Headspace injection precision

[christenseng]

Any ideas as to what could cause a static headspace injection to produce peak areas roughly 60-80% higher than the other injections in the series?

For example, in a 6-injection series of a std. solution containing 12 residual solvents, 5 out of the 6 injections will be right on top of each other with %RSDs <5 for all analytes. The odd injection will have peak areas up to 80% higher than the other five.

Replacing the syringe seemed to cