Professional Documents
Culture Documents
INTRODUCTION
u Injector septum
u Injector temperature
u Sample vials
assembly
u Installing the fiber assembly into the
SPME holder.
injection
u Building an SPME method
TROUBLESHOOTING
SUPPLIES
REFERENCES
Figure 3. Injection unit in the “FiberExp” position Installation of the SPME adapter
with the right edge just touching the left rear edge of 1. Press “F1/Menu” and then F1/Chang Syr. The injection unit will
the agitator
move to a position that will facilitate installation of the SPME
adapter.
2. If the injection unit is directly over a sample tray, the “Chang
Syr” position should be moved.
3. Press “Continue” and then:
>Utilities
>Syringe
4. Press “F3/Set Pos” and set the x y z positions to a location where
there is a clear space under the fiber. Then Press “F4/Home” and
repeat Step 1.
5. Install the plunger holder into the injection unit (Figure 4 left).
6. Install a fiber assembly in the fiber holder and place it in the
SPME adapter (Figure 4 right).
Pull up the plunger so that the fiber is completely withdrawn
into the protective needle.
7. Place the SPME adapter, partially into the injection unit. In order
P001189
to do this, tilt the top of the SPME adapter foreword slightly
(Figure 5A) and thread the protective needle carefully through
Figure 4. Left: injector unit with plunger holder in- the upper and lower needle guides at the bottom of the injec-
stalled (A). The arrows marked “B” are pointing to the tion unit. This process is made easier by lifting the magnet so
upper and lower needle guides; Center: unassembled it is level with the needle guide, then push the needle through
kit components; Right: fiber holder and fiber assem- the opening (Figure 5B)
bly properly installed in the CombiPal SPME adapter 8. Push the plunger down so that approximately 1.5 to 2 cm of
the fiber and fiber support rod are exposed
9. Place the plunger crosspiece into the plunger holder. Allow the
syringe adapter to “click” into place by magnetic force, against
the syringe carrier.
10. Tighten the plunger retaining screw against the plunger cross-
piece (Figure 5C) and press
“Continue”.
P001190
P001191
P001192
Figures 5A, 5B and 5C Need titles Checking alignment of CombiPAL in the injectors
The PAL should have been initially aligned with the injection port
during setup. This accomplished without an needle in the injector.
The needle guide is used to position the unit. For SPME, the needle
alignment is very critical.
1. Place SPME adaptor container holder and assembly into the PAL
injector
2. Follow sequence in PAL
menu setup Injectors
3. Select appropriate injector and continue
4. The needle will puncture the injection port (septum) nut open-
ing
5. Using X, Y, Z positions, align the PAL injector so the needle is
P001193 straight in the x and y positions. This may require very slightly
alignment, but it will assure that the needle will properly hit the
center of the septum nut opening
P001194
Setting fiber depth in the sampling vial. 3. Check the fiber depth in the sample. For direct immersion in
The main parameter for controlling the depth into the vial is “vial liquid you want the entire fiber, except the top 1 mm, immersed
penetration”. Vial penetration is the distance the plunger moves in the liquid as shown in the photo.
after the needle pierces the vial. The distance that the needle pierces 4. Adjust the “vial penetration” setting in the software until the
a vial is fixed at 11 mm. fiber is in the proper position. This may take several attempts.
The depth will remain the same in the agitator.
Figure 7. Showing the “Vial Penetr” parameter 5. For headspace, you want to make sure that the fiber is not
touching the liquid and is centered in the headspace.
6. Follow step 4 until desired depth is obtained.
For most samples, heat will be applied. For liquid samples, agitation Building an SPME method
may be used along with heating. For solid samples, agitation may See the “CombiPAL System User Manual” for details on how to
not be necessary with heating of the sample. Macro “Test SPME build a method.
with Heat and No Agitation” can be used. Optimize heating and
time by designing multiple methods. First select a fixed time and The parameters in the SPME method are discussed below:
vary temperature. Typical temperature range is 35 °C to 70 °C.
