Antioxidant cytotoxic antitumor and protective DNAFX抗氧化细胞毒等.docx

Antioxidant cytotoxic antitumor and protective DNAFX抗氧化细胞毒等.docx

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AntioxidantcytotoxicantitumorandprotectiveDNAFX抗氧化细胞毒等

窗体顶端

窗体底端

PharmacognosyRes.2011Jul-Sep;3（3）:

160–165.

doi:

 10.4103/0974-8490.85000

PMCID:

PMC3193615

Copyright:

©PharmacognosyResearch

Antioxidant,cytotoxic,antitumor,andprotectiveDNAdamagemetabolitesfromtheredseabrownalgaSargassumsp

Seif-EldinN.Ayyad,SalehT.Ezmirly,SalimA.Basaif,WaliedM.Alarif,1AdelF.Badria,2andFaridA.Badria3

DepartmentofChemistry,FacultyofScience,KingAbdulAzizUniversity,P.O.80203,Jeddah21589,KSA

1DepartmentofMarineChemsitry,FacultyofMarineSciences,KingAbdulazizUniversity,Jeddah21589,P.O.80207,KSA

2TissueEngineeringLab.,FacultyofDentistry,AlexandriaUniversity,Alexandria,Egypt

3DepartmentofPharmacognosy,FacultyofPharmacy,MansouraUniversity,Mansoura35516,Egypt

Addressforcorrespondence:

Dr.Seif-EldinNAyyad,FacultyofSciences,KingAbdulazizUniversity,Jeddah,SaudiArabia.E-mail:

snayyad2@

ReceivedFebruary1,2011;RevisedApril6,2011;AcceptedSeptember16,2011.

Thisisanopen-accessarticledistributedunderthetermsoftheCreativeCommonsAttribution-Noncommercial-ShareAlike3.0Unported,whichpermitsunrestricteduse,distribution,andreproductioninanymedium,providedtheoriginalworkisproperlycited.

Abstract

Background:

Macroalgaecanbeviewedasapotentialantioxidantandanti-inflammatorysourcesowingtotheircapabilityofproducingcompoundsforitsprotectionfromenvironmentalfactorssuchasheat,pollution,stress,oxygenconcentration,andUVradiations.

Objective:

Toisolatemajorcompoundswhicharemainlyresponsibleforthepharmacologicalactivityofbrownalgaunderinvestigation,Sargassumsp.

MaterialsandMethods:

Algalmaterialwasairdried,extractedwithamixtureoforganicsolvents,andfractionatedwithdifferentadsorbents.Thestructuresofobtainedpurecompoundswereelucidatedwithdifferentspectroscopictechniques,andtwopurematerialsweretestedforprotectionofDNAfromdamage,antioxidant,antitumor,andcytotoxicity.

Results:

Fourpurecompoundswereobtained,ofwhichfucosterol

（1）andfucoxanthin（4）weretested;itwasfoundthatfucoxanthinhasstrongantioxidantandcytotoxicityagainstbreastcancer（MCF-7）withIC50=11.5μg/ml.

Conclusion:

Thenaturallyhighlyconjugatedsafecompoundfucoxanthincouldbeusedasantioxidantandasanantitumorcompound.

Keywords:

Antioxidant,cytotoxicity,fucoxanthin,Sargassumsp

∙ OtherSections▼

oAbstract

oINTRODUCTION

oMATERIALSANDMETHODS

oRESULTSANDDISCUSSION

oREFERENCES

INTRODUCTION

ThegenusSargassumbelongstobrownalgaeofclassPhaeophyceaeintheorderFucales.Numerousspeciesaredistributedthroughoutthetropicalandsubtropicaloceansoftheworld,wheretheygenerallyinhabitshallowwaterandcoralreefs.However,thegenusmaybewellknownforitsplanktonic（free-floating）species.[1]Itwasclearfromtheliteraturesthatmorethan62speciesofthegenusSargassumhasbeeninvestigated,[2]andindicateditsproductivitywithimpressivediversityofnaturalcompounds.Forinstances,thesecondarymetabolitescontaindifferentstructuralclassessuchasplastoquinones,[3–5]chromanols,[6]chromenes,[7,8]steroids,[9]andglycerides.[10]Thepublicationsshowedthat,Sargassummetaboliteshavewiderangeofbiologicalactivities,whichincludeantibiotic,anti-HIV,anticoagulant,anticonvulsant,anti-inflammatory,antineoplastic,andantitumor.[11–14]

