Margatoxin(MgTx)isacomponentofthevenomofScorpioCentruroidesmargaritatus.Margatoxinpreferentiallyinhibitsvoltage-dependentpotassiumchannelsKv1.3withanIC50valuearound50pM(20foldmorepotentthanCharyBDotoxin)andirreversIBLyinhibitstheproliferationresponseofhumanT-cellsat20µMconcentration.MargatoxinisknowntobelesspotentonKv1.3expressedinXenopusOocytes(IC50around1nM).MargatoxinwasalsodescribedtobeapotentinhibitorofhumanvascularsmoothmusclecellmigrationwithanIC50of85pM.
FreesampleDescription:
AAsequence:Thr-Ile-Ile-Asn-Val-Lys-Cys7-Thr-Ser-Pro-Lys-Gln-Cys13-Leu-Pro-Pro-Cys17-Lys-Ala-Gln-Phe-Gly-Gln-Ser-Ala-Gly-Ala-Lys-Cys29-Met-Asn-Gly-Lys-Cys34-Lys-Cys36-Tyr-Pro-His-OH
(DisulfidebondsbetweenCys7-Cys29,Cys13-Cys34andCys17-Cys36)
Length(aa):39
Formula:C178H286N52O50S7
MolecularWeight:4179.03Da
Appearance:Whitelyophilizedsolid
Solubility:waterandsalinebuffer
CASnumber:[145808-47-5]Source:Synthetic
Purityrate:>97%
Reference:
PotentsuppressionofvascularsmoothmusclecellmigrationandhumanneointimalhyperplasiabyKV1.3channelblockers
AIM:
TheaimofthestudywastodeterminethepotentialforK(V)1potassiumchannelblockersasinhibitorsofhumanneoinitimalhyperplasia.
METHODSANDRESULTS:
Bloodvesselswereobtainedfrompatientsormiceandstudiedinculture.Reversetranscriptase-polymerasechainreactionandimmunocytochemistrywereusedtodetectgeneexpression.Whole-cellpatch-clamp,intracellularcalciummeasurement,cellmigrationassays,andorganculturewereusedtoassesschannelfunction.K(V)1.3wasuniqueamongtheK(V)1channelsinshowingpreservedandup-regulatedexpressionwhenthevascularsmoothmusclecellsswitchedtotheproliferatingphenotype.Therewasstrongexpressioninneointimalformations.Voltage-dependentpotassiumcurrentinproliferatingcellswassensitivetothreedifferentblockersofK(V)1.3channels.Calciumentrywasalsoinhibited.Allthreeblockersreducedvascularsmoothmusclecellmigrationandtheeffectswerenon-additive.Oneoftheblockers(margatoxin)washighlypotent,suppressingcellmigrationwithanIC(50)of85pM.Twooftheblockersweretestedinorgan-culturedhumanveinsamplesandbothinhibitedneointimalhyperplasia.
CONCLUSION:
K(V)1.3potassiumchannelsarefunctionalinproliferatingmouseandhumanvascularsmoothmusclecellsandhavepositiveeffectsoncellmigration.Blockersofthechannelsmaybeusefulasinhibitorsofneointimalhyperplasiaandotherunwantedvascularremodellingevents.
CheongA.,etal.(2011)PotentsuppressionofvascularsmoothmusclecellmigrationandhumanneointimalhyperplasiabyKV1.3channelblockers.CardiovascRes.PMID20884640
Kv1.3channelsinpostganglionicsympatheticneurons:expression,function,andmodulation
DocziMA.etal.(2008)Kv1.3channelsinpostganglionicsympatheticneurons:expression,function,andmodulation.AmJPhysiolRegulIntegrCompPhysiol.PMID18614767
PotentsuppressionofKv1.3potassiumchannelandIL-2secretionbydiphenylphosphineoxide-1inhumanTcells
ZhaoN.,etal.(2013)PotentsuppressionofKv1.3potassiumchannelandIL-2secretionbydiphenylphosphineoxide-1inhumanTcells.PLoSOne.PMID23717641
TheeffectsofKv1.3andIKCa1potassiumchannelinhibitiononcalciuminfluxofhumanperipheralTlymphocytesinrheumatoidarthritis
OBJECTIVE:
Thetransientincreaseofthecytoplasmicfreecalciumlevelplaysakeyroleintheprocessoflymphocyteactivation.Kv1.3andIKCa1potassiumchannelsareimportantregulatorsofthemaintenanceofcalciuminfluxduringlymphocyteactivationandpresentapossibletargetforselectiveimmunomodulation.
