高壓乳勻法制備中藥固體脂質納米粒作者：厲英超1；董蕾2；賈皚1；萇新明1；薛揮1(西安交通大學醫學院1第一附屬醫院消化內科，陜西西安710061；2第二附屬醫院消化內科，陜西西安710004)摘要：目的采用高壓乳勻法將中藥有效成分包載于固體脂質納米粒(SLN)，并研究制備的納米粒的主要性質。方法選擇水飛薊賓(SIL)和漢防己甲素(TET)為模型藥物，采用高壓乳勻法將其分別包載于SLN。在電鏡下觀察其形態，以粒度分析儀和Zeta電位分析儀測定其粒徑和Zeta電位，用葡聚糖凝膠柱層析法和HPLC測定其包封率和載藥量，還觀察了SLN的穩定性。結果高壓乳勻法制備的SIL-SLN呈球狀，形態規則，平均粒徑為(1578)nm，Zeta電位為(-35.362.68)mV，包封率為95.64%，載藥量為4.63%；TET-SLN呈片狀存在，不規則，粒徑較小，平均粒徑為(473)nm，Zeta電位為(-32.992.54)mV，包封率為97.82%，載藥量為4.76%。SIL-SLN和TET-SLN有較高穩定性。結論高壓乳勻法適于制備包載中藥的SLN。關鍵詞：水飛薊賓；漢防己甲素；固體脂質納米粒；高壓乳勻法；中藥現代化中圖分類號：R944文獻標識碼：A文章編號：1673-4254(2006)05-0541-04PreParationofsolidlipidnanoParticlesloadedwithtraditionalChinesemedicinebyhigh-pressurehomogenizationLIYing-chao1；DONGLei2；JIAAi1；CHANGXin-ming1；XUEHui11DePartmentofGastroenterology,FirstAffiliatedHospital,MedicalCollegeofXianJiaotongUniversity,Xian710061,China；2DePartmentofGastroenterology,SecondAffiliatedHospital,MedicalCollegeofXianJiaotongUniversity,Xian710004,ChinaAbstract:ObjectiveToinvestigatethepreParationofsolidlipidnanoParticles(SLN)loadedwithtraditionalChinesemedicinesbyhigh-pressurehomogenization,andstudythephysicochemicalcharacteristicsoftheParticlesproducedbythismethod.MethodsThemodeltraditionalChinesemedicines,silibinin(SIL)andtetrandrine(TET),wereincorporatedintoSLNseParatelybyhigh-pressurehomogenization.TransmissionelectronmicroscopewasemployedtostudytheshapeoftheParticles.ParticlecharacterizationsystemandzetapotentialanalyzerwereusedtostudythediameterandzetapotentialofSLNinthesuspension.Theentrapmentefficiencyanddrugloadingweredeterminedwiththesephadexgelchromatographyandhigh-performanceliquidchromatography.ThestabilityofSLNwasalsostudied.ResultsTheSIL-SLNspreParedbyhigh-pressurehomogenizationweresphericalandregular.ThemeandiameterandzetapotentialofSIL-SLNindistilledwaterwere1578nmand-35.362.68mV,respectively.Theentrapmentefficiencywas95.64%,andthedrugloadingwas4.63%.TheTET-SLNwasplatelet-shaped,irregularandsmaller.ThemeandiameterandzetapotentialofTET-SLNwere473nmand-32.992.54mV,respectively,withdrugloadingof4.76%,andupto97.82%ofTETwasincorporated.SIL-SLNandTET-SLNhadgoodstability.ConclusionHigh-pressurehomogenizationisfeasibleforpreParingSLNloadedwithtraditionalChinesemedicines.Keywords:silibinin；tetrandrine；solidlipidnanoParticles；high-pressurehomogenization；traditionalChinesemedicinesSupportedbyScienceandTechnologyProjectofXianCity(GG04133).LIYing-chao(1974-),PhD,attendingphysician,specializedinpharmaceuticalresearchofliverfibrosis,TelE-mail:l_163.comCorrespondingauthor:DONGLei,medicalprofessor,Tel29368,E-mail:SolidlipidnanoParticles(SLNs)areParticlesmadefromsolidlipidswithameandiameterofapproximately50to1000nmtoserveasanalternativecolloidalcarriersystemforcontrolleddrugdelivery1.ComParedwithotherParticulatecarriersSLNhasseveraladvantagesfordrugdeliverysuchasitsgoodbiocomPatibility2,biodegradability3,highbioavailability4,andeffectstargetingtheliverandspleen.Inrecentyears,markedlyincreasingstudiesonSLNhavebeenreported,especiallywiththemethodofhigh-pressurehomogenization5.Nevertheless,onlyafewinvestigationshavebeenconductedinregardwiththeincorporationofeffectivecomponentsoftraditionalChinesemedicinesintoSLN.SilymarinisapurifiedextractfromthemilkthistleSilybummarianum(L.)Gaertn,whichiscomposedofamixtureof4isomericflavonolignans,namelysilibinin(orsilybin,SIL),isosilibinin,silidianinandsilychristin.SIL,whichconstitutes60%-70%ofthesilymarinmixture,hasbeenidentifiedasthemajoractivecomponent6.Tetrandrine(TET)isabisbenzylisoquinolinealkaloidextractedfromthetraditionalChinesemedicinalherbRadixstephaniatetrandrae.SILandTETpossesswidespectrumsofpharmacologicalactivities7891011.ThesetwoeffectivecomponentsofthetraditionalChinesemedicineshavehighlipophilicityandareexcellentcandidatesforSLNencapsulation.Byusingthisdrugdeliverysystem,ahighbioavailabilityandanintravenousadministrationarepossible.Inthepresentstudy,SIL-SLNandTET-SLNwerepreParedbyhigh-pressurehomogenization,andthephysicochemicalcharacteristicsoftheParticlesproducedbythismethodwereanalyzed.MATERIALSANDMETHODSDrugsandreagentsSIL(95%)waspurchasedfromPanjinGreenBiologicalDevelopmentCo.Ltd.,China.TET(98%)waspurchasedfromShanchuanBiologicalCo.Ltd.,Xian.Cholesterin(obtainedfromZhengxiangChemicalResearchInstitute,Shanghai)andstearicacid(TiandaChemicalIndustryLtd.,Tianjin)wereusedseParatelyasthelipidmaterialsofSLN.SoybeanlecithinwasobtainedfromAuboxingCo.Ltd.,Beijing.Sephadexgel-50waspurchasedfromTianjinChemicalIndustryLtd.Methanol(HPLCgrade)andabsolutealcoholwassuppliedbyXianChemicalIndustryLtd.Glycerin(AmoyGlycerinIndustryLtd.)wasusedasacoemulsifierinwaterphase.PreParationofSIL-SLNSIL(75mg),cholesterin(1.5g)andsoybeanlecithin(1.0g)wereweighedpreciselywithelectronicbalance(BP-121S,sartoriusLtd.,Germany)anddissolvedin10mlabsolutealcoholinwaterbathat70.AnaqueousphasewaspreParedbydissolving45mlglycerinin75mldistilledwater.Theresultantorganicsolutionwasrapidlyinjectedintothestirredaqueousphase(80).Theresultingsuspensionwasstirredcontinuouslyat80for2h.TheoriginalSIL-SLNsuspensionwasthenloadedintoahigh-pressurehomogenizer(15M-8BA,APV,UK,5cyclesat50MPa)andthesampleswerekeptat4.PreParationofTET-SLNTET(75mg),stearicacid(1.5g)andsoybeanlecithin(1.0g)wereweighedpreciselyandpreParedintoTET-SLNsuspensionaccordingtothemethoddescribedabove.TransmissionelectronmicroscopyThemorphologyofSIL-SLNandTET-SLNwasexaminedwithtransmissionelectronmicroscope(H-600,Hitachi,JaPan).Thesampleswerestainedwith2%(m/V)phosphotungsticacidfor30sandplacedoncoppergridswithfilmsforviewing.MeandiameterandzetapotentialParticlecharacterizationsystem(Mastersizer2000,MalvernInstruments,UK,20nm-2000m)andzetapotentialanalyzer(ZetasizerNano,MalvernInstruments,UK)wereusedtostudythediameterandzetapotentialofSLNindistilledwater.ThreesamplesofSIL-SLN/TET-SLNwerepreParedaccordingtothepreviouslydescribedmethodandeachsamplewasmeasured3timestocalculatethemeandiameterandzetapotential.Entrapmentefficiency(EE)anddrugloading(DL)ofSIL-SLN.Chromatographiccondition:ThechromatographiccolumnofPlanetsilC18(4.6mm15cm)wasusedwithmobilephaseofmethanol/0.1mol/Lphosphatebuffer(35/65,V/V,pH3.0),flowrateof1.0ml/min,columntemperatureof40,anddetectionwavelengthof288nm.Thecontrolsolutions(0.050,0.161,1.605,14.19,28.38,56.75,113.50g/ml)waspreParedbydissolvingpreciselyweighedSILinthemobilephase.TheamountofSILenteringthereceptorcomPartmentwasdeterminedwithhigh-performanceliquidchromatogram(HPLC,LC-2010,Shimadzu,JaPan).Theintegralcalculusofthechromatographicpeakarea(A)wasrecordedastheYaxis,andtheconcentrationofSIL(C)astheXaxis.Drugrecoverywascalculatedfromthefollowingequation:Drugrecovery=measureddrugweightinSLN100%/theoreticaldrugweightloadedinthesystem.TheSIL-SLNsuspensionwasseParatedbySephadexgel-50columnchromatography.TheconcentrationsofSILinthesuspension(n1)andfreedrug(n2)wereassayedbyHPLCafterdilutionwithmethanol.EEandDLcouldbecalculatedaccordingtothefollowingequations:EE%=(n1-n2)/n1100%,DL=Wdrugloadedinsystem/Wlipidmatrix100%.EEandDLofTET-SLNChromatographiccondition:ThechromatographiccolumnofSpherisorbODSC18(250mm4.6mm,5m)wasusedwithmobilephaseofmethanol/ether/ethylamine(volumeproportionof100:1:0.05)andflowrateof1.0ml/minatroomtemperatureanddetectionwavelengthof282nm.Theregressionequation,percentagerecoveriesofTET,EEandDLofTET-SLNweredeterminedandcalculatedaccordingtothemethodsandequationsdescribedpreviously.EvaluationofstabilitySIL-SLNandTET-SLNwerestoredat37andtheParticlesizesweredeterminedafter7,45and90days,respectively,toevaluatetheirstability.StatisticalanalysisTheresultswerepresentedasMeanSD.StatisticalanalysiswasperformedusingStudentsttestwithP0.05)afterstorageat37for90days.DISCUSSIONSLNsareacolloidalcarriersystemforcontrolleddrugdelivery,anditisclaimedthatSLNcombinestheadvantagesandavoidsthedisadvantagesofothercolloidalcarriers.Itsadvantagesincludethepossibilityofcontrolleddrugreleaseanddrugtargeting,increaseddrugstability,absenceofcarrierbiotoxicity,andlargescaleproductionandsterilization12.High-pressurehomogenizationhasemergedasareliableandpowerfultechniqueforSLNpreParation12.Inthepresentstudy,thismethodprovedtobefeasibleforpreParingSIL-SLNandTET-SLN,whicharesmall,steadyandhighlyincorporated.ThissuccessindicatesthepossibilityofincorporatingvariouslipophiliceffectivecomponentsextractedfromthetraditionalChinesemedicinesinSLNbythismethod,whichmakepossiblehighbioavailability,controlleddrugrelease,drugtargeting,decreaseddrugtoxicitiesandminimizedsideeffects,andrepresentsasucc