Rarely, is the extraction temperature set above 70 °C, because the Parameter Value Comments
increased temperature will desorb analytes off the fiber and may drive Cycle SPME
more water into the fiber and needle sheath. After the optimum Syringe Fiber
temperature is obtained select appropriate extraction time. Keep in Allows the sample to be pre-
mind the total GC cycle time when setting extraction times. Pre Inc Time 00:00:00-23:59:59 heated prior to insertion of the
fiber.
Optional Bakeout of the fiber after injection “OFF” for “Sampling out of the
With some fibers, a high temperature is necessary to desorb the 30.0 °C –200 °C Tray” at ambient temperature
Incubat Temp
analytes completely. Often the GC injector cannot be set to a high or OFF without agitation. A maximum
of 80 °C is suggested.
enough temperature because a column with a low temperature
limit is installed. With the CombiPAL the user can bake the fiber Set “0s” to turn off agitation.
Agi On Time 0s - 99s
Used for “Sampling out of Tray”
after desorption in a separate bakeout station (Figure 8). This is an
optional piece of hardware. Agi Off Time 0s - 99s
Distance from top of vial septum
Vial Penetr 22.0 – 31.0 mm
Figure 8. Optional bake-out station. The baking occurs to end of fiber (Figure 7)
with a flow of inert purge gas Sampling time in liquid or
Extract Time 00:00:10-23:59:59
headspace
Normally an injector such as
Desorb to None - Waste 2 GC Inj1, GC Inj2 or “Front” or
“Rear” is entered here.
Distance from top of injector nut
Inj Penetr 44.0 mm – 67.0 mm
to end of fiber (Figure 6B)
Desorb Time 00:00:10-23:59:59 Time in injector
For baking out the fiber after de-
Fiber Bake- sorption in the optional bake-out
00:00:00-23:59:59
out station. If “0” , the unit does not
move to the bake-out Station.
Enter the complete GC cycle
time including cool-down and
GC Runtime 00:00:30-23:59:59
re-equilibration to coordinate the
CombiPAL and GC cycles
> Objects
> Injector
> NdlHeatr
0 2 4 6 8 10 12 14
Min G003780
Troubleshooting
Symptom Possible Cause Recommended Action
Verify (see above) that the bottom of the SPME fiber syringe is
Fiber breaks in injector Improper depth in injector
not less than 5 mm into the insert.
Septum corings or other particles are in the Replace insert. If septum particles are present, consider using a
Fiber breaks in injector
injector. seal such as the Merlin Microseal to eliminate the septum.
Verify that the cap can not be turned after sealing. Reduce the
Poor precision Vials are leaking extraction temperature to see if the precision improves.
Temperature > 80 °C are not recommended.
See the Advantage Note on SPME method development in the
Poor precision Poor sample handling.
“SPME Application and Advantage Note” section of this manual.
Increase desorption time and/or temperature or bake out the fiber
Sample carryover Fiber is not fully desorbed.
after each injection.
Fiber support rod is submerged in liquid Reduce the fiber penetration depth in vial or reduce the amount
Sample carryover
sample. of sample in the vial.
Verify that the GC is clean by making a run without injecting a
Extraneous peaks in blanks Contamination is in the GC.
sample.
Sample an empty vial without a septum installed.
Contamination is in the sample vial septa.
Sample an empty vial with a septum installed.
If the contamination is from the septum, bake the septum in a
laboratory oven at 150 °C overnight. This will minimize extrane-
ous peaks.
Extraneous peaks in blanks
Vial is not completely released from the needle when the injection
unit moves away. Replace short tension cord (PM)
2. Extraneous peaks in 1. Septa used in sampling vial or injection port is 1. Prebake the vial septa for 2 hours at 68°C
outgassing organic contaminants (S/A) prior to use. Use low-bleed LB-2 septa to
analysis minimize injection port septum bleed.