DESIGN:
Case-controlstudy.
SUBJECTSANDMETHODS:
Wetookperipheralbloodsamplesfrom10healthyindividualsand9recentlydiagnosedrheumatoidarthritis(RA)patientsreceivingnoanti-rheumatictreatment.WeevaluatedcalciuminfluxkineticsfollowingactivationinCD4,Th1,Th2andCD8cellsapplyinganovelflowcytometryapproach.WealsoassessedthesensitivityoftheabovesubsetstospecificinhibitionoftheKv1.3andIKCa1potassiumchannels.
RESULTS:
ThepeakofcalciuminfluxinlymphocytesisolatedfromRApatientsisreachedmorerapidly,indicatingthattheyrespondmorequicklytostimulationcomparedtocontrols.Inhealthyindividuals,theinhibitionoftheIKCa1channeldecreasedcalciuminfluxinTh2andCD4cellstoalowerextentthaninTh1andCD8cells.Onthecontrary,theinhibitionofKv1.3channelsresultedinalargerdecreaseofcalciumentryinTh2andCD4thaninTh1andCD8cells.NodifferencewasdetectedbetweenTh1andTh2orCD4andCD8cellsinthesensitivitytoIKCa1channelinhibitionamonglymphocytesofRApatients.However,specificinhibitionoftheKv1.3channelactsdifferentiallyoncalciuminfluxkineticsinRAlymphocytesubsets.Th2andparticularlyCD8cellsareinhibitedmoredominantlythanTh1andCD4cells.
CONCLUSION:
TheinhibitionofKv1.3channelsdoesnotseemtobespecificenoughinperipheralRAlymphocytes,sinceanti-inflammatoryTh2cellsarealsoaffectedtoanoteworthyextent.
ToldiG.,etal.(2013)TheeffectsofKv1.3andIKCa1potassiumchannelinhibitiononcalciuminfluxofhumanperipheralTlymphocytesinrheumatoidarthritis.ImmunoBIOLOGy.PMID22705192
OverexpressionofDelayedRectifierK(+)ChannelsPromotesInsituProliferationofLeukocytesinRatKidneyswithAdvancedChronicRenalFailure
Leukocytes,suchaslymphocytesandmacrophages,predominantlyexpressdelayedrectifierK(+)channels(Kv1.3),andthechannelsplaycrucialrolesintheactivationandproliferationofthecells.Sincelymphocytesareactivatedinpatientswithend-stagerenaldisease(ESRD),thechannelsexpressedinthosecellswouldcontributetotheprogressionofrenalfibrosisinadvanced-stagechronicrenalfailure(CRF).Inthepresentstudy,usingaratmodelwithadvancedCRFthatunderwent5/6nephrectomyfollowedbya14-weekrecoveryperiod,weexaminedthehistopathologicalfeaturesofthekidneysandtheleukocyteexpressionofKv1.3-channelsandcellcycleMarkers.Age-matchedsham-operatedratswereusedascontrols.InthecorticalinterstitiumofadvancedCRFratkidneys,leukocytesproliferatedinsituandoverexpressedKv1.3channelproteinintheircytoplasm.Treatmentwithmargatoxin,aselectiveKv1.3-channelinhibitor,significantlysuppressedthenumberofleukocytesandtheprogressionofrenalfibrosiswithasignificantdecreaseinthecorticalcellcyclemarkerexpression.ThisstudydemonstratedforthefirsttimethatthenumberofleukocyteswasdramaticallyincreasedinratkidneyswithadvancedCRF.TheoverexpressionofKv1.3channelsintheleukocyteswasthoughttocontributetotheprogressionofrenalfibrosisbystimulatingcellcyclingandpromotingcellularproliferation.
