2/5/2017 HowtooperateHPLC

HowtooperateHPLC(HighPerformanceLiquidChromatography)

1.Turnontheswitchesofthepumpanddetector.

2.Turnonthecomputerfirstandthenprinter.

3.Theword"HewlettPackardSystemHardwareTest"willbeshownonmonitorandthatmeansthecomputer
ischeckingthesystemforexamplesharddisk.Ifthecomputerdoesnotfindanyproblems,themonitorwill
showtheword"Pass".

4.Next,type"C:\>win"togetintotheHPLCsystem.

5.Theword"HPChemistation"willappearsonthemonitorandthendoubleclickat"InstrumentonlineHPLC
system".

6.Themonitorwillshowtheword"HPLCsystem"whichgivesyou"peak"(whenthesampleisinjected),the
flowrateofmobilephaseorsolvent,wavelengthandpressure.

7.Forthepump,clickthebutton"pumpon"andthen"enter".

8.Forthedetector,clickthebutton"lampon"andthen"enter".Afterthat,adjustthemagnitudeofwavelength
asyouprefer.

9.Adjusttheconditionofthepumpbykeyinginthevaluestothecomputerattheicon"theInstrument"and
"Setuppump".

9.1Adjusttheflowrateofmobilephasestartingfrom0.1ml/min

9.2Selecttheproperkindofsolventtobethemobilephaseforexamples

A.Acetonitrile

B.Water

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2/5/2017 HowtooperateHPLC

C:Aceticacid

D.Methanol

Themobilephasecanbemixedtogether,forexampleB10%,D90%

10.Graduallyincreasetheflowrateofthemobilephasebythestepof0.1ml/min.Asthetimepasses,the
pressureincreases.Whentheword"Ready"appearsonthemonitor,thepressureisconstant.Thenincreasethe
flowrateofmobilephaseonemorestep.Repeatthesamewayuntilreachingthepreferedflowrate.Waituntilthe
baselinesteadythenpreparetoinjectthesample.

11.CleanthesyringewiththesamplebeforeinjectingthesampleintoHPLC.Theninjectandloadthesample
intoHPLC,thecomponentsofthesamplewillbedetectedbythedetectorandthetimeiscounted.

12.Peakofeachcomponentwillbeshownonthemonitoratthedifferentretentiontime.Whenthereareno
morepeaksdetected,pressF8buttontostopdetectingthesample.

13.Computerwillcalculateandintegratetheareaofeachpeak.Thentheresultswillbepresentedintherelation
betweenareaandtheretentiontime.Peaksandotherresultscanbeviewedbyclickingatthefollowingicons
"View","Dataanalysis","Loadsignal"and"Integrationresult".

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