外文文献透析袋Tech Notes参考资料 The Dialysis Process Permeation Rate Influence on Permeation Rate Dialysis Mem

TheDialysisProcess

Thedialysisprocessofasoluteoccursatthemembranesurface,incontexttotheconcentrationdifferencesoftwosolutionsoneithersideofthemembrane.Thisprocessisaffectedbythevariablesoftemperature,viscosityandmixingrateofasolution.

Themovementofasoluteacrossasemipermeablemembraneistheresultofrandommolecularmotion.Asthesolutemoleculesinasolutionmove,theywillcollidefromtimetotimewiththemembraneuntiltheydiffuse.(Seediagram1.)

Thepermeationofagivensolute(Y),fromasolutionontheleft(L)ofthemembranetotheright(R),andbackagain,willdependuponthefrequencyofcollisionsbetweenYmoleculesoneithersideofthemembrane.

Forexample,iftheconcentrationofsolute-Y,inthesolutionLis100mM,andsolutionRis10mM,theprobabilityofY-solutecollidingwiththeY-soluteinthesolutionL,willbemuchhigherthanthechanceofthesolute-YcollidingwiththeRsideofthemembrane.Therefore,thenetrateoftransferofagivensolute,(atacertaintemperature,viscosityandmixingrate)willincreasewithgreaterconcentrationdifferencesbetweenthetwosolutions.

PermeationRate

Therateoftransportacrossasemipermeablemembranedependsmainlyontheshape,charge,andsizeofthesolute.Speedandsizeareonlytwooutofnumerousinteractingfactors.Theflux,orpermeationacrossasemipermeablemembraneofsolutesinsolution,isinverselyrelatedtothemolecularweight.

Smallmoleculescollidemoreoftenwiththemembrane,thus,theirrateofmolecularmigrationthroughthemembranewillbehigh.Largemolecules,movingatlowvelocitiescollideinfrequentlywiththemembrane.Therefore,theirrateofmigrationthroughthesemi-permeablemembranewillbelow(eventhosethatfitthroughthemembranepores).
Thesizeofasolutecorrelateshighlywiththemolecularweight.Asthemolecularsizeapproachesandexceedsthesizeofthemembranepores(MWCO),passageofsoluteswillcompletelyorpartiallybeprevented.

InfluenceonPermeationRate

ParameterConcentrationGradientMolecularSizeTemperatureWallThicknessMembraneSurfaceArea

IncreasedLargeConcentrationDifferentialSmallSphericalMoleculesHighTemperatureThin(<20μm)Large

DecreasedSmallConcentrationDifferentialLargeFibrousMoleculesLowTemperatureThick(>20μm)Small

DialysisMembraneMaterials

Syntheticandnaturalmembranesarecommonlyusedforfiltrationapplications.Membranematerialsmostoftenusedincluderegeneratedcellulose,celluloseacetate,polysulfone,polycarbonate,polyethylene,polyolefin,polypropylene,andpolyvinylidenefluoride.
TheregeneratedcelluloseinCelluSepisderivedfromcotton:Cottonlintersaredissolvedinasolutionandspreadintoflatsheetsorextrudedintotubes.Thematerialistreatedwithglycerintopreventtheporesfromcollapsingandairdriedatacertaintemperatureandpressuretoformarigidmembrane.CelluSepregeneratedcellulosemembranehasasymmetricporestructurewhichallowssmallmoleculestomigrateineitherdirection,makingitidealforexperimentalpurposes.

TheregeneratedcelluloseinCelluSepisderivedfromcotton:Cottonlintersaredissolvedinasolutionandspreadintoflatsheetsorextrudedintotubes.Thematerialistreatedwithglycerintopreventtheporesfromcollapsingandairdriedatacertaintemperatureandpressuretoformarigidmembrane.CelluSepregeneratedcellulosemembranehasasymmetricporestructurewhichallowssmallmoleculestomigrateineitherdirection,makingitidealforexperimentalpurposes.

T-SeriesInstructions

Formostdialysisapplications,CelluSepmembrane
canbeuseddirectlyfromtherollafterabriefrinseindistilled
water.

HowLongtoDialyze...

Low-molecularweightsaltsandbuffers(e.g.,TrisClandKPO4)equilibratewithin3hourswithstirring.Equilibrationtimesforviscoussampleswillbelonger.
Changethedialysisbufferasnecessary.Usuallytwodialysisbufferchangesaresufficient.WhenCsClisremovedfromequilibriumdensitygradient-bandedDNA,twoequilibrationsagainsta1000-foldvolumeexcessofbufferwilldecreaseCsClconcentration106-fold,toastill-significant5M,anditmaybenecessarytochangebufferathirdtime.

ProteinAdsorption

CelluSepmembranesexhibitaverylownon-specificproteinadsorptionwhencomparedwithcompetitivemembranes.CytochromeCsolutionsexhibitalargeadsorptivelosswithcompetitivemembranesfollowingafivefoldconcentrationof150g/mlsolution.

MembraneArea:14.5cm2(43mmdisk),StartingConcentration:150g/ml(7.5mgCytochromeC)

AdsorptionLossg/cm2RetentionCelluSep0.002%1.0098.5%CelluloseEster0.800%4.1397.0%CelluloseAcetate2.300%11.897.0%

Inadditiontoverylownon-specificadsorptiontheCelluSepmembraneshaveexcellentchemicalresistanceagainstorganicsolvents.

SensitiveAssayMembranePre-treatment

Ifglycerol,sulfurcompounds,orsmallamountsofheavymetalswillinterferewithsubsequentsteps,themembrane首ldbepreparedasdescribedbelow.Forconvenience,pre-treated,highlycleanCelluSepH1HighGradewetmembranes,designedforsensitiveapplications,mayalsobeused.

Wearinggloves,cutthedialysismembraneintothedesiredlengthandsoakindistilledwaterfor15minutes.Heatthepre-cutmembranefor30minutes,whilestirring,to80°Cinalargevolumeof10mMsodiumbicarbonate.Transferthemembranesintoa10mMNa2EDTAsolutionandsoakfor30minutes.Replacesolutionwithdistilledwaterandstirfor30minutesat80°C.Allowmembranetocooldown,andstoreinarefrigeratorina0.05%sodiumazidesolution,ora0.10%sodiumbenzoatesolution.Alternatively,a20-50%ethanolsolutionmaybeused.Tubingmustalwaysremainsubmersed.Beforeuse,washtubinginsideandoutwithdistilledwaterandconditionindialysisbuffer(ifnecessary,tubingmaybesterilized).Secureclampononeendofmembrane.Bufferorwater首ldbeplacedinsidethebagtoensureintegrityoftheseal.Checkintegrityofthetubingandclamps.Pourouttestsolutionandloadsample(Forconcentratedsaltsamples,leavespaceinthetubingtoallowfornetflowofwaterintothesampleandtopreventtubingfrombursting.)Immersedialysistubinginbeakerorflaskcontainingalargevolume(usually100to1000-foldthatofthesample)ofthedesiredbufferanddialyzeforseveralhoursatthedesiredtemperaturewithgentlestirring.
NOTE:Low-molecularweightsaltsandbuffers(e.g.,TrisClandKPO>4)equilibratewithin3hourswithstirring.Equilibrationtimesforviscoussampleswillbelonger.Changethedialysisbufferasnecessary.Usuallytwodialysisbufferchangesaresufficient.WhenCsClisremovedfromequilibriumdensitygradient-bandedDNA,twoequilibrationsagainsta1000-foldvolumeexcessofbufferwilldecreaseCsClconcentration106-fold,toastill-significant5M,anditmaybenecessarytochangebufferathirdtime.Removedialysistubingfrombuffer,removeclampfromoneend,andremovesamplewithapipet.

MembraneSterilization

Thecommonmethodofmembranesterilizationisexposuretoethyleneoxidegas.Alternativesterilizationmethodsaregammairradiationandsteamautoclaving.Severalpublishedabstractsillustrateadecreaseinpermeabilityafterasterilizationprocess,dependingupontheappliedmethod.Belowaresomeofthetechniquesdescribed.
Suggestedpreparationofmembranesbeforesterilizationistosoakthemembranefor30minutesindistilledwater.Afterwashing,themembranemightbestoredina0.85%sodiumchloridesolutioncontaining0.1%formaldehyde.

