HighPerformanceLiquidChromatography(HPLC)andPharmaceutical Analysis
Highperformanceliquidchromatography(HPLC)hasbecomeanessential techniqueinpharmaceuticalanalysisfortheseparation,identification,and quantificationofpharmaceuticalsDuetoitshighresolutionandsensitivity,itis usedfortestingdrugpurity,monitoringdegradation,studying pharmacokinetics,andexaminingdrugstabilityWithrecentadvancesin instrumentation,thetechniquehasbecomemoreefficientintermsof separatingmultiplecomponentsandanalyzingtrickychemicalcompounds TheuseofHPLCinpharmaceuticalanalysishasimproveddrugsafetyand efficacy,providingavaluabletoolforpharmaceuticaldevelopmentandquality control.Inconclusion,HPLChasbecomeacriticaltoolforpharmaceutical analysisduetoitshigh-resolutionandunparalleledsensitivityinseparating andquantifyingcomplexchemicalcompounds.
Inthepast30years,gaschromatography(GC)hasshownitsunique applicationinpharmaceuticalanalysisItisgenerallyusedfortheinspectionof residualsolventsinvariousrawmaterialsandpreparations,andthequalitative andquantitativeanalysisofvariousvolatilecomponentsThechromatographic techniquethathasalwaysoccupiedthemainstreampositionindruganalysisis HPLC,whichhasbeenwidelyusedintheanalysisandidentificationof metabolitesDependingonthemechanismofaction,variousHPLCtechniques suchasadsorptionchromatography,partitionchromatography(normalphase, reversedphase),ionexchangechromatography,affinitychromatography,gel filtrationchromatography(alsoknownassizeexclusionchromatography)and gelpermeationchromatographycanallbeusedThereareawiderangeof applications,ofwhichthemostwidelyusedisstillthereversed-phasemode (reversed-phaseHPLC,RP-HPLC)
Inrecentyears,newbrancheshavebeendevelopedonthebasisofpartition chromatography,suchashydrophilicinteractionliquidchromatography(HILIC) andhydrophobicinteractionchromatographyAsavariationofnormalphase chromatography,HILICusesahydrophilicstationaryphase,waterand water-miscibleorganicsolventsasmobilephases,anditsapplicationtendsto increase.Itsadvantagesaremainlyreflectedinthefactthatitisespecially suitableforseparationofdrugsandmetabolitesandcompatibilitywithmass spectrometry
Thecurrentresearchprogressfocusesontheuseofmonolithiccolumnsor small-porepackingcolumnstoreducesampleanalysistimeandincrease samplethroughput,thatis,toachievethepurposeof"rapid"or "high-throughput"analysis.Inanultra-highpressureliquidchromatography (UPLC)systemthatcanwithstandabout1×108Pa,chromatographiccolumns
withaporesizeof15and3mmcanbeused,whichismoresuitableforrapid analysisofcomplexdrugsamples.However,underultra-highpressure conditions,thenonlinearheatingandretentioneffectsintroducedbythe compressibilityoftheliquidcannotbeignored,andspecialattentionshouldbe paidtothispointduringmethoddevelopment.
Inaddition,theintroductionofasecondorganicadditiveintothemobilephase, usedasion-pairingreagents,micelles,orsilanolmaskingreagentsforthe separationofextremelyacidicandextremelybasicsubstanceshasalso receivedattentioninHPLCmethoddevelopmentRP-HPLCcanalsobeused tostudytheencapsulationandreleaseofactiveingredientsinnanoparticles. Forexample,theRP-HPLCmethodwith01%trifluoroaceticacidasamobile phasecanbeusedtocharacterizethebindingandreleaseefficienciesof alginate-chitosannanoparticlesoninsulininasimulatedgastrointestinal environment.Therearealsoreportsthat2DLC(suchasnormal-phasediol narrow-borecolumn-reversed-phasemonolithiccolumn)and multi-dimensionalLCareusedtoseparateandanalyzedrugmixturesor extracts,whichcaneffectivelyimprovepeakcapacityThenew chromatographictechnologyalsoincludesthedevelopmentofvariousnew molecularlyimprintedchromatographiccolumnsandthecolumnswitching technologyininvivodruganalysis,whichisusedtoachievethepurposeof fastpre-columnswitchingpurification.
IntermsofdetectorsapplicabletoHPLCtechnology,theapplicationof evaporativelightscatteringdetectors(ELSD)inHPLCtendstoincrease, especiallyfortheanalysisofvariousantibiotics,naturalproductsand phosphateestersOtherdetectorsusedincludedifferentialdetectors, fluorescence,chemiluminescenceandelectrochemicaldetectors,andcharged aerosoldetectors(CAD)ELSD,CADanddifferentialdetectorsareall universaldetectors,generallyusedforthedetectionofsubstancescontaining noorlittleultravioletchromophoresComparedwithELSD,CADhasthe characteristicsofhighersensitivity,widelinearrangeupto4ordersof magnitude,andconstantsignalresponsefactorThesetwodetectorsare mass-dependentdetectors,whichgenerateuniversalresponsesignals. Therefore,thecalibrationcurveobtainedbyusingareferencesubstanceis alsoapplicabletootherdrugsandtheirimpurities.Thisoffersthepossibilityof accuratequantitationintheabsenceofcontrolsorradiolabeledcompounds ThecommondisadvantageofCADandELSDisthatthemobilephase compositionaffectsthedetectionsignal,whichcanbesolvedbyaddinga mobilephasewiththeoppositegradientcompositiontocompensate