KazamaI.,etal.(2012)OverexpressionofDelayedRectifierK(+)ChannelsPromotesInsituProliferationofLeukocytesinRatKidneyswithAdvancedChronicRenalFailure.IntJNephrol.PMID22701172
CharacteristicsofACh-inducedhyperpolarizationandrelaxationinrabbitjugularvein
BACKGROUNDANDPURPOSE:
Therolesplayedbyendothelium-derivedNOandprostacyclinandbyendothelialcellhyperpolarizationinACh-inducedrelaxationhavebeenwellcharacterizedinarteries.However,themechanismsunderlyingACh-inducedrelaxationinveinsremaintobefullyclarified.
EXPERIMENTALAPPROACH:
ACh-inducedsmoothmusclecell(SMC)hyperpolarizationandrelaxationweremeasuredinendothelium-intactand-denudedpreparationsofrabbitjugularvein.
KEYRESULTS:
Inendothelium-intactpreparations,ACh(≤10⁻⁸M)marginallyincreasedtheintracellularconcentrationofCa²⁺([Ca²⁺](i))inendothelialcellsbutdidnotaltertheSMCmembranepotential.However,ACh(10⁻¹⁰-10⁻⁸M)inducedaconcentration-dependentrelaxationduringthecontractioninducedbyPGF(2α)andthisrelaxationwasblockedbytheNOsynthaseinhibitorN(ω)-nitro-l-arginine.ACh(10⁻⁸-10⁻⁶M)concentration-dependentlyincreasedendothelial[Ca²⁺](i)andinducedSMChyperpolarizationandrelaxation.TheseSMCresponseswereblockedinthecombinedpresenceofapamin[blockerofsmall-conductanceCa²⁺-activatedK⁺(SK(Ca),K(Ca)2.3)channel],TRAM34[blockerofintermediate-conductanceCa²⁺-activatedK⁺(IK(Ca),K(Ca)3.1)channel]andmargatoxin[blockerofsubfamilyofvoltage-gatedK⁺(K(V))channel,K(V)1].
CONCLUSIONSANDIMPLICATIONS:
Inrabbitjugularvein,NOplaysaprimaryroleinendothelium-dependentrelaxationatverylowconcentrationsofACh(10⁻¹⁰-10⁻⁸M).Athigherconcentrations,ACh(10⁻⁸-3×10⁻⁶M)inducesSMChyperpolarizationthroughactivationofendothelialIK(Ca),K(V)1and(possibly)SK(Ca)channelsandproducesrelaxation.TheseresultsimplythatAChregulatesrabbitjugularveintonusthroughactivationoftwoendothelium-dependentregulatorymechanisms.
ItohT.etal.(2012)CharacteristicsofACh-inducedhyperpolarizationandrelaxationinrabbitjugularvein.BrJPharmacol.PMID22595036
Charybdotoxinandmargatoxinactingonthehumanvoltage-gatedpotassiumchannelhKv1.3anditsH399Nmutant:anexperimentalandcomputationalcomparison
Theeffectofthepore-blockingpeptidescharybdotoxinandmargatoxin,bothscorpiontoxins,oncurrentsthroughhumanvoltage-gatedhK(v)1.3wild-typeandhK(v)1.3_H399Nmutantpotassiumchannelswascharacterizedbythewhole-cellpatchclamptechnique.Inthemutantchannels,bothtoxinshardlyblockedcurrentthroughthechannels,althoughtheydidpreventC-typeinactivationbyslowingdownthecurrentdecayduringdepolarization.Moleculardynamicssimulationssuggestedthatthefastcurrentdecayinthemutantchannelwasaconsequenceofaminoacidreorientationsbehindtheselectivityfilterandindicatedthattherigidity-flexibilityinthatregionplayedakeyroleinitsinteractionswithscorpiontoxins.Achannelwithaslightlymoreflexibleselectivityfilterregionexhibitsdistinctinteractionswithscorpiontoxins.Ourstudiessuggestthatthetoxin-channelinteractionsmightpartiallyrestorerigidityintheselectivityfilterandtherebypreventthestructuralrearrangementsassociatedwithC-typeinactivation.