Incontinuationofoursearchprogram,whichinterestedintheisolationofsecondarymetabolitesfrommarinesources,especiallymacroalgaecollectedfromRedSea,[15–17]amarinemacroalgae,identifiedasSargassumsp.,collectedfromEl-Shuaibalagoon80kmsouthofJeddah,wasinvestigated.Thetotalextract（Pet.ether:

Chloroform:

Methanol[1:

1:

1]）hadbeenfractionatedusingdifferentchromatographictechniquesandaffordedfourmetabolites;fucosterol

（1）,saringosterone

（2）,saringosterol（3）,andfucoxanthin（4）.1and4hadbeentestedtowardtheBleomycin-dependentDNAdamage,cytotoxicityagainstHepG2（humanhepatocellularlivercarcinomacellline）,WI38（Skincarcinomacellline）,Vero（celllinewasinitiatedfromthekidneyofanormaladultAfricangreenmonkey）,MCF-7（breastcancercelllines）,antitumor,andantioxidantusing2,2‘-azino-bis-3-ethylbenzthiazoline-6-sulfonicacid（ABTS）.

∙ OtherSections▼

oAbstract

oINTRODUCTION

oMATERIALSANDMETHODS

oRESULTSANDDISCUSSION

oREFERENCES

MATERIALSANDMETHODS

Apparatusandmaterial

Chromatographicmaterial:

Aluminumoxidetype60-120meshwasusedforcolumnchromatography.Thin-layerchromatography（TLC）silicagelGF254wasusedforTLC.Preparativethin-layerchromatography（PTLC）wasperformedonaluminumoxideplates（20×20cm）of250-mmthickness.Electronimpactmassspectraweredeterminedat70eVonaKratosMS-25instrument.1Dand2DNMR（NuclearMagneticResonance）spectrawererecordedonBrukerAVANCEIIIWM600MHzspectrometersand13CNMRat150MHzChemicalshiftsaregiveninδ（ppm）relativetoTMS（Tetramethylsilane）asinternalstandard.TheInfraRed（IR）spectrawererecordedonaPerkinElmerspectrometermodel100.Thesprayreagentusedis50%-sulfuricacidinmethanolassprayingreagent.Thechromatoplatewasheatedaftersprayingat100to105°Cuntilthespotsattainedmaximumcolorintensity.ThealgawasdescribedasSargassum（FamilySargassaceae）andwascollectedbyhandfromEl-Shuaibalagoon80kmsouthofJeddah,SaudiArabia,intheRedSea,inJune2010.

ExtractionofSargassumsp

Theair-driedalgalmaterial（350g）wasextractedbyequalvolumeofmixtureofPet.ether,chloroform,andmethanol（2×6l,24hoursforeachbatch）atroomtemperature.Theextractwasconcentratedunderreducedpressuretoobtain10gresidue.ThismaterialwaschromatographedonacolumnofSilicagel.

Columnchromatography

Silicagelcolumn（500g,80×2.5cm）wasusedforresolution.Theresidue（10g）washomogenizedwithsmallamountofsilicagel（50g）andpouredontothetopofthecolumnwhichwaspackedinPet.ether（40:

60）.Theeluentwereusedsuccessively（Pet.ether-Ether,Pet.ether-Ethylacetate）.Fractionswerecollected（50ml）;Pet.ether-EthermixtureofincreasingpolaritybydiethyletherandthenethylacetateandfollowedbyTLCusingsilica-gelchromatoplates,appropriatesolventsystemand50%sulfuricacidinmethanolassprayingreagent.Ifthematerialwasnotpure,preparativeTLCwasappliedusingtheappropriatesolventsystemandtheadsorbentaluminumoxideforpurification.

Isolatedcompounds

ThefractionAelutedbyPet.ether-Ether（6:

4）wascollectedandrechromatographedoverPTLCofsilicagelusingPet.Ether-ethylacetate（8:

2）togivethreecompounds:

1,2,and3.ThefractionBelutedbyPet.ether-Ethylacetate（6:

4）wascollectedandrechromatographedonSephadexLH-20usingamixtureofMeOH-CHCl3（9:

1）andthenfinallypurifiedbypreparativeTLCofsilicagelusingchloroformmethanol（9.5:

0.5）toaffordapurecompound4.