2. Fiber is not preconditioned prior to sampling (P) 2. Precondition fiber at the recommended
conditioning temperature in the fiber
instruction sheet. Once fiber is precondi-
tioned, only 1-2 minutes is required to clean
the fiber prior to sampling.
3. Inlet liner is contaminated or contains 3. Replace the inlet liner. Use pre-drilled
septa particles (D) septum or a septumless injector system (e.g.
Merlin Microseal)
4. GC column is collecting analytes on the front 4. Complete GC analysis temperature program
of the column because it is not heated high before injecting another SPME extract and
Normal enough in sample analysis (A) keep column at 150°C when not in use.
5. Interfering peaks coelute with analytes of 5. Change GC column or temperature program
interest (A) rate
6. Carryover from previous analysis of the fiber (P) 6. Bake out fiber at the recommended
conditions for several additional minutes
7. Cross-contamination from laboratory air (S) 7. Do not expose the fiber to the laboratory
environment at any time during the sampling
or injection steps. Analyze control blanks
using the same handling process as the
sample to determine if technique or
laboratory cross-contamination is present.
Problem 8. Cross-contamination during transport from the 8. Move the depth adjusting lever of the
field (S) portable field sampler to the top-most
locking slot, so the end of the septum-
piercing needle is totally withdrawn into the
sealing septum of the sampler. If the fiber
will be stored for more than one day we
recommend that it be stored at subambient
temperature. This reduces the chance of
sample cross-contamination.
1. The end of the needle is plugged with a piece 1. Injection port septum nut is overtightened.
3. Fiber will not retract or of septum (D) Loosen the nut slightly to allow for improved
sticks in holder needle injection. Use pre-drilled injection port septa
or a septumless injector system (e.g. Merlin
Microseal)
2. The fiber was exposed to solvents that caused 2. Do not expose PDMS coated fibers to non-
swelling of coating (S) polar solvents such as pentane, methylene
chloride, or diethyl ether. Do not expose
Carbowax fibers to polar solvents.
3. The top screw in the holder assembly is too 3. Loosen the top screw on the holder assembly
tight (P) slightly to allow for free movement of the
plunger.
4. Needle bends during 1. Improper manual sampling technique (S) 1. To prevent the needle from bending when
injection into sample vial doing manual SPME sampling, follow this
or GC injection port procedure:
Adjust the SPME needle to the 0.2 depth
gauge setting on the plunger (first tick
mark). This will expose about 3mm of the
needle through the end of the black holder.
Hold the SPME assembly on top of the
sampling vial with the bottom of the black
holder flush with the top of the vial cap.
Hold the sampling vial and black SPME
holder base securely with one hand
Twist the stainless steel plunger clockwise
with the other hand
Keep turning the plunger until the desired
depth setting is achieved (you will usually
hear a pop when the needle pierces the
septum).
Expose the fiber and perform sampling as
usual
2. Improper manual desorption (injection) 2. To prevent the needle from bending when
technique (D) doing manual SPME injections, follow this
procedure:
Adjust the SPME needle to the 0.2 depth
gauge setting on the plunger (first tick
mark). This will expose about 3mm of the
needle through the end of the black holder.
Position the SPME assembly on top of the
GC injection port or in the SPME inlet guide
with the bottom of the black holder flush
with the top of the injector or guide.
Hold the black SPME holder base securely
with one hand
Twist the stainless steel plunger clockwise
with the other hand
Keep turning the plunger until the desired
depth setting is achieved (you will usually
hear a pop when the needle pierces the
septum).
Expose the fiber and perform sample
desorption as usual
3. Vial or injection port septum is too tight 3. Loosen slightly the vial closure or injection
(S/D) port nut
4. Septa in sample vial/injection port is too thick 4. Use LB-2 septa for injection port or silicone
or coated with thick Teflon® coating (S/D) septa with <10mil Teflon on sampling vials.
