, collagenase, thiab hyaluronidase, yog li daim tawv nqaij lub zog thiab yooj ploj, thiab wrinkles tshwm [40] . Qhov inhibition ntawm collagenase thiab elastase kev ua ub no yog ib qho ntawm cov tswv yim zoo los tiv thaiv daim tawv nqaij ntawm daim tawv nqaij laus tshwm sim [41]. Raws li tau hais meej hauv daim duab 3A, qhov sib xyaw ua ke tau pom cov cuab yeej tiv thaiv kab mob sib kis los ntawm ICso tus nqi ntawm 419.10 ug / mL, piv rau phenanthroline (IC50=182}.80ug / mL) raws li tus qauv. Hais txog cov nyhuv elastase inhibitory (Daim duab 3B), nws tau pom cov txiaj ntsig zoo kawg nkaus nrog tus nqi ICso ntawm 1.06 ug / mL, thaum muaj kev tswj hwm zoo, N-methoxysuccinyl-Ala-Pro-Val-chloromethyl ketone, exerted tus nqi ICso ntawm 0.63 ug. /mL.Ua raws li cov txiaj ntsig no, kev tshawb fawb ua ntej tau tshaj tawm tias methanol nplooj extract ntawm txiv nkhaus taw yog 10-fold muaj zog elastase inhibitor dua li tus qauv tocopherol ascribed rau cov khoom tsis sib tw inhibition ntawm mangiferin[42]. Tsis tas li ntawd, cov ntaub ntawv dhau los tau ua pov thawj qhov kev tiv thaiv elastase ntawm cov tshuaj polyphenolic vim muaj cov pab pawg hydrophilic xws li hydroxyl lossis carboxyl uas tuaj yeem cuam tshuam zoo rau kev sib tw inhibition ntawm enzymes [43].

2.3.2.Kev txiav txim siab ntawm Anti-Hyaluronidase thiab Antityrosinase Kev Ua Haujlwm

Hyaluronic acid yog qhov nthuav dav glycosaminoglycan ntawm daim tawv nqaij uas khaws nws cov ntsiab lus moist-ture [44]. Nws yog enzymatically hydrolyzed los ntawm hyaluronidase uas ua rau kev ua txhaum ntawm kev ncaj ncees ntawm daim tawv nqaij qauv thiab cuam tshuam cov ntaub so ntswg permeability [45]. Lub sup-pression ntawm hyaluronidase tswj cov tawv nqaij kev ncaj ncees, ncua kev loj hlob ntawm daim tawv nqaij laus, thiab khaws cov tawv nqaij hydration [13]. Cov kev tshawb fawb yav dhau los tau qhia tias ntuj hyaluronidase inhibitors tuaj yeem ua haujlwm zoo li cov tshuaj tiv thaiv kev laus rau cov khoom siv kho mob ntawm daim tawv nqaij txhim kho [46]. Raws li tau piav qhia hauv daim duab 4A, cov khoom sib cais tuaj yeem ua rau txo qis hyaluronidase kev ua haujlwm nrog ICso tus nqi ntawm 1.65 ug / mL, piv rau 6-O-palmitoyl L-ascorbic acid raws li kev tswj zoo (2.55 ug / mL).

Ntawm cov kev kho mob ntawm daim tawv nqaij senescence tshwm sim, daim tawv nqaij hyperpigmentation tuaj yeem ua rau ac-celerated los ntawm tyrosinase enzyme nyob rau hauv lub xub ntiag ntawm reactive oxygen hom (ROS) [46]. Tyrosinase, tooj liab-muaj glycoprotein, ua haujlwm tseem ceeb hauv melanin synthesis los ntawm hydroxylation ntawm L-tyrosine mus rau 3, 4-dihydroxyphenylalanine (DOPA), tom qab ntawd los ntawm oxidation ntawm DOPA rau DOPAquinone [47]. Yog li ntawd, kev tawm tsam ntawm tyrosinase kev ua haujlwm tuaj yeem txo cov tawv nqaij hyperpigmentation [48]. Los ntawm qhov kev pom no, cov compound poten-tially inhibited tyrosinase nrog tus nqi ICso ntawm 0.48 ug / mL, piv rau kojic acid raws li kev tswj zoo (0.82 ug / mL) raws li qhia hauv Daim duab 4B. Ib txoj kev tshawb fawb yav dhau los ntawm Mangifera indica noob extract tau pom zoo los tiv thaiv tyrosinase thiab anti-hyaluronidase cov txiaj ntsig cuam tshuam nrog cov ntsiab lus polyphenolic [47]. Ntawm qib molecular, ob qho tib si tyrosine thiab DOPA muaj cov pab pawg hydroxyl uas qhia tias yog qhov tseem ceeb proton pub rau tyrosinase ua kom [49]. Ib yam li ntawd, lub compound tuav plaub hydroxyl moieties nyob rau hauv nws lub cev pob txha uas muaj peev xwm coj li kev sib tw inhibitors los ntawm reacting li substrates rau tyrosinase.