NikoueeA.etal.(2012)Charybdotoxinandmargatoxinactingonthehumanvoltage-gatedpotassiumchannelhKv1.3anditsH399Nmutant:anexperimentalandcomputationalcomparison.JPhysChemB.PMID22490327
Voltage-dependentbiphasiceffectsofchloroquineondelayedrectifierK(+)-channelcurrentsinmurinethymocytes
LymphocytesareofrichindelayedrectifierK(+)-channels(Kv1.3)intheirplasmamembranes,andthechannelsplaycrucialrolesinthelymphocyteactivationandproliferation.Sincechloroquine,awidelyusedanti-malarialdrug,exertsimmunosuppressiveeffects,itwillaffectthechannelcurrentsinlymphocytes.Inthepresentstudy,employingthestandardpatch-clampwhole-cellrecordingtechnique,weexaminedtheeffectsofchloroquineonthechannelsexpressedinmurinethymocytes.Publishedpapersreportthatchloroquinewillinhibitvoltage-dependentK(+)-channelcurrentsbypluggingintotheopen-pore.Weobserved,indeed,thatchloroquinesuppressedthepulse-endcurrentsofKv1.3-channelsathighervoltagesteps.Surprisingly,however,wefoundthatthedrugenhancedthepeakcurrentsatbothhigherandlowervoltagesteps.SincechloroquineshowedsuchbiphasiceffectsonthethymocyteK(+)-channels,andsincethoseeffectswerevoltagedependent,weexaminedtheeffectsofchloroquineontheactivationandtheinactivationofthechannelcurrents.Wenotedthatchloroquineshiftedboththeactivationandtheinactivationcurvestowardthehyperpolarizingpotential,andthatthoseshiftsweremoreemphasizedatlowervoltagesteps.WeconcludethatchloroquinefacilitatesboththeactivationandtheinactivationofKv1.3-channelcurrentsinthymocytes,andthatthoseeffectsarevoltagedependent.
KazamaI.etal.(2012)Voltage-dependentbiphasiceffectsofchloroquineondelayedrectifierK(+)-channelcurrentsinmurinethymocytes.JPhysiolSci.PMID22328488
DifferentpotassiumchannelsareinvolvedinrelaxationofratrenalarteryinducedbyP1075
TheATP-sensitiveK(+)channelsopener(K(ATP)CO),P1075[N-cyano-N’-(1,1-dimethylpropyl)-N″-3-pyridylguanidine],hasbeenshowntocauserelaxationofvariousisolatedanimalandhumanbloodvesselsbyopeningofvascularsmoothmuscleATP-sensitiveK(+)(K(ATP))channels.Inadditiontothewell-knowneffectontheopeningofK(ATP)channels,ithasbeenreportedthatvasorelaxationinducedbysomeoftheK(ATP)COsincludessomeotherK(+)channelsubtypes.GiventhatthereisstillnoinformationonothertypesofK(+)channelspossiblyinvolvedinthemechanismofrelaxationinducedbyP1075,thisstudywasdesignedtoexaminetheeffectsofP1075ontheratrenalarterywithendotheliumandwithdenudedendotheliumandtodefinethecontributionofdifferentK(+)channelsubtypesintheP1075actiononthisbloodvessel.OurresultsshowthatP1075inducedaconcentration-dependentrelaxationofratrenalarteryringspre-contractedbyphenylephrine.Glibenclamide,aselectiveK(ATP)channelsinhibitor,partlyantagonizedtherelaxationofratrenalarteryinducedbyP1075.Tetraethylammonium(TEA),anon-selectiveinhibitorofCa(2+)-activatedK(+)channels,aswellasiberiotoxin,amostselectiveblockeroflarge-conductanceCa(2+)-activatedK(+)(BK(Ca))channels,didnotabolishtheeffectofP1075onratrenalartery.Incontrast,anon-selectiveblockerofvoltage-gatedK(+)(K(V))channels,4-aminopyridine(4-AP),aswellasmargatoxin,apotentinhibitorofK(V)1.3channels,causedpartialinhibitionoftheP1075-inducedrelaxationofratrenalartery.Inaddition,inthisstudy,P1075relaxedcontractionsinducedby20mMK(+),buthadnoeffectoncontractionsinducedby80mMK(+).OurresultsshowedthatP1075inducedstrongendothelium-independentrelaxationofratrenalartery.ItseemsthatK(ATP),4-AP-andmargatoxin-sensitiveK(+)channelslocatedinvascularsmoothmusclemediatedtherelaxationofratrenalarteryinducedbyP1075.