Shorten the amount of exposed needle on
SPME holder to 0.5 inch or (~1cm) before
puncturing the vial septa. Adjust the holder
needle setting to the desired depth for
sampling. Do not use butyl rubber style septa.
5. GC inlet liner is too narrow or packed with 5. Use larger splitless inlet liners (0.75mm ID or
adsorbent material (D) larger) without glass wool or adsorbents.
Needle bends with automated injection systems 6. Needle is out of alignment with injection port 6. Reference autoinjector manual on alignment
or sample vial (S/D)
5. Fiber breaks 1. The fiber was not retracted into the protective 1. Retract fiber into protective needle during
needle after removal from sample vial or insertion into vial/injection port and removal
injection port (S/D)
2. The end of the needle is plugged with a piece 2. Injection port septum nut is overtightened.
of septum (D) Loosen the nut slightly to allow for improved
injection. Use pre-drilled injection port septa
or a septumless injector system (e.g. Merlin
Microseal)
1. Time and temperature variations during 1. Time of extraction and temperature are the
6. Reproducibility is poor sampling (S) two most critical conditions to control. Use
timing device and calibrated thermometer to
ensure reproducible results. Remember that
room temperature fluctuations will influence
the ambient sample temperature.
2. Not consistently positioning the fiber at the 2. Position fiber just below sample surface for
same depth during sampling (S) immersion sampling and at a consistent
position above the sample during headspace
sampling.
3. pH or salt conditions varying during sampling 3. Apply uniformly across all extractions any pH
(S) or salt adjustments made to the samples.
4. Equilibrium is not reached during extraction 4. Determine minimum time for equilibrium
(S) using a standard mixture and controlled
extraction conditions. Note that full equilib-
Normal
rium is not required to be reached for all
applications to achieve reproducible results.
5. Varying organic content in the samples (S) 5. Dilute samples to minimize solvent interfer-
ence or use headspace sampling to minimize
solvent effect. Use internal standards,
surrogates, or the standard addition
technique to compensate for variations in
sample matrix.
6. Varying headspace in sample vials during 6. Minimize headspace volume to 50% or less
headspace extraction (S) and agitate the sample. Maintain the same
headspace volume and agitation conditions
Problem across all extractions.
7. Solid samples not releasing analytes for 7. Grind solid into small particles, add to water,
extraction (S) and apply heat and agitation.
8. Competing analyte displaces compound of 8. Reduce the extraction time to minimize
interest/or interferes (S) displacement/or interference
9. Not reproducing desorption conditions (D) 9. Verify that the fiber position (depth),
desportion time, temperature, and splitless
conditions are consistent. Use an automated
SPME system to improve reproducibility.
10. Not using agitation during sampling or apply it 10. Use a stir bar or soncication system to agitate
inconsistently (S) the sample during sampling. Maintain
consistent agitation conditions for all
standards and samples.
11. Sample volumes are inconsistent (S) 11. Maintain consistent volumes for all standards
and samples.
7. Fiber discolored 1. Fiber is oxidized during fiber conditioning or 1. Minimize oxygen in carrier gas, condition
sample injection into GC (S/D) fibers in oxygen free gas flow. Reduce the
injection port temperature to the recom-
mended maximum setting. Carbowax/DVB
coatings are especially sensitive to tempera-
ture (<260°C is recommended).
2. Heating during injection (D/P) 2. Does not usually affect the performance of
the fiber. Always minimize the oxygen
content in the GC carrier gas to avoid
oxidizing the fiber coating. The polyacrylate
coated fiber will discolor above 280°C.
Carbowax/DVB may slightly darken during
use, however, if the fiber turns brown, lower
the injection port temperature (265°C is the
recommended maximum) and check the
system for leaks.
8. Number of injections from 1. Fiber is oxidized during fiber conditioning or 1. Minimize oxygen in carrier gas, condition
the fiber is less than sample injection into GC (S/D) fibers in oxygen free gas flow. Reduce
previously obtained injection port temperature to the recom-
mended fiber maximum.