3. Cov khoom siv thiab cov txheej txheem

3.1.Cov khoom siv cog

Cov nplooj tshiab ntawm Mangifera indica L.leaves tau txais ntawm cov txiv hmab txiv ntoo los ntawm ib lub vaj ntiag tug hauv cheeb tsam Abo-Zabal (N 30 degree 1743.5336"E 3128.254), Qualiobya tsoom fwv, Egypt, thaum 20 Lub Xya hli ntuj 2021. Lawv tau ua siab zoo lees paub los ntawm Mrs. Treize Labib, tus kws tshaj lij taxonomy ntawm El-Orman Botanical Garden, Giza, Tim lyiv teb chaws. Voucher cov qauv nrog tus lej PHG-P-MI-362 tau muab tso rau ntawm Pharmacognosy Department gallery, Kws qhia ntawv ntawm Pharmacy, Ain Shams University.

3.2.Extraction thiab Chromatographic Isolation

Cov nplooj ziab cua ntawm Mangifera indica L. (1.56 kg) tau percolated hauv 70 feem pua ​​methanol (27L) rau 5 hnub, ces lim. Cov lim tau tag nrho evaporated nyob rau hauv vacuo ntawm 47 degree kom qhuav kom qhuav residue (9{25}}.36 g; 5.79 feem pua ​​w/w); cov txiaj ntsig ntawm kev rho tawm tau suav raws li qhov sib npaug hauv qab no: [tag nrho qhov hnyav ntawm qhuav extract / tag nrho qhov hnyav ntawm cov nroj tsuag] × 100. Tom qab ntawd, cov extract tau muab siv rau ntawm Diaion HP-20 fractionation. Cov gradient elution (dej / methanol) tau siv. Tsib feem loj tau txais: 100 feem pua ​​​​dej, 25 feem pua ​​​​methanol, 50 feem pua ​​​​methanol, 75 feem pua ​​​​methanol, thiab 100 feem pua ​​​​methanol. 50 feem pua ​​​​methanol-soluble feem (26.38 g) tau raug polyamide thiab eluted pib nrog 100 feem pua ​​dej, ces gradient eluted nrog dej / methanol (los ntawm 100: 0 mus rau 0: 100, w / o) kom tau rau 6 subfractions: 100 feem pua ​​dej, 20 feem pua, 40 feem pua, 60 feem pua, 80 feem pua ​​thiab 100 feem pua ​​methanol. Cov 40 feem pua ​​​​methanol-soluble subfraction (2.36 g) tau siv rau ntawm Sephadex LH-20 siv MeOH (isocratic elution) Cov feem zoo sib xws tau ua ke thiab evaporated kom tau txais plaub lub ntsiab subfractions (A1-A4), Cov subfraction A3(0. daj amorphous hmoov.

3.3.Nuclear Magnetic Resonance (NMR) Spectrometer

Lub Bruker Ascend 400/R (Burker Avance Ⅲ, Fallanden, Switzerland) spectrometer tau siv nyob rau ntawm Lub Chaw Saib Xyuas Tshuaj, Kev Tshawb Fawb, thiab Kev Txhim Kho, Kws Qhia Tshuaj, Ain Shams University. 3.4.Mass Spectrometry

Lub Finnigan LCQ-DECA huab hwm coj spectrometer (San Jose, CA, USA) txuas nrog lub PDA ntes tau siv rau kev tshuaj xyuas huab hwm coj. Cov qauv tau yaj hauv HzO: MeOH raws li kev sib xyaw thiab txhaj ncaj qha rau hauv HPLC / ESI-MS system. Ob qho tib si tsis zoo thiab zoo ESI ionization ion hom tau siv nyob rau hauv cov xwm txheej hauv qab no: ziab thiab nebulizing roj, N2; capillary kub, 250 degree; Txau hluav taws xob, 4.48 kV; capillary voltage, 39.6V; tube lens voltage, 10.00 V; thiab puv scan hom hauv huab hwm coj ntau m/z 100-2000. ESI-MS kev tshuaj xyuas rau cov kev sib cais tau ua tiav ntawm Waters Xevo TQD huab hwm coj spectrometer nrog UPLC Acquity hom (Milford, CT, USA) ntawm Center for Drug Discovery, Research and Development, Kws Qhia Ntawv ntawm Pharmacy, Ain Shams University.