NovakovicA.,etal.(2012)DifferentpotassiumchannelsareinvolvedinrelaxationofratrenalarteryinducedbyP1075.BasicClinPharmacolToxicol.PMID22225832
Chemicalsynthesisandstructure-functionstudiesofmargatoxin,apotentinhibitorofvoltage-dependentpotassiumchannelinhumanTlymphocytes
The39aminoacidpeptide,margatoxin(MgTX),apotentinhibitorofthevoltage-activatedpotassiumchannel(Kv1.3)inhumanTlymphocytes,wassynthesizedbyasolidphasetechnique.FormationofthedisulfidebridgeswasrapidatpH8.2.Thefinalproductwaspurifiedtohomogeneityandwasphysicallyandbiologicallyindistinguishablefromthetoxinpreparedbiosynthetically.Thedisulfidebridgepairingwassimilartothatfoundpreviouslyfortherelatedtoxin-charybdotoxin(3):fromCys7toCys29,fromtestedforinhibitionof125Imargatoxinbindingtovoltage-activatedpotassiumchannels.TheresultsindicatethatthethreeC-terminalresiduesofMgTXareimportantfortheefficienttoxinbindingtoKv1.3.
Bednarek,M.A.,etal.(1994)Chemicalsynthesisandstructure-functionstudiesofmargatoxin,apotentinhibitorofvoltage-dependentpotassiumchannelinhumanTlymphocytes,BiochemBiophysResCommun.PMID: 8297371
Determinationofthethree-dimensionalstructureofmargatoxinby1H,13C,15Ntriple-resonancenuclearmagneticresonancespectroscopy
Thesolution structure ofthe39-residuepeptide margatoxin,ascorpiontoxinthatselectivelyblocksthevoltage-gatedpotassium-channelKv1.3,hasbeendeterminedbyNMR spectroscopy.Thetoxinwasisotopicallylabeledwith 13C and 15N andstudiedusingtwo-dimensionalhomonuclearandthree-andfour-dimensionalheteronuclearNMR spectroscopy.Thefinal structure wasdeterminedusing501constraints,comprising422NOEconstraints,60dihedralangleconstraints,9disulfideconstraints,and10hydrogenbondconstraints.StructureswereinitiallydeterminedwiththeprogramPEGASUSandsubsequentlyrefinedwithX-PLOR.Theaveragermsdeviationfromacalculatedaverage structure forthebackboneatomsofresidues3-38is0.40A.Ahelixispresentfromresidues11to20andincludestwoprolineresiduesatpositions15and16.Aloopatresidues21-24leadsintoatwo-strandantiparallelsheetfromresidues25to38withaturnatresidues30-33.Residues3-6runadjacenttothe33-38strandbutdonotformacanonicalbeta-strand.Thetwoadditionalresiduesof margatoxin,relativetotherelatedtoxinscharybdotoxinandiberiotoxin,insertinamannerthatextendsthebeta-sheetbyoneresidue.Otherwise,theglobal structure isverysimilartothatofthesetwoothertoxins.Thelongersheetmayhaveimplicationsforchannelselectivity.
