高压乳匀法制备中药固体脂质纳米粒.doc

高压乳匀法制备中药固体脂质纳米粒作者厉英超1；董蕾2；贾皑1；苌新明1；薛挥1西安交通大学医学院1第一附属医院消化内科，陕西西安710061；2第二附属医院消化内科，陕西西安710004摘要目的采用高压乳匀法将中药有效成分包载于固体脂质纳米粒SLN，并研究制备的纳米粒的主要性质。方法选择水飞蓟宾SIL和汉防己甲素TET为模型药物，采用高压乳匀法将其分别包载于SLN。在电镜下观察其形态，以粒度分析仪和ZETA电位分析仪测定其粒径和ZETA电位，用葡聚糖凝胶柱层析法和HPLC测定其包封率和载药量，还观察了SLN的稳定性。结果高压乳匀法制备的SILSLN呈球状，形态规则，平均粒径为1578NM，ZETA电位为3536268MV，包封率为9564，载药量为463；TETSLN呈片状存在，不规则，粒径较小，平均粒径为473NM，ZETA电位为3299254MV，包封率为9782，载药量为476。SILSLN和TETSLN有较高稳定性。结论高压乳匀法适于制备包载中药的SLN。关键词水飞蓟宾；汉防己甲素；固体脂质纳米粒；高压乳匀法；中药现代化中图分类号R944文献标识码A文章编号16734254200605054104PREPARATIONOFSOLIDLIPIDNANOPARTICLESLOADEDWITHTRADITIONALCHINESEMEDICINEBYHIGHPRESSUREHOMOGENIZATIONLIYINGCHAO1；DONGLEI2；JIAAI1；CHANGXINMING1；XUEHUI11DEPARTMENTOFGASTROENTEROLOGY,FIRSTAFFILIATEDHOSPITAL,MEDICALCOLLEGEOFXI’ANJIAOTONGUNIVERSITY,XI’AN710061,CHINA；2DEPARTMENTOFGASTROENTEROLOGY,SECONDAFFILIATEDHOSPITAL,MEDICALCOLLEGEOFXI’ANJIAOTONGUNIVERSITY,XI’AN710004,CHINAABSTRACTOBJECTIVETOINVESTIGATETHEPREPARATIONOFSOLIDLIPIDNANOPARTICLESSLNLOADEDWITHTRADITIONALCHINESEMEDICINESBYHIGHPRESSUREHOMOGENIZATION,ANDSTUDYTHEPHYSICOCHEMICALCHARACTERISTICSOFTHEPARTICLESPRODUCEDBYTHISMETHODMETHODSTHEMODELTRADITIONALCHINESEMEDICINES,SILIBININSILANDTETRANDRINETET,WEREINCORPORATEDINTOSLNSEPARATELYBYHIGHPRESSUREHOMOGENIZATIONTRANSMISSIONELECTRONMICROSCOPEWASEMPLOYEDTOSTUDYTHESHAPEOFTHEPARTICLESPARTICLECHARACTERIZATIONSYSTEMANDZETAPOTENTIALANALYZERWEREUSEDTOSTUDYTHEDIAMETERANDZETAPOTENTIALOFSLNINTHESUSPENSIONTHEENTRAPMENTEFFICIENCYANDDRUGLOADINGWEREDETERMINEDWITHTHESEPHADEXGELCHROMATOGRAPHYANDHIGHPERFORMANCELIQUIDCHROMATOGRAPHYTHESTABILITYOFSLNWASALSOSTUDIEDRESULTSTHESILSLNSPREPAREDBYHIGHPRESSUREHOMOGENIZATIONWERESPHERICALANDREGULARTHEMEANDIAMETERANDZETAPOTENTIALOFSILSLNINDISTILLEDWATERWERE1578NMAND3536268MV,RESPECTIVELYTHEENTRAPMENTEFFICIENCYWAS9564,ANDTHEDRUGLOADINGWAS463THETETSLNWASPLATELETSHAPED,IRREGULARANDSMALLERTHEMEANDIAMETERANDZETAPOTENTIALOFTETSLNWERE473NMAND3299254MV,RESPECTIVELY,WITHDRUGLOADINGOF476,ANDUPTO9782OFTETWASINCORPORATEDSILSLNANDTETSLNHADGOODSTABILITYCONCLUSIONHIGHPRESSUREHOMOGENIZATIONISFEASIBLEFORPREPARINGSLNLOADEDWITHTRADITIONALCHINESEMEDICINESKEYWORDSSILIBININ；TETRANDRINE；SOLIDLIPIDNANOPARTICLES；HIGHPRESSUREHOMOGENIZATION；TRADITIONALCHINESEMEDICINESSUPPORTEDBYSCIENCEANDTECHNOLOGYPROJECTOFXI’ANCITYGG04133LIYINGCHAO1974,PHD,ATTENDINGPHYSICIAN,SPECIALIZEDINPHARMACEUTICALRESEARCHOFLIVERFIBROSIS,TEL02985324001,EMAILL_163COMCORRESPONDINGAUTHORDONGLEI,MEDICALPROFESSOR,TEL0298727693629368,EMAILDONGLEI4488SOHUCOMSOLIDLIPIDNANOPARTICLESSLNSAREPARTICLESMADEFROMSOLIDLIPIDSWITHAMEANDIAMETEROFAPPROXIMATELY50TO1000NMTOSERVEASANALTERNATIVECOLLOIDALCARRIERSYSTEMFORCONTROLLEDDRUGDELIVERY1COMPAREDWITHOTHERPARTICULATECARRIERSSLNHASSEVERALADVANTAGESFORDRUGDELIVERYSUCHASITSGOODBIOCOMPATIBILITY2,BIODEGRADABILITY3,HIGHBIOAVAILABILITY4,ANDEFFECTSTARGETINGTHELIVERANDSPLEENINRECENTYEARS,MARKEDLYINCREASINGSTUDIESONSLNHAVEBEENREPORTED,ESPECIALLYWITHTHEMETHODOFHIGHPRESSUREHOMOGENIZATION5NEVERTHELESS,ONLYAFEWINVESTIGATIONSHAVEBEENCONDUCTEDINREGARDWITHTHEINCORPORATIONOFEFFECTIVECOMPONENTSOFTRADITIONALCHINESEMEDICINESINTOSLNSILYMARINISAPURIFIEDEXTRACTFROMTHEMILKTHISTLESILYBUMMARIANUMLGAERTN,WHICHISCOMPOSEDOFAMIXTUREOF4ISOMERICFLAVONOLIGNANS,NAMELYSILIBININORSILYBIN,SIL,ISOSILIBININ,SILIDIANINANDSILYCHRISTINSIL,WHICHCONSTITUTES6070OFTHESILYMARINMIXTURE,HASBEENIDENTIFIEDASTHEMAJORACTIVECOMPONENT6TETRANDRINETETISABISBENZYLISOQUINOLINEALKALOIDEXTRACTEDFROMTHETRADITIONALCHINESEMEDICINALHERBRADIXSTEPHANIATETRANDRAESILANDTETPOSSESSWIDESPECTRUMSOFPHARMACOLOGICALACTIVITIES7891011THESETWOEFFECTIVECOMPONENTSOFTHETRADITIONALCHINESEMEDICINESHAVEHIGHLIPOPHILICITYANDAREEXCELLENTCANDIDATESFORSLNENCAPSULATIONBYUSINGTHISDRUGDELIVERYSYSTEM,AHIGHBIOAVAILABILITYANDANINTRAVENOUSADMINISTRATIONAREPOSSIBLEINTHEPRESENTSTUDY,SILSLNANDTETSLNWEREPREPAREDBYHIGHPRESSUREHOMOGENIZATION,ANDTHEPHYSICOCHEMICALCHARACTERISTICSOFTHEPARTICLESPRODUCEDBYTHISMETHODWEREANALYZEDMATERIALSANDMETHODSDRUGSANDREAGENTSSIL95WASPURCHASEDFROMPANJINGREENBIOLOGICALDEVELOPMENTCOLTD,CHINATET98