VALVES
141
APPLICATIONNOTE
FluidicMovement inTubes
Q:
“Why can I loadonlyup tohalf of the volumeof the loop
inpartial-fillingmethod?”
A:
Sampleoccupies2µLof loop for every1µL loaded from the syringe.
For example, 10µLof sample spreadsout over theentire lengthof a
20µL loop.Anyadditional sample loadedwill overflow theendof the
loopandexit out towaste. Reproducibility ispoorbecause the
volumeof sample in the loop isdifferent from theknown volume
originally loadedby your syringe.
Fluid spreads inaparabolic shape througha tube insteadofmoving in
oneplugbecause the velocity isdifferent at thecenter of the tube than
at thewalls. The velocityat thecenter of the tube is twice theaverage
velocity, andnear thewall the velocity isalmost zero, creatingaparabolic
shape. Thisfluidicmovement is called laminar flow. SeeFigure1.
Indualmode injection
valves (see“SampleLoop
Loading”ApplicationNote
onpage129) thesample
from thesyringeneedle loads
directly into thesample loop.
Thesamplevolume isknown
since there isnosamplewaste.
The laminarflowphenomenon
accounts for theshapeof the
plotasshown inFigure2.Note
that theplothas three regions:
a) Partial-FillingRegion.When
thevolumedispensed is less than
half the loopvolume, thecurve
is linear.Samplehasnot reached
theendof the loop.Within this
region,performancedependson
thesyringeandoperator.
b)NonlinearRegion.When the
volumedispensed isbetween
half the loopvolumeandabout two loopvolumes, thecurve isnonlinear.
Sample is lost from the loop, so reproducibility ispoor. If youdispensea
volumeequal to the loopsize, youare in this regionofpoorperformance.
c)Complete-FillingRegion.When thevolumeof sampledispensed isseveral
loopvolumes, the loopcontainsonlypuresample, undilutedby residual
mobilephase.Within this region, reproducibility ishighest.
In thesinglemode injectionvalves thesamplemustpass throughaconnecting
passagebefore it reaches thesample loop. Sincesomeof thesample
dispensed from thesyringe remains in theconnectingpassageway, an
unknownamountenters thesample loop. Therefore, singlemode injection
valvesachievehigh reproducibilityonlybyusing thecomplete-fillingmethod.
APPLICATIONNOTE
How toFindandFixCommonSample Injector Leaks
Leaks cause valuable sample loss.Nobodywants that. Thekey to the
valveholdingpressure is the integrityof the sealing surfaces. If there is
a scratchon the sealing surface, or theneedle seal in the rotor seal is
damaged, a leakmayappear. It isalso important to realizewhat appears
tobea leakcan insteadbea result of siphoning. The followingare the
threemost common situations inwhichfluid leaksoccur.
1. If fluid leaksoutof theneedle
portonlywhile loading the loop
(i.e.,whilepushingdownon the
plungerof the syringe), theproblem
ismost likely that theneedle seal or
theneedleport fitting in the loop
fillerport isnotgripping the syringe
needle tightlyenough. Tighten the
needle seal gripbypushingwith
theeraserendof thepencil on the
needleport (SeeFigure1). The
tightening reduces theholediameter
of theneedle seal andport fitting.
2. If fluid leakscontinuously from theneedleportor vent linesand/or from
the stator-to-stator ring interface, replace the rotor seal and/or stator face
assembly. Scratcheson the rotor seal or cracks in the stator faceassembly
allowmobilephase toescapeandcausecrossport leakage.Genuine
Rheodyne replacement rotor sealsare listedonpage137.
3. If fluid leaks from theneedleport and/or vent linesbut eventually
stops, thecause ismost likely siphoningandnot a leak. Siphoningoccurs
if the vent linesare lower or higher than theneedleport.Adjust the vent
line(s) so that theoutlet isat the samehorizontal level as theneedleport
toprevent siphoning. (SeeFigure2).
Forother leakageor injection troubleshooting, refer to theRheodyne
TroubleshootingGuide forHPLC InjectionProblems. Youmaydownload
theGuide from theRheodyneweb site:www.idex-hs.comunderSupport.
Figure2
Needleport level compared to the level of vent lineoutlet:
(A) siphoningoccurswhen the vent lineoutlet is above theneedleport level
(B) siphoningdoes not occur if the vent lineoutlet is the samehorizontal level
as theneedleport
Sample
Flow
TubeWall
MobilePhase
Figure1
Schematicof sampleflow throughmobilephasebetween tubingwalls
Areaof Peak
SampleDispensed (loop volumes)
1
2
3
4
40
80
linear
nonlinear
constant
Figure2
Samplemass (observedpeakarea)
vs. volumeof sampledispensed from the
syringe, inunitsof loop volumes, injected
onto the column fromaRheodyne
®
dual
mode injector suchasmodel 7725
Figure1
To reform theneedle seal,
push theeraser endof apencil
against theneedleport
ApplicationNotes