ChemicalSterilizationwithEtO

Placethepreparedmembraneinanopenpolyethylenebaginavacuumoven.Evacuateandfilltheovenwithagasmixtureof20%EtO/80%CO2byatotalpressureof1atm.Treatthemembranefor5hoursat40°C.Evacuatethesterilizinggasandadmit50%ofrelativehumidityofair.Aslightreduction,approximately10%permeabilitycharacteristic,hasbeenreportedwiththeuseofthisEtOmethod.

Gamma-irradiation

Sealthemembraneinapolyethylenebagandexposetoagammaraysourceforatotaldoseof2.5Megarads.Duringexposurethetemperature首ldnotincreasetomorethan10°C.Thepermeabilitycharacteristicafterthetreatmentisapproximately75%.

Autoclaving

Thelengthoftheautoclavingcycle首ldbekeptasshortaspossible.Membranesmaybesafelyautoclavedat121°Cat100kPa(1bar)for10minutesifsubmersedindistilledwater.They首ldnotbepermittedtodryoutafterwards.Dryheatingoveraperiodof48hoursat80°Cdropsthepermeabilitycharacteristictoabout50%.

Micro-organismEffects

Membranes,whenwet,aremostsusceptibletomicrobesandfungi.Suchmicrobialgrowthswillimpairthedialysispropertiesofthematerialandcausedecreasedyieldandinfectionofthesample.Therefore,tubing首ldnotbeleftwithoutsatisfactoryprotectionagainstbacteriaandfungi.
Sodiumazideandsodiumbenzoatearecommonlyusedpreservativesforserologicalmaterial.Aconcentrationof0.05-0.10%sodiumazideisrecommendedforthispurpose.However,aconsiderablylowerconcentration,around0.02%,seemstogivesatisfactoryprotection,althoughstreptococcigrowthmayoccuratthisconcentration.

Precaution

SodiumAzide(NaN3)isastandardpreservativeforlaboratoryreagents.AlthoughNaN3isconsideredhighlytoxicitsLD50inratsis45mg/kgorally.
MFPIrecommendsthefollowingSodiumBenzoate(C7H5NaO2)solutioninadditiontoSodiumAzide(NaN3)forsuppressingmicrobiologicalgrowthinwetstoreddialysismembranes.SodiumBenzoateisconsiderablylesstoxicthanSodiumAzide(LD50inratsofC7H5NaO2is4.07g/kgorally).
SodiumBenzoateiscommonlyusedasapreservativeinpharmaceuticalsandinfoodproducts(notmorethan1in1000partsbeingpermitted).Althoughitspreservativeeffectisbestexhibitedinslightlyacidicmedia;inneutralmediaitprovidessatisfactoryprotectionofthemembrane.

Shortinstructionforpreparationofa0.5Molar100mlSodiumBenzoateSolution:

Reagent:C7H5NaO2,MolecularWeight:144.04
Dissolve7.2gramsofC7H5NaO2in100mlofwater.ThesolutionmaybebroughttoaboiltospeedthedissipationoftheC7H5NaO2.

PackingandStorage

AllCelluSepdialysismembranesarepackagedinuniquedispenserboxestofacilitateeaseinhand领andmeasuring.Themembranesareenclosedinpolyethylenebags,containingdesiccantpouchestopreventexcessmoistureandreducetheriskofcontamination.Themembranescontainglycerol,addedasahumidifierandplasticizertomaintainproductintegrity.
Topreventdryingandsubsequentbrittleness,itisvitalforthetubingtobestoredproperly.Moisturelosscancauselossofmembraneflexibilityandresultinpinholesduringhand领.Inordertoavoidthispossibility,itisrecommendedthatunusedtubingbestoredintheoriginalpolyethylenebagoranyotherairtightmoisture-proofcontainerinacoolplace(refrigerator),inaminimumof35%relativehumidity.

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2004-10-18