WASPURCHASEDFROMSHANCHUANBIOLOGICALCOLTD,XIANCHOLESTERINOBTAINEDFROMZHENGXIANGCHEMICALRESEARCHINSTITUTE,SHANGHAIANDSTEARICACIDTIANDACHEMICALINDUSTRYLTD,TIANJINWEREUSEDSEPARATELYASTHELIPIDMATERIALSOFSLNSOYBEANLECITHINWASOBTAINEDFROMAUBOXINGCOLTD,BEIJINGSEPHADEXGEL50WASPURCHASEDFROMTIANJINCHEMICALINDUSTRYLTDMETHANOLHPLCGRADEANDABSOLUTEALCOHOLWASSUPPLIEDBYXIANCHEMICALINDUSTRYLTDGLYCERINAMOYGLYCERININDUSTRYLTDWASUSEDASACOEMULSIFIERINWATERPHASEPREPARATIONOFSILSLNSIL75MG,CHOLESTERIN15GANDSOYBEANLECITHIN10GWEREWEIGHEDPRECISELYWITHELECTRONICBALANCEBP121S,SARTORIUSLTD,GERMANYANDDISSOLVEDIN10MLABSOLUTEALCOHOLINWATERBATHAT70℃ANAQUEOUSPHASEWASPREPAREDBYDISSOLVING45MLGLYCERININ75MLDISTILLEDWATERTHERESULTANTORGANICSOLUTIONWASRAPIDLYINJECTEDINTOTHESTIRREDAQUEOUSPHASE80℃THERESULTINGSUSPENSIONWASSTIRREDCONTINUOUSLYAT80℃FOR2HTHEORIGINALSILSLNSUSPENSIONWASTHENLOADEDINTOAHIGHPRESSUREHOMOGENIZER15M8BA,APV,UK,5CYCLESAT50MPAANDTHESAMPLESWEREKEPTAT4℃PREPARATIONOFTETSLNTET75MG,STEARICACID15GANDSOYBEANLECITHIN10GWEREWEIGHEDPRECISELYANDPREPAREDINTOTETSLNSUSPENSIONACCORDINGTOTHEMETHODDESCRIBEDABOVETRANSMISSIONELECTRONMICROSCOPYTHEMORPHOLOGYOFSILSLNANDTETSLNWASEXAMINEDWITHTRANSMISSIONELECTRONMICROSCOPEH600,HITACHI,JAPANTHESAMPLESWERESTAINEDWITH2M/VPHOSPHOTUNGSTICACIDFOR30SANDPLACEDONCOPPERGRIDSWITHFILMSFORVIEWINGMEANDIAMETERANDZETAPOTENTIALPARTICLECHARACTERIZATIONSYSTEMMASTERSIZER2000,MALVERNINSTRUMENTS,UK,20NM2000ΜMANDZETAPOTENTIALANALYZERZETASIZERNANO,MALVERNINSTRUMENTS,UKWEREUSEDTOSTUDYTHEDIAMETERANDZETAPOTENTIALOFSLNINDISTILLEDWATERTHREESAMPLESOFSILSLN/TETSLNWEREPREPAREDACCORDINGTOTHEPREVIOUSLYDESCRIBEDMETHODANDEACHSAMPLEWASMEASURED3TIMESTOCALCULATETHEMEANDIAMETERANDZETAPOTENTIALENTRAPMENTEFFICIENCYEEANDDRUGLOADINGDLOFSILSLNCHROMATOGRAPHICCONDITIONTHECHROMATOGRAPHICCOLUMNOFPLANETSILC1846MM15CMWASUSEDWITHMOBILEPHASEOFMETHANOL/01MOL/LPHOSPHATEBUFFER35/65,V/V,PH30,FLOWRATEOF10ML/MIN,COLUMNTEMPERATUREOF40℃,ANDDETECTIONWAVELENGTHOF288NMTHECONTROLSOLUTIONS0050,0161,1605,1419,2838,5675,11350ΜG/MLWASPREPAREDBYDISSOLVINGPRECISELYWEIGHEDSILINTHEMOBILEPHASETHEAMOUNTOFSILENTERINGTHERECEPTORCOMPARTMENTWASDETERMINEDWITHHIGHPERFORMANCELIQUIDCHROMATOGRAMHPLC,LC2010,SHIMADZU,JAPANTHEINTEGRALCALCULUSOFTHECHROMATOGRAPHICPEAKAREAAWASRECORDEDASTHEYAXIS,ANDTHECONCENTRATIONOFSILCASTHEXAXISDRUGRECOVERYWASCALCULATEDFROMTHEFOLLOWINGEQUATIONDRUGRECOVERYMEASUREDDRUGWEIGHTINSLN100/THEORETICALDRUGWEIGHTLOADEDINTHESYSTEMTHESILSLNSUSPENSIONWASSEPARATEDBYSEPHADEXGEL50COLUMNCHROMATOGRAPHYTHECONCENTRATIONSOFSILINTHESUSPENSIONN1ANDFREEDRUGN2WEREASSAYEDBYHPLCAFTERDILUTIONWITHMETHANOLEEANDDLCOULDBECALCULATEDACCORDINGTOTHEFOLLOWINGEQUATIONSEEN1N2/N1100,DLWDRUGLOADEDINSYSTEM/WLIPIDMATRIX100EEANDDLOFTETSLNCHROMATOGRAPHICCONDITIONTHECHROMATOGRAPHICCOLUMNOFSPHERISORBODSC18250MM46MM,5ΜMWASUSEDWITHMOBILEPHASEOFMETHANOL/ETHER/ETHYLAMINEVOLUMEPROPORTIONOF1001005ANDFLOWRATEOF10ML/MINATROOMTEMPERATUREANDDETECTIONWAVELENGTHOF282NMTHEREGRESSIONEQUATION,PERCENTAGERECOVERIESOFTET,EEANDDLOFTETSLNWEREDETERMINEDANDCALCULATEDACCORDINGTOTHEMETHODSANDEQUATIONSDESCRIBEDPREVIOUSLYEVALUATIONOFSTABILITYSILSLNANDTETSLNWERESTOREDAT37℃ANDTHEPARTICLESIZESWEREDETERMINEDAFTER7,45AND90DAYS,RESPECTIVELY,TOEVALUATETHEIRSTABILITYSTATISTICALANALYSISTHERESULTSWEREPRESENTEDASMEANSDSTATISTICALANALYSISWASPERFORMEDUSINGSTUDENTSTTESTWITHP005INDICATINGSIGNIFICANTDIFFERENCERESULTSTRANSMISSIONELECTRONMICROSCOPYTHEELECTRONMICROSCOPYMICROGRAPHSOFSLNLOADEDWITHTRADITIONALCHINESEMEDICINESPREPAREDBYHIGHPRESSUREHOMOGENIZATIONWERESHOWNINFIG1THESILSLNWASSPHERICALANDREGULARA,ANDTHETETSLNAPPEAREDPLATELETSHAPED,IRREGULARANDSMALLERBMEANDIAMETERANDZETAPOTENTIALTHEMEANDIAMETEROFSILSLNWAS1578NM,ANDTHEZETAPOTENTIALWAS3536268MVINDISTILLEDWATERTHEMEANDIAMETEROFTETSLNWAS473NM,ANDTHEZETAPOTENTIAL3299254MVFIG1ELECTRONMICROGRAPHSOFTHEPREPAREDSLNLOADEDWITHTRADITIONALCHINESEMEDICINESORIGINALMAGNIFICATION40000ASILSLN；BTETSLNEEANDDLOFSILSLNTHEREGRESSIONEQUATIONOFSILWASA33071C99102THEASSAYWASLINEARR09999INTHECONCENTRATIONRANGEOF005011350ΜG/MLTHEPERCENTAGERECOVERIESATHIGHANDLOWCONCENTRATIONSWERE9899AND9849,RESPECTIVELY,WITHAMEANOF9896THEEEOFSILSLNWAS9564133ANDTHEDLWAS463021EEANDDLOFTETSLNTHELINEARCALIBRATIONCURVEOFTETWASOBTAINEDINTHERANGEOF12525ΜG/MLR09999THEREGRESSIONEQUATIONOFTETWASA18526C83422THEPERCENTAGERECOVERIESRANGEDFROM9872TO10180MEAN9946UPTO9782145OFTETWASINCORPORATEDINSLN,ANDTHEDLWAS476026STABILITYTAB1SHOWSTHEDATAOFPARTICLESIZESOFSILSLNANDTETSLNAFTER7,45AND90DAYSOFSTORAGEAT37℃THESETWOSLNSUSPENSIONSSHOWEDSUFFICIENTLONGTERMSTABILITYWITHONLYSLIGHTPARTICLEGROWTHP005AFTERSTORAGEAT37℃FOR90DAYSDISCUSSIONSLNSAREACOLLOIDALCARRIERSYSTEMFORCONTROLLEDDRUGDELIVERY,ANDITISCLAIMEDTHATSLNCOMBINESTHEADVANTAGESANDAVOIDSTHEDISADVANTAGESOFOTHERCOLLOIDALCARRIERSITSADVANTAGESINCLUDETHEPOSSIBILITYOFCONTROLLEDDRUGRELEASEANDDRUGTARGETING,INCREASEDDRUGSTABILITY,ABSENCEOFCARRIERBIOTOXICITY,ANDLARGESCALEPRODUCTIONANDSTERILIZATION12HIGHPRESSUREHOMOGENIZATIONHASEMERGEDASARELIABLEANDPOWERFULTECHNIQUEFORSLNPREPARATION12INTHEPRESENTSTUDY,THISMETHODPROVEDTOBEFEASIBLEFORPREPARINGSILSLNANDTETSLN,WHICHARESMALL,STEADYANDHIGHLYINCORPORATEDTHISSUCCESSINDICATESTHEPOSSIBILITYOFINCORPORATINGVARIOUSLIPOPHILICEFFECTIVECOMPONENTSEXTRACTEDFROMTHETRADITIONALCHINESEMEDICINESINSLNBYTHISMETHOD,WHICHMAKEPOSSIBLEHIGHBIOAVAILABILITY,CONTROLLEDDRUGRELEASE,DRUGTARGETING,DECREASEDDRUGTOXICITIESANDMINIMIZEDSIDEEFFECTS,ANDREPRESENTSASUCCESSFULATTEMPTOFNOVELAPPROACHTOTHEMODERNIZATIONOFTRADITIONALCHINESEMEDICINESVARIOUSFACTORSMAYINFLUENCETHEPARTICLESIZEINHIGHPRESSUREHOMOGENIZATION,INCLUDING,FORINSTANCE,HOMOGENIZATIONPRESSURE,NUMBEROFCYCLES,LIPIDSANDEMULSIFIERS/COEMULSIFIERSUSED,ANDOPERATINGTEMPERATUREINTHEPRESENTSTUDY,THETWOKINDSOFSLNWEREPREPAREDUNDERIDENTICALCONDITIONSWITHALMOSTTHESAMEMATERIALSEXCEPTTHEDRUGSINCORPORATEDINSLNANDTHELIPIDMATERIALTHEDIFFERENCEOFTHELIPIDSCHOLESTERINANDSTEARICACIDANDTHEINTERACTIONSBETWEENTHEDRUGSANDTHELIPIDSMIGHTENORMOUSLYCONTRIBUTETOTHEDIFFERENCESOFPARTICLESIZESHOWEVER,THEDETAILEDMECHANISMSREMAINTOBEFURTHERINVESTIGATEDTHESHAPEOFSLNMAYSIGNIFICANTLYDIFFERF