Johnson,B.A.,etal.(1994)Determinationofthethree-dimensionalstructureofmargatoxinby1H,13C,15Ntriple-resonancenuclearmagneticresonancespectroscopy,Biochemistry.PMID: 7999764
Purification,characterization,andbiosynthesisofmargatoxin,acomponentofCentruroidesmargaritatusvenomthatselectivelyinhibitsvoltage-dependentpotassiumchannels
AnovelpeptidylinhibitorofK+channelshasbeenpurifiedtohomogeneityfromvenomofthenewworldscorpionCentruroidesmargaritatus.Theprimarystructureofthis39-amino-acidpeptide,whichwetermmargatoxin(MgTX),wasdeterminedbyaminoacidcompositionalanalysisandpeptidesequencing.MargatoxinpotentlyinhibitsbindingofrADIolabeledcharybdotoxin(ChTX)tovoltage-activatedchannelsinbrainsynapticplasmamembranes.LikeChTX,MgTXblocksthen-typecurrentofhumanT-lymphocytes(Kv1.3channel),butcomparedtoChTX,is20-foldmorepotent(half-blockatapproximately50pM),hasaslowerdissociationrate,andhasnoeffectoncalcium-activatedchannels.Todemonstratethatthesecharacteristicsareduesolelytothepurifiedtoxin,recombinantMgTXwasexpressedinEscherichiacoliaspartofafusionprotein.Aftercleavageandfolding,purifiedrecombinantMgTXdisplayedthesamepropertiesasnativepeptide.ReplacementoftheCOOH-terminalhistidineresidueofMgTXwithasparagineresultedinapeptidewitha10-foldreductioninpotency.Thiswasduetoafasterapparentdissociationrate,suggestingthattheCOOH-terminalaminoacidmayplayanimportantroleinthebindingofMgTXtotheKv1.3channel.MgTXdisplayssignificantsequencehomologywithpreviouslyidentifiedK+channelinhibitors(e.g.ChTX,iberiotoxin,noxiustoxin,andkaliotoxin).However,givenitspotencyanduniqueselectivity,MgTXrepresentsanespeciallyusefultoolwithwhichtostudythephysiologicroleofKv1.3channels.
Garcia-Calvo,etal.(1993)Purification,characterization,andbiosynthesisofmargatoxin,acomponentofCentruroidesmargaritatusvenomthatselectivelyinhibitsvoltage-dependentpotassiumchannels,JBiolChem. PMID: 8360176
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由弱酸及其盐、弱碱及其盐组成的混合溶液，能在一定程度上抵消、减轻外加强酸或强碱对溶液酸碱度的影响，从而保持溶液的pH值相对稳定。这种溶液称为缓冲溶液。
是否可以理解为纯化水得PH范围为6.3-7.6？能否直接用pH计测量？谢谢!
pH(1)=pKa+lg[c(CH₃COONa)/c(CH₃COOH)]=pKa=4.74
通HCl后,溶液是c(CH₃COOH)=0.2mol/L、c(NaCl)=0.1mol/L的混合溶液,溶液pH按照弱酸溶液pH的求法求.
c(H⁺)=√[Ka*c(CH₃COOH)]=√(10^-4.74*0.2)=0.00191(mol/L)（采用了近似公式）
pH(2)=-lg{c(H⁺)}=2.72
两个pH求得,那么pH的变化量也就可得了.pH的变化量=|pH(2)-pH(1)|=|2.72-4.74|=2.02
1）PH缓冲溶液作用原理和pH值
当往某些溶液中加入一定量的酸和碱时,有阻碍溶液pH变化的作用,称为缓冲作用,这样的溶液叫做缓冲溶液.弱酸及其盐的混合溶液（如HAc与NaAc）,弱碱及其盐的混合溶液（如NH3·H2O与NH4Cl）等都是缓冲溶液.
由弱酸HA及其盐NaA所组成的缓冲溶液对酸的缓冲作用,是由于溶液中存在足够量的碱A-的缘故.当向这种溶液中加入一定量的强酸时,H离子基本上被A-离子消耗：
所以溶液的pH值几乎不变；当加入一定量强碱时,溶液中存在的弱酸HA消耗OH-离子而阻碍pH的变化.
