Kev npaj ntawm Ferulic Acid- Phospholipid Complex los txhim kho Solubility, Dissolution, Thiab B16F10 Cellular Melanogenesis Inhibition Activity

Abstract

Keeb kwm

Ferulic acid(FA; 4-hydroxy-3-methoxy cinnamic acid) muaj nyob rau hauv ntau yam khoom noj, xws li nplej, mov, barley, oats, citrus txiv hmab txiv ntoo, thiab txiv lws suav [1]. FA tau pom tias muaj peev xwm tiv thaiv tawv nqaij tiv thaiv UV-induced oxidative stress [2]. Nws thim rov qab UVB-induced oxidative kev puas tsuaj nyob rau hauv nas ntawm daim tawv nqaij hlav los ntawm modulating cov kev qhia ntawm vascular endothelial kev loj hlob yam (VEGF), inducible nitric oxide synthase (iNOS), qog necrosis factor (TNF)- , thiab interleukin (IL) {{7} } [3]. Nws kuj hloov pauv qhov kev qhia ntawm mutated p53, Bcl-2, thiab Bax hauv UVB-induced nas tawv nqaij hlav [4]. Ntau qhov kev tshawb fawb tau tsim tias FA inhibits qhov kev qhia ntawm cytotoxic thiab o ntawm cov enzymes [5] thiab matrix metalloproteinases (MMPs), thiab attenuates degradation ntawm collagen fibers [6].

Raws li kev tshawb fawb cuam tshuam,cistancheyog ib hom tshuaj ntsuab uas hu ua "cov tshuaj ntsuab miracle uas ua rau lub neej ntev". Nws lub ntsiab tivthaiv yogcistanoside, uas muaj ntau yam teebmeem xws li antioxidant, anti-inflammatory, thiab kev tiv thaiv kev ua haujlwm nce qib. Lub mechanism ntawmcistanchethiabtawv nqaij dawbnyob rau hauv cov nyhuv antioxidant ntawm cistanche glycosides. Melanin nyob rau hauv tib neeg daim tawv nqaij yog tsim los ntawm oxidation ntawm tyrosine catalyzed los ntawm tyrosinase, thiab cov tshuaj tiv thaiv oxidation yuav tsum muaj kev koom tes ntawm oxygen, yog li cov pa-dawb radicals nyob rau hauv lub cev ua ib qho tseem ceeb cuam tshuam rau melanin ntau lawm.Cistanchemuaj cistanoside, uas yog ib qho antioxidant thiab tuaj yeem txo qhov tsim ntawm cov dawb radicals hauv lub cev, yog li inhibiting melanin ntau lawm.

Tsis tas li ntawd, cistanche kuj muaj cov haujlwm ntawm kev txhawb nqa collagen ntau lawm, uas tuaj yeem ua rau kom elasticity thiab luster ntawm daim tawv nqaij thiab pab kho cov tawv nqaij puas. Cistanche Phenylethanol Glycosides muaj qhov cuam tshuam tseem ceeb ntawm kev ua haujlwm ntawm tyrosinase, thiab cov nyhuv ntawm tyrosinase tau pom tias muaj kev sib tw thiab thim rov qab inhibition, uas tuaj yeem muab lub hauv paus kev tshawb fawb los tsim thiab siv cov khoom xyaw whitening hauv Cistanche. Yog li ntawd, cistanche muaj lub luag haujlwm tseem ceeb hauv kev ua kom tawv nqaij. Nws tuaj yeem cuam tshuam melanin ntau lawm kom txo cov xim thiab dullness; thiab txhawb collagen ntau lawm los txhim kho daim tawv nqaij elasticity thiab radiance. Vim muaj kev lees paub dav dav ntawm cov teebmeem ntawm cistanche, ntau cov khoom siv tawv nqaij dawb tau pib infuse cov khoom xyaw tshuaj ntsuab xws li Cistanche kom tau raws li cov neeg siv khoom xav tau, yog li nce kev lag luam tus nqi ntawm Cistanche hauvcov khoom siv tawv nqaij dawb. Hauv cov ntsiab lus, lub luag haujlwm ntawm cistanche hauvtawv nqaij dawbyog qhov tseem ceeb. Nws cov nyhuv antioxidant thiab collagen-tsim cov nyhuv tuaj yeem txo cov xim thiab dullness, txhim kho daim tawv nqaij elasticity thiab luster, thiab yog li ua tiav cov nyhuv whitening. Tsis tas li ntawd, daim ntawv thov dav ntawm Cistanche hauv cov khoom siv tawv nqaij ua kom pom tseeb tias nws lub luag haujlwm hauv kev lag luam tsis tuaj yeem kwv yees.