3.5.In-Silico Pharmacokinetics Prediction

Lipinski txoj cai ntawm tsib, physicochemical tsis xws li lipophilicity, solubility, thiab pharmacokinetic zog raws li GIT nqus, kev faib tawm, metabolism, thiab daim tawv nqaij perme-ation ntawm cov khoom sib cais tau ua los ntawm SwissADME online server [36]. 3.6. Kev soj ntsuam ntawm Anti-Skin Aging Properties

3.6.1.Kev txiav txim siab ntawm Anti-Collagenase Activity

Lub peev xwm inhibitory ntawm cov tshuaj sib cais tawm tsam collagenase kev ua haujlwm tau soj ntsuam siv cov khoom siv tshuaj fluorometric collagenase inhibitor (BioVision, catalog no. # K833-100) raws li txoj kev tau piav qhia yav dhau los [50]. Kev tswj xyuas yog (1,10)-phenanthroline. Cov tshuaj ntsuam xyuas thiab kev tswj xyuas tau npaj nyob rau hauv qhov ntau ntawm 1,10,100, thiab 1000 ug / mL rau kev tshuaj xyuas. Cov tshuaj ntsuam xyuas tau npaj rau hauv 96- lub phaj zoo nrog lub tiaj tiaj. Ua ntej, collagenase substrate tau yaj hauv collagenase assay buffer (CAB). Cov qauv rau kev soj ntsuam tau npaj los ntawm kev sib xyaw ua ke nrog collagenase thiab CAB. Kev npaj inhibitor tswj cov qauv tau ua los ntawm kev sib xyaw cov inhibitor ((1,10)-phenanthroline (80 mM) nrog cov enzyme diluted collagenase thiab CAB tsis. Enzyme tswj tau npaj los ntawm kev sib xyaw cov diluted collagenase nrog CAB.Lub CAB tsis yog siv los ua tus tswj keeb kwm yav dhau. Tag nrho cov qauv tau incubated nyob rau hauv chav tsev kub rau 15 min. Lub sijhawm no, cov tshuaj tiv thaiv sib xyaw tau npaj los ntawm kev sib xyaw collagenase nrog CAB. Nyob rau hauv cov kauj ruam nram qab no, cov tshuaj tiv thaiv sib tov sib xyaw kom huv si nrog cov qauv npaj. Cov fluorescence tau ntsuas tam sim ntawm 490 nm excitation wavelength thiab 520 nm emission wavelength siv lub microplate nyeem ntawv (FilterMax F5, Thermo Fisher). Kev ntsuas tau ua nyob rau hauv hom kinetic ntawm 37 degree rau 60 min. Cov qauv tau npaj rau hauv duplicate thiab cov nyhuv collagenase inhibitory ntawm cov tshuaj ntsuam xyuas tau suav nrog siv cov lej hauv qab no:

3.6.2.Kev txiav txim siab ntawm Anti-Elastase Activity

Kev ua haujlwm los tiv thaiv elastase ntawm qhov sib xyaw tau tshawb xyuas siv cov khoom siv EnzChek§Elastase Assay (E-12056) raws li txoj kev qhia yav dhau los [51]. Cov tshuaj ntsuam xyuas thiab kev tswj xyuas tau npaj rau hauv qhov ntau ntawm 0.1,10, thiab 100 ug / mL rau kev tshuaj xyuas. Cov khoom sib xyaw tau npaj rau hauv 96- cov phiaj zoo nrog cov hauv qab kom meej rau qhov kev ntsuam xyuas fluorometric. Luv luv, elastase enzyme daws, elastase substrate, thiab inhibitor tswj tau npaj raws li kev pom zoo. Tom qab ntawd diluted elastase tov tau ntxiv rau hauv qhov dej. Cov tshuaj ntsuam xyuas, inhibitor tswj, thiab enzyme tswj tau ntxiv rau cov dej tom qab. Cov qauv tau muab sib xyaw rau ntawm lub shaker thiab tom qab ntawd incubated rau 5 feeb ntawm 37 degree. Assay buffer tau tov nrog substrate los npaj cov tshuaj fluorometric cov tshuaj tiv thaiv, uas tau ntxiv thiab sib xyaw kom huv si nrog txhua tus qauv. Cov fluorescence tau ntsuas tam sim ntawm 400 nm excitation wavelength thiab 505 nm emission wavelength siv lub microplate nyeem ntawv (FilterMax F5, Thermo Fisher, Waltham, MA, USA).Qhov ntsuas tau ua hauv hom kinetic ntawm 37 degree rau 30min tiv thaiv los ntawm lub teeb. Tag nrho cov qauv kuaj tau raug npaj thiab soj ntsuam hauv qhov sib npaug thiab cov nyhuv elasstase inhibitory ntawm cov tshuaj ntsuam xyuas tau suav nrog siv cov lej hauv qab no:

qhov twg:

△RFU=kev hloov pauv hauv cov txheeb ze fluorescence units EC=enzyme tswj

3.6.3.Kev txiav txim siab ntawm Anti-Tyrosinase Activity

Cov kev ua haujlwm inhibitory ntawm cov roj tau sim tawm tsam collagenase siv Abcam@Tyrosinase Inhibitor Kev Tshuaj Ntsuam Xyuas Colorimetric Assay Kit (catalog # ab204715). Qhov kev ntsuam xyuas tau ua raws li phau ntawv qhia. Tus qauv tyrosinase inhibitor yog kojic acid. Cov tshuaj ntsuam xyuas tau npaj rau hauv cov concentration ntawm 0.1 txog 10 ug / mL rau kev tshuaj xyuas. Kwv yees li 2 l ntawm kev muab tyrosinase yog tov nrog 48 uL ntawm tyrosinase assay tsis. Cov qauv ntawm cov roj yam tseem ceeb los yog sawv ntsug (20 uL) tau tov nrog cov enzyme sib tov (50 μL) thiab incubated ntawm 25 degree rau 10 min. Tom qab ntawd, 30μL ntawm tyrosinase substrate tov tau ntxiv rau hauv qhov dej ntawm cov roj yam tseem ceeb thiab cov qauv. Tom qab ntawd, lub phaj tau ntsuas spectrophotometrically ntawm lub wavelength ntawm 510 nm rau 30-60 min ntawm 25 degree .

3.6.4.Kev txiav txim siab ntawm Anti-Hyaluronidase Kev Ua Haujlwm

Hyaluronidase kev ua ub no tau soj ntsuam spectrophotometrically los ntawm kev ntsuas tus nqi ntawm N-acetylglucosamine tsim los ntawm sodium hyaluronate [52]. Cov tshuaj ntsuam xyuas thiab kev tswj xyuas tau npaj nyob rau hauv qhov ntau ntawm 0.1 txog 10 ug/mL rau kev tshuaj xyuas. Tsib caug microliters ntawm bovine hyaluronidase (7900 units mL-1, (Sigma, St. Louis, MO, USA) yaj nyob rau hauv 0.1 M acetate buffer (pH=3.5) yog tov nrog 100 μL ntawm ib tug xaiv concentration ntawm cov compound yaj nyob rau hauv 5 feem pua ​​DMSO thiab ces incubated nyob rau hauv ib tug da dej ntawm 37 degree rau 20min. Cov pab pawg neeg tswj tau kho nrog 100 μL ntawm 5 feem pua ​​DMSO es tsis txhob ntawm cov qauv.Qhov optical ceev ntawm cov tshuaj tiv thaiv sib tov yog ntsuas ntawm 585 nm. los ntawm spectrophotometric.

3.7.Kev txheeb xyuas qhov tseeb

Txhua qhov kev sim tau ua hauv triplicate. Cov ntaub ntawv rau ICso ntawm enzyme nyob rau hauv-hibitory kev ua ub no tau soj ntsuam statistically siv ib txoj kev tsom xam ntawm variance (ANOVA). Kev txheeb cais tau ua tiav siv GraphPad Prism 5.0. 4. Cov lus xaus thiab cov kev xav yav tom ntej

Cov khoom lag luam los ntawm cov nroj tsuag yog nyob rau hauv kev xav tau siab hauv kev lag luam thoob ntiaj teb rau kev txhim kho ntawm cov neeg ua haujlwm tshiab rau kev tsim kho kom zoo nkauj[4]. Nyob rau hauv txoj kev tshawb no, ib tug tshiab xanthone compound, 1.3.5, 6-tetrahydroxyxanthone-C-4- -D-glucopyranoside, raug rho tawm los ntawm nplooj extract ntawm M. indica. Raws li kev soj ntsuam enzyme inhibitory, qhov sib xyaw no tuaj yeem ua rau pom zoo los tiv thaiv collagenase, anti-elastase, aanti-hyaluronidase, thiab anti-tyrosinase prop-erties, yog li conferring ib qho kev ua kom muaj kev cuam tshuam rau ntawm daim tawv nqaij aging-txog enzymes. Tsis tas li ntawd, ib qho hauv silico physicochemical parameter thiab ADME txoj kev tshawb fawb txhawb nqa nws kev koom ua ke rau hauv cov ntaub ntawv tshuaj pleev ib ce vim nws qhia tau hais tias muaj kev cog lus aqueous solubility thiab tsim nyog kwv yees cov tawv nqaij nkag mus. Yog li ntawd, qhov sib xyaw no tuaj yeem siv tau los ua ib qho txiaj ntsig zoo ntawm cov khoom siv bioactive rau nutraceutical thiab cosmeceutical formula-tions. Txawm li cas los xij, kev ntsuam xyuas toxicological thiab kev kho mob muaj peev xwm ua tau zoo.

Kab lus no yog muab rho tawm los ntawm Molecules 2022, 27, 2609. https://doi.org/10.3390/molecules27092609 https://www.mdpi.com/journal/molecules