2. Coating on fiber deteriorated (P) 2. Replace fiber. Fibers are reusable and will
last for 50 injections on average.
3. Fiber was exposed to solvents that cause 3. Do not expose PDMS coated fibers to non-
swelling of coating (S) polar solvents such as pentane, methylene
chloride, or diethyl ether. Do not expose
Carbowax fibers to polar solvents.
A=Analysis related D=Desportion related P=Product related S=Sampling related
SUPPLIES REFERENCES
Sampling Vials for CombiPAL Headspace Useful Web pages:
Based on studies involving Supelco SPME fiber durability with the Supelco (SPME literature) http://www.sigma-aldrich.com/supelco
CombiPAL system It is important that thinner septa be used to seal University of Waterloo (SPME research) http://spme.uwaterloo.ca/,
vials for SPME use. We recommend a 1.5 mm silicone PTFE lined
septa with SPME neck vials or screw cap vials. A steel cap should be University of Texas (SPME bibliography) http://www.cm.utexas.
used with screw top vials and either aluminum/tin or steel can be edu/~brodbelt/spme_refs.html
used with the SPME neck vial. A 1 mm thick Viton septa works best
BOOKS
with screw cap vials. For SPME use, we strongly recommend that 1. Pawliszyn, J., Solid Phase Microextraction: Theory and Practice, Wiley-VCH, Inc,
caps with a 8 mm hole be used instead of the 5 mm hole. New York, 1997
2. Wercinski, S. ed, Solid Phase Microextraction: a Practical Guide, Marcel Dekker,
New York, 1999
Description Color Qty. Cat. No.
Screw neck Vial and closure SPME APPLICATIONS
10 mL round bottom Clear 100 SU860099
10 mL round bottom Amber 100 SU860100 SPME Literature and References
20 mL round bottom Clear 100 SU860097 (add picture of CD marked as
20 mL round bottom Amber 100 SU860098 6th edition)
Steel Magnetic Cap 8mm Hole
With 1.3 mm thick silicone blue/PTFE white septa 100 SU860101
Applications of SPME are con-
With 1.5 mm thick silicone blue/PTFE white septa 100 SU860103 tained on the newly released
With 1.6 mm thick butyl red/PTFE gray septa 100 SU860102 6th edition of the SPME CD.
SPME flat neck Vial and closure This CD is a compilation of
20 mL round bottom Clear 100 SU860104 literature references, applica-
Steal Magnetic Cap, Gold, 8mm Hole tion notes, and helpful hints
With 1.5 mm thick silicone white/PTFE blue septa 100 SU860053 to using SPME.
With 1.0 mm thick Viton black septa 100 SU860106
20 mm x 1.5 mm thick silicone /PTFE septa 100 27541 Over 600+ new literature
20 mm x 0.75 mm thick Viton septa 100 27247 references have been added to the Application Guide (total
Crimper of >2200) and is easily searched by analyte or sample ma-
Hand crimper, for 20 mm crimp seals 22316-U trix. The SPME CD is presented in Adobe Acrobat format for
easy review and access. Request a copy of the SPME CD at
Pre-Drilled Thermogreen LB-2 Septa for SPME the Sigma-Aldrich web site at www.sigmaaldrich.com/Brands/
Easier needle penetration and high puncture tolerance – ideal for Supelco_Home or call Supelco at 814-359-3041
autosamplers. Reduce septum coring that can cause extraneous
peaks. Already conditioned, ready-to-use. Extremely low bleed over
a wide range of inlet temperatures – from 100 °C to 350 °C. Rubber
formulation exclusive to Supelco.
Description Cat. No.
9.5 mm, pk. of 25 23161
9.5 mm, pk. of 50 23162-U
11 mm, pk. of 25 23167
11 mm, pk. of 50 23168