Nyem rau Cistanche Rau Whitening

Nug ntxiv:

david.deng@wecistanche.com WhatApp:86 13632399501

Phospholipid complexes yog siv dav hauv kev lag luam kws tshuaj. Lawv muaj qhov zoo permeability thiab kev nyab xeeb thiab tau txais kev saib xyuas ntxiv rau kev siv tshuaj pleev ib ce. Vim tias phospholipids yog biofunctional surfactants nrog cov khoom zoo solubilizing, lawv tuaj yeem siv los ua cov khoom siv rau cov tshuaj soluble tsawg [7], txhim kho transdermal permeation thiab tsub zuj zuj ntawm cov tshuaj pleev xim [8]. Transdermal permeation ntawm cov tshuaj cuam tshuam nrog kev sib cais, kev faib tawm, thiab kev tawm mus rau hauv daim tawv nqaij. Lub cev thiab tshuaj lom neeg, tshwj xeeb tshaj yog cov roj-dej faib coefficient ntawm cov tshuaj yuav tsum tau tswj hwm, cuam tshuam rau cov txheej txheem no [9].

Cov txheej txheem

Khoom siv

Cell kab lis kev cai

Kev npaj ntawm FA-PC siv cov kuab tshuaj evaporation

Kev tshuaj xyuas kom pom qhov zoo ntawm ferulic acid thiab phospholipids

FA thiab phospholipids ntawm molar ratios ntawm 2: 1, 1: 1, 1: 2, 1: 3, thiab 1: 4 tau ntxiv rau 100 mL puag ncig-hauv qab khob thiab yaj hauv anhydrous ethanol (FA, 2.0 mg / ml). Cov tshuaj sib tov tau nplawm tas li ntawm 40 degree rau 1 h, thiab tom qab ntawd cov tshuaj anhydrous ethanol raug tshem tawm los ntawm kev sib hloov evaporation. Cov qhuav FA-PC complexes tau muab tso rau hauv lub desiccator rau 24 teev.

Kev tshuaj ntsuam xyuas qhov ntsuas kub zoo rau kev npaj FA-PC

Kev tshuaj xyuas rau lub sijhawm pom zoo rau kev npaj FA-PC

Kev tshuaj xyuas kom pom FA concentration

Cov yam ntxwv ntawm FA-PC

Sib txawv scanning calorimetry (DSC)

Solubility thiab roj-dej faib coefficient

Solubility

Lub solubilities ntawm hmoov FA thiab FA-PC tau txiav txim los ntawm kev ntxiv ib qho piv txwv rau 10.0 mL [10] dej los yog n-octanol thiab ces co ntawm lub viav vias txaj rau 3 h ntawm 37 degree. Cov sib tov tau centrifuged ntawm 15, 000 rpm rau 10 min kom tshem tawm cov FA insoluble. Tom qab ntawd, cov supernatants tau lim los ntawm 0.45 µm daim nyias nyias. Tom qab ntawd, cov filtrates tau diluted tenfold nrog methanol, thiab FA cov ntsiab lus tau txiav txim siab siv lub UV spectrophotometer (UV-3150; Shimadzu; Nyiv).

Cov qauv (10 mL) ntawm FA thiab FA-PC hauv dej-saturated n-octanol tau npaj thiab shaken. n-octanol-saturated dej (10 mL) tau ntxiv rau txhua tus qauv, thiab cov kua miscible tau agitated rau 24 teev. Tom qab ntawd, cov qauv raug tso cai sawv ntsug rau txheej txheej. Qhov FA concentration hauv txhua theem tau txiav txim siab los ntawm UV spectrophotometric (UV-3150; Shimadzu; Nyiv). Kev tshuaj xyuas tau ua tiav hauv triplicate.

Hauv vitro diffusion

Kev tshawb fawb hauv vitro diffusion tau ua los ntawm kev siv Franz diffusion hlwb (TK-20A; Shanghai Xie Kai Cov Ntaub Ntawv Nyiaj Txiag Co., Ltd.; Tuam Tshoj). Tsis tas li ntawd, peb siv Strat-M® daim nyias nyias, uas yog cov khoom siv hluavtaws nrog cov yam ntxwv sib txawv uas cuam tshuam rau tib neeg cov tawv nqaij ntau dua li cov qauv tawv nqaij [11]. Cov membranes tau clamped ntawm tus pub thiab txais cov chav ntawm ntsug diffusion hlwb, thiab cov chav txais khoom tau ntim nrog phosphate-buffered saline (PBS; pH 7.4) kom solubilize FA lossis FA-PC thiab xyuas kom cov dej hauv lub dab dej.