2）PH缓冲溶液的缓冲能力
在缓冲溶液中加入少量强酸或强碱,其溶液pH值变化不大,但若加入酸,碱的量多时,缓冲溶液就失去了它的缓冲作用.这说明它的缓冲能力是有一定限度的.
缓冲溶液的缓冲能力与组成缓冲溶液的组分浓度有关.0.1mol·L-1HAc和0.1mol·L-1NaAc组成的缓冲溶液,比0.01mol·L-1HAc和0.01mol·L-1NaAc的缓冲溶液缓冲能力大.关于这一点通过计算便可证实.但缓冲溶液组分的浓度不能太大,否则,不能忽视离子间的作用.
组成缓冲溶液的两组分的比值不为1∶1时,缓冲作用减小,缓冲能力降低,当c（盐）/c（酸）为1∶1时△pH最小,缓冲能力大.不论对于酸或碱都有较大的缓冲作用.缓冲溶液的pH值可用下式计算：
此时缓冲能力大.缓冲组分的比值离1∶1愈远,缓冲能力愈小,甚至不能起缓冲作用.对于任何缓冲体系,存在有效缓冲范围,这个范围大致在pKaφ（或pKbφ）两侧各一个pH单位之内.
弱酸及其盐（弱酸及其共轭碱）体系pH=pKaφ±1
弱碱及其盐（弱碱及其共轭酸）体系pOH=pKbφ±1
例如HAc的pKaφ为4.76,所以用HAc和NaAc适宜于配制pH为3.76～5.76的缓冲溶液,在这个范围内有较大的缓冲作用.配制pH=4.76的缓冲溶液时缓冲能力最大,此时（c（HAc）/c（NaAc）=1.
3）PH缓冲溶液的配制和应用
为了配制一定pH的缓冲溶液,首先选定一个弱酸,它的pKaφ尽可能接近所需配制的缓冲溶液的pH值,然后计算酸与碱的浓度比,根据此浓度比便可配制所需缓冲溶液.
以上主要以弱酸及其盐组成的缓冲溶液为例说明它的作用原理、pH计算和配制方法.对于弱碱及其盐组成的缓冲溶液可采用相同的方法.
PH缓冲溶液在物质分离和成分分析等方面应用广泛,如鉴定Mg2离子时,可用下面的反应：
白色磷酸铵镁沉淀溶于酸,故反应需在碱性溶液中进行,但碱性太强,可能生成白色Mg（OH）2沉淀,所以反应的pH值需控制在一定范围内,因此利用NH3·H2O和NH4Cl组成的缓冲溶液,保持溶液的pH值条件下,进行上述反应.
这就是说不用酸碱预处理吗？
Whatman的网站上没有DE52最大耐受压力，请问又经验的战友应该是多少？
Whatman的网站上：
DE32DryMicrogranularDEAECellulose
SimilarperformancecharacteristicsafterprecyclingasDE52.
DE52PreswollenMicrogranularDEAECellulose
ProbablythemostwidelyusedDEAEcelluloseintheworld;usedforbiopolymerswithlowtohighnegativecharges;exhibitsexcellentresolutionwithgoodflowrates.
附件是一本图书(MethodsinMolecularMedicine,)的章节,上面说:
WhatmanDEAE52comesalreadypreswollenandonlyneedstobetransferred
totherunningbuffer50mMTE8.
lAntibodiesUsingIonExchangeChromatography.pdf(87.06k)
1.直接用固体磷酸钠配制成50mM的磷酸钠溶液，再调pH到7.4；（我们试着用这个做了下，发现挂不上柱）
2.配置磷酸钠盐缓冲液：按NaH2PO4：Na2HPO4以19：81的摩尔比配制成pH7.4的缓冲液？（附一张百度出来的配方
）
3.如果是磷酸钠盐缓冲液，可以直接将50mM的NaH2PO4的水溶液用NaOH调成pH7.4吗？
再者，2和3这两个方法配制的磷酸钠盐缓冲液有什么区别？最终效果是一样的吗？如果不一样，有什么理论的知识支撑呢？个人感觉是分析化学中酸碱理论中的缓冲液那里的知识。求帮忙解答这些疑问。
另外，我还想问一下，pH对于Ni柱对His-tagged的蛋白的分离纯化影响大吗？是怎么影响的？谢谢大家了！
拼音名：Chunhuashui
英文名：PurifiedWater
【性状】本品为无色的澄清液体；无臭，无味。
【检查】酸碱度取本品10ml，加甲基红指示液2滴，不得显红色；另取10ml，加溴麝香草酚蓝指示液5滴，不得显蓝色。氯化物、流酸盐与钙盐取本品，分置三支试管中，每管各50ml。第一管中加硝酸5滴与硝酸银试液1ml，第二管中加氯化钡试液2ml，第三管中加草酸铵试液2ml，均不得发生浑浊。
硝酸盐取本品5ml置试管中，于冰浴中冷却，加10%氯化钾溶液0.4ml与0.1%二苯胺硫酸溶液0.1ml，摇匀，缓缓滴加硫酸5ml，摇匀，将试管子50℃水浴中放置15分钟，溶液产生的蓝色与标准硝酸盐溶液［取硝酸钾0.163g，加水溶解并稀释至100ml，摇匀，精密量取1ml，加水稀释成100ml，再精密量取10ml，加水稀释成100ml，摇匀，即得（每1ml相当于1pgNO3）0.3ml，加无硝酸盐的水4.7ml，用同一方法处理后的颜色比较，不得更深（0.000006%）。
亚硝酸盐取本品10ml，置纳氏管中，加对氨基苯磺酰胺的稀盐酸溶液（1→100）lml与盐酸菜乙H肢溶液（0．l＋100）1ml，产生的粉红色，与标准亚硝酸盐溶液〔取亚硝酸钠0.750g（按干燥品计算），加水溶解，稀释至100ml，摇匀，精密量取1ml，加水稀释成100ml，摇匀，再精密量取1ml，加水稀释成50ml，摇匀，即得（每1ml相当于1μgNO2）]0.2ml，加无亚硝酸盐的水9.8ml，用同一方法处理后的颜色比较，不得更深（0.000002%）。
氨取本品50ml，加碱性碘化汞钾试液2ml，放置15分钟；如显色，与氯化铵溶液（取氯化铵31.5mg，加无氨水适量使溶解并稀释成1000ml）1.5ml，加元氨水48ml与碱性碘化汞钾试液2ml制成的对照液比较，不得更深（0.00003%）。
二氧化碳取本品25ml，置50ml具塞量筒中，加氢氧化钙试液25ml，密塞振摇，放置，小时内不得发生浑浊。
易氧化物取本品100ml，加稀硫酸10ml，煮沸后，加高锰酸钾滴定液（0.02mol/L）0.10ml，再煮沸10分钟，粉红色不得完全消失。
不挥发物取本品100ml，置105℃恒重的蒸发皿中，在水浴上蒸干，并在105℃干燥至恒重，遗留残渣不得过1mg。
重金属取本品50ml，加水18.5ml，蒸发至20ml，放冷，加醋酸盐缓冲液（pH3.5）2ml与水适量使成25ml，加硫代乙酰胺试液2ml，摇匀，放置2分钟，与标准铅溶液1.5ml加水18.5ml用同一方法处理后的颜色比较，不得更深（0.00003%）。
微生物限度取本品，采用薄膜过滤法处理后，依法检查（附录ⅪJ），细菌、霉菌和酵母菌总数每1ml不得过100个。
【贮藏】密闭保存。
【化学成分】本品为蒸馏法、离子交换法、反渗透法或其他适宜的方法制得的供药用的水，不含任何附加剂。
【分子式与分子量】H2O18.02
【药理作用】溶剂、稀释剂
这里药典纯化水标准中并无PH值项目，请问对纯化水有PH值的要求吗，范围应在多少？请说明出处？
在纯化水检测中，检验酸碱度合格，但是发现PH在8左右。如果按以上标准检验合格，是否要考虑PH值？请知道的解答，谢谢！
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