Fucosterol

（1）,（24E）-Stigmasta-5,24（28）-diene-3β-ol:

whitesolid（30mg,0.008%drywt）m.p.133-135°C.IR（cm-1）:

3480（OH）,1945（C=C-H）,1640（C=C）,1370（gem.Di-Me）;EI-MSm/z:

412[M,C29H48O]+,397[M-CH3]+,379[M-CH3-H2O]+,314（100）[M-C6H10O]+.1HNMR（CDCl3）δ（ppm）0.86-0.88（d,J=6.6Hz,6H,Me-26andMe-27）,0.91（d,J=6.6Hz,3H,Me-21）,0.68（s,3H,Me-18）,1.00（s,3H,Me-19）,1.59（d,J=6.6Hz,3H,Me-29）,3.53（m,H,H-3）,5.35（m,H,H-6）,5.11（q,H,H-28）,2.22（sep,J=6.6Hz,H,H-25）.13CNMR（CDCl3）δ（ppm）（39.75,C-1）,（35.20,C-2）,（71.80,C-3）,（42.33,C-4）,（140.74,C-5）,（121.70,C-6）,（36.14,C-7）,（35.78,C-8）,（50.10,C-9）,（39.72,C-10）,（28.23,C-11）,（42.28,C-12）,（42.30,C-13）,（56.74,C-14）,（31.64,C-15）,（34.78,C-16）,（56.72,C-17）,（11.85,C-18）,（19.40,C-19）,（39.50,C-20）,（24.32,C-21）,（37.23,C-22）,（31.90,C-23）,（146.00,C-24）,（33.90,C-25）,（22.50,C-26）,（22.23,C-27）,（115.94,C-28）,（18.90,C-29）.

Saringosterone

（2）,24-vinylcholest-4-ene-3-one:

acolorlessoil（10mg,0.003%drywt）.IR（cm-1）:

3420（OH）,1675（C=O）,1630（C=C）;EIMSm/z（rel.int.）:

426（12）[M,C29H46O2]+,383（21）[M+-C3H7],313（30）,271（35）,269（100）.HREIMS:

m/z426.3413（calcd.426.3349）C29H46O2;1H-NMR（CDCl3）δ5.76（dd,J=17,12Hz,H,H-28）,5.73（brs,H,H-4）,5.27（d,J=12Hz,H,H-29）,5.15（d,J=17Hz,H,H-29）,1.16（s,3H,Me-19）,0.95（d,J=6.5Hz,3H,Me-21）,0.86（d,J=7.0Hz,3H,Me-26）,0.84（d,J=7.0Hz,3H,Me-27）,0.72（s,3H,Me-18）.13C-NMRδ（36.54,C-1）,（34.65,C-2）,（200.35,C-3）,（124.43,C-4）,（172.35,C-5）,（33.64,C-6）,（32.65,C-7）,（36.32,C-8）,（54.41,C-9）,（39.25,C-10）,（21.65,C-11）,（40.22,C-12）,（43.08,C-13）,（56.26,C-14）,（24.85,C-15）,（28.86,C-16）,（56.55,C-17）,（12.63,C-18）,（17.34,C-19）,（36.84,C-20）,（18.35,C-21）,（32.54,C-22）,（28.95,C-23）,（89.76,C-24）,（29.05,C-25）,（19.46,C-26）,（18.07,C-27）,（137.78,C-28）,（117.12,C-29）.

Saringosterol（3）,24-vinylcholest-5-ene-3β,24-diol,saringosterol:

colorlessoil（5mg,0.001%drywt.）.IR（cm-1）:

3445（OH）,1644（C=C）;EIMSm/z（rel.int.）:

428（12）[M,C29H48O2]+,410（6）[M+-H2O],314（40）,273（20）,271（100）,255（28）,228（22）,213（40）,145（64）.1HNMR（CDCl3）d5.73（dd,J=17,12Hz,H,H-28）,5.34（brs,H,H-6）,5.28（d,J=12Hz,H,H-29）,5.17（d,J=17Hz,H,H-29）,3.53（m,H,H-3）,1.02（s,3H,Me-19）,0.97（d,J=6.6Hz,3H,Me-21）,0.88（d,J=6.6Hz,3H,Me-26],0.86（d,J=6.6Hz,3H,Me-27）,0.67（s,3H,Me-18],13C-NMRd（37.17,C-1）,（32.54,C-2）,（72.43,C-3）,（37.94,C-4）,（141.43,C-5）,（122.31,C-6）,（32.35,C-7）,（36.84,C-8）,（50.74,C-9）,（37.93,C-10）,（21.75,C-11