Kev sib cais chromatographic tau ua tiav siv Agilent 1260LC series system (Agilent Technologies, Palo Alto, CA, USA) nruab nrog lub tshuab nqus tsev degasser, quaternary twj tso kua mis, autosampler, thermostatted column compartment, thiab diode array detection (DAD ). Agilent Technologies ChemStation software rau kua chromatography (LC; B.02.01) tau siv, thiab HPLC kev sib cais tau ua tiav siv Eclipse plus-C18 kem (4.6 × 250 mm, 5 μm). Kev kuaj pom wavelength yog 0.05 feem pua ​​​​acetic acid (A) thiab methanol (B) (40:60, v/v). Tus nqi ntws yog 1.0 mL / min. Qhov kub thiab txias tau teem rau ntawm 30 degree. Cov kev kho ntau ntxiv tau ua kom paub tseeb tias FA tso tawm ntawm txhua lub sijhawm. Tag nrho cov kev ntsuas tau ua nyob rau hauv triplicate, thiab feem pua ​​​​ntawm cov FA cumulative uas permeated los ntawm daim nyias nyias (feem pua ​​Q) tau npaj raws li lub sijhawm ua haujlwm.

Inhibition ntawm melanogenesis

B16F10 cell viability assay

Cell viability thiab cell proliferation raug soj ntsuam siv ib tug {{{0}}}(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) kev ntsuas [2]. B16F10 hlwb raug pretreated nrog 0.25, 0.5, 1.0, 2.0, thiab 4.0 mg/mL concentrations ntawm FA thiab FA–PC. Tom qab incubation rau 48 h, MTT tov (qhov kawg concentration: 5 mg / mL) tau ntxiv, thiab cov hlwb raug incubated rau 3 h ntawm 37 degree. Thaum kawg, qhov nqus ntawm txhua tus qauv tau ntsuas ntawm tus nyeem ntawv microplate ntawm 570 nm kom tau txais qhov feem pua ​​​​ntawm cov hlwb siv tau.

Kev ntsuas cov ntsiab lus melanin

Cov ntsiab lus melanin tau ntsuas raws li tau piav qhia yav dhau los [6] nrog qee qhov kev hloov kho. B16F10 melanoma hlwb tau noob (2 × 105 hlwb / zoo nyob rau hauv 3 mL ntawm nruab nrab) nyob rau hauv rau-zoo kab lis kev cai daim hlau thiab incubated ib hmos kom cov hlwb ua raws. Thaum kawg ntawm kev kho mob, cov hlwb raug ntxuav nrog PBS thiab lysed nrog 1 M NaOH muaj ing 10 feem pua ​​​​dimethyl sulfoxide (DMSO) rau 30 min ntawm 80 degree. Lub absorbance (optical ceev; OD) tau ntsuas ntawm 475 nm siv lub microplate nyeem ntawv. Cov ntsiab lus melanin tau suav nrog cov qauv hauv qab no:

Cov ntaub ntawv tsom xam

Qhov tseem ceeb ntawm qhov sib txawv ntawm qhov ntsuas qhov nruab nrab ntawm txhua pab pawg kho mob thiab pawg tswj hwm tau txiav txim siab siv Dunnett's t-test. P tus nqi<0.05 were considered statistically significant.

Cov txiaj ntsig thiab kev sib tham

Kev npaj zoo tshaj plaws ntawm FA-PC

Solubility thiab roj-dej faib coefficient ntawm FA-PC

Table 1 qhia txog kev solubility ntawm FA, PM, thiab FA-PC hauv dej thiab hauv n-octanol. FA-PC tau pom zoo solubility hauv dej thiab n-octanol (1.68 thiab 7.77 mg / mL, raws li), thiab solubility ntawm FA-PC hauv n-octanol tau pom tias muaj ntau dua li ntawm FA (1.34 mg / mL) .

Lipophilicity feem ntau yog ntsuas raws li qhov sib faib coefficient (P) ntawm ob theem immiscible. Nws yog feem ntau qhia raws li log P. Qhov pom tseeb muab faib coefficients ntawm FA thiab FA-PC hauv n-octanol / dej system tau txiav txim siab. Cov txiaj ntsig tau qhia tias log P siab dua rau FA–PC (1.21) dua li rau FA (0.99) thaum ntsuas pH 5.0. Lub cav nce me ntsis P tau cuam tshuam nrog kev txhim kho zoo n-octanol solubility ntawm FA-PC piv rau FA. Qhov nce solubility ntawm FA-PC hauv n-octanol tuaj yeem piav qhia los ntawm cov yam ntxwv amorphous ntawm FA-PC. Raws li lipophilicity thiab permeability muaj kev sib raug zoo, cov txiaj ntsig no qhia tau tias kev hloov pauv ntawm cov permeability ntawm FA tuaj yeem txhim kho los ntawm kev siv nws ua phospholipid complex.

DSC 

DSC yog ib txoj hauv kev txhim khu kev qha rau kev tshuaj xyuas cov tshuaj-excipient compatibility thiab muab cov ntaub ntawv siab tshaj plaws txog kev sib cuam tshuam ntawm cov tshuaj thiab cov khoom siv ntxiv [10]. Lub xub ntiag ntawm kev sib cuam tshuam tuaj yeem txiav txim siab los ntawm kev tshem tawm cov peaks endothermic, cov tsos ntawm cov peaks tshiab, kev hloov pauv hauv cov duab ncov thiab qhov pib, qhov kub thiab txias / melting point, thiab thaj chaw txheeb ze lossis enthalpy. Daim duab 1 qhia txog DSC thermograms ntawm FA (Fig. 1a), PC (Fig. 1b), PM (Fig. 1c), thiab FA–PC (Fig. 1d). Lub thermal nkhaus rau ntshiab FA muaj ib tug ntse ntse endothermic melting ntawm txog 172.7 degree, qhia txog nws cov anhydrous thiab crystalline xeev, thaum uas phospholipids muaj ib tug me me endothermal ncov ntawm 231.7-248.6 degree. Lub DSC nkhaus rau PM, muaj xws li cov superimposed thermal profiles rau FA thiab phospholipids, qhia tsis muaj qhov tseem ceeb.Kev hloov pauv tshwj tsis yog hloov me me mus rau qhov kub siab dua (175.9 degree), qhia tias tsis muaj kev cuam tshuam ntawm cov khoom. FA-PC muaj ib lub ncov loj ntawm 158.2 degree, uastxawv ntawm FA ncov, qhia txog kev sib cuam tshuam ntawm FA thiab PC. Peb cov txiaj ntsig tau qhia tias qhov sib txawv ntawm kev sib cuam tshuam thiab / lossis amorphization sib txawvsib tov lossis complexes tuaj yeem tau txais nyob ntawm lawv txoj kev npaj thiab qhov no yog txuam nrog qhov sib txawv ntawm solubility.

FTIR

FTIR spectroscopy tuaj yeem lees paub qhov tsim ntawm FA-PC los ntawm kev sib piv FA-PC spectrum nrog cov ntshiab FA. Daim duab 2 qhia txog FTIR spectra ntawm FA, PC, PM, thiab FA–PC. FA spectrum (Daim duab 2a) qhia txog tus yam ntxwv hydroxyl stretching band ntawm 3436 cm−1. Nws tag nrho dhau los ua ib qho dav dav hauv spectrums FA–PC, PM, thiab phospholipid (Fig. 2b–d). FA spectrum (Fig. 2a) nthuav tawm cov yam ntxwv ncov ntawm 1620 cm−1 (C=C stretching) thiab 1450 cm−1 (C=C aromatic nplhaib stretching). Nyob rau hauv FA–PC spectrum (Fig. 2b), ob lub ncov no tsis pom, tej zaum vim tsis muaj zog los yog tshem tawm, los yog tiv thaiv los ntawm phospholipid molecule.

FA spectrum (Fig. 2a) nthuav tawm cov yam ntxwv tsis zoo ntawm carboxyl peaks ntawm 1691 cm−1 (C=O stretching), thiab 1664 cm−1 (C=C stretching). Nyob rau hauv FA-PC spectrum (Fig. 2b), ob lub ncov no tsis pom, tej zaum vim muaj lub zog txaus nyiam ntawm cov nqi tsis zoo carboxyl hauv FA thiab cov nqi nitrogen zoo hauv phospholipids. phospholipid spectrum (Fig. 2d) tau nce siab ntawm 1733 cm−1 (C=O stretching), 1238 cm−1 (P=O stretching), thiab 1087 cm−1 (P–O -C stretching). Cov

SEM

Qhov chaw morphologies ntawm FA, PC, PM, thiab FA-PC tau tshawb xyuas siv SEM (Fig. 3). Nyob rau hauv daim duab 3c, FA zoo nkaus li crystalline, yuav luag cov duab plaub, thaum FA-PC hais (Daim duab 3a) tshwm sim tsis tu ncua nrog ib tug du nto. FA–PC muaj cov duab sib txawv thiab qhov chaw saum npoo av piv rau FA thiab PC (Fig. 3b). Qhov no tej zaum tshuav qhov ua tiav miscibility ntawm FA hauv PC. Hauv PM scan (Fig. 3d), ob qho tib si FA thiab phospholipids tau yooj yim cais.

Kev tshawb fawb hauv vitro diffusion

Tsis ntev los no, cov hluavtaws Strat-M® membrane tau qhia los ua ib qho kev hloov rau tib neeg cov tawv nqaij hauv vitro diffusion kev tshawb fawb [11]. Strat-M® daim nyias nyias yog tsim los ntawm ob txheej ntawm polyethersulfone uas yog resistant rau diffusion. Cov txheej polyethersulfone pw saum ib txheej ntawm polyolefin, uas yog qhib dua thiab diffusive. Qhov no synthetic membrane yog tus yam ntxwv los ntawm tsawg batch-rau-batch variability, yog li muab cov ntaub ntawv zoo ib yam. Ntxiv mus, nws tau pom tias cov ntaub ntawv diffusion rau Strat-M® daim nyias nyias muaj kev sib raug zoo nrog cov neeg ntawm daim tawv nqaij [11].

Txhawm rau ntsuas qhov cuam tshuam ntawm PC ntawm cov khoom hauv vitro diffusion ntawm FA, feem pua ​​​​Q ntawm FA thiab FA-PC tau npaj rau lub sijhawm. Cov txiaj ntsig hauv daim ntawv tam sim no tau pom qhov sib txawv uas phospholipids nce FA permeation rau hauv Strat-M® daim nyias nyias. Tsis tas li ntawd, FA-PC nyob ntev dua ntawm Strat-M® membrane dua li FA (Fig. 4). Yog li ntawd, kev koom ua ke ntawm phospholipids rau hauv FA tej zaum yuav ua rau nws lub sijhawm nyob hauv stratum corneum thiab ua rau nws tsim nyog rau daim tawv nqaij permeation. Hais txog lub sijhawm permeation, txawm hais tias Strat-M® daim nyias nyias tau tshaj tawm tias muaj kev sib raug zoo nrog tib neeg cov tawv nqaij [11], nws kuj muaj qhov sib txawv ntawm cov tawv nqaij piv nrog tib neeg cov tawv nqaij. Cov kev cuam tshuam ntawm lub sij hawm permeation yuav suav nrog cov kuab tshuaj, qhov concentration ntawm cov tebchaw, pH tus nqi, thiab al. Ntau qhov kev sim yuav tsum tau ua hauv kev tshuaj xyuas yav tom ntej.

Inhibition ntawm melanogenesis

Cov nyhuv ntawm FA-PC ntawm melanin synthesis

Raws li cov ntaub ntawv, FA tuaj yeem inhibit cellular tyrosinase kev ua haujlwm thiab melanogenesis hauv B16F10 melanoma hlwb los ntawm kev txo qis ntawm cov cellular proteins c-kit thiab ERK1/2 [12]. Hauv txoj kev tshawb fawb tam sim no, FA-PC txo qis melanin cov ntsiab lus hauv B16F1{{10}} cov hlwb ntau dua li FA ntawm qhov ntsuas ntawm 0.25, 0.5, thiab 1.0 mg / mL (Table 2). Yog li, peb txiav txim siab tias FA-PC zoo dua FA ntawm inhibiting melanin synthesis hauv B16F10 hlwb.

Xaus

Cov neeg sau ntawv pab txhawb

LL thiab LY tau ua ntau yam kev pab cuam rau kev xav thiab tsim, lossis tau txais cov ntaub ntawv, lossis kev tshuaj xyuas thiab kev txhais cov ntaub ntawv; XY thiab WX tau koom nrog hauv kev sau cov ntawv sau los yog hloov kho nws cov ntsiab lus tseem ceeb ntawm kev txawj ntse; thiab ZJ thiab DY tau txais kev pom zoo zaum kawg ntawm cov ntawv luam tawm. Txhua tus kws sau ntawv tau nyeem thiab pom zoo cov ntawv sau kawg.

Kev lees paub

Txoj haujlwm no tau txhawb nqa los ntawm National Natural Science Foundations ntawm Tuam Tshoj (31501402).

Kev nyiam sib tw

Cov kws sau ntawv tshaj tawm tias lawv tsis muaj kev sib tw nyiam.

Tau txais: 27 Kaum Ob Hlis 2016 Txais: 14 Lub Peb Hlis 2017

Published on: 22 March 2017

Cov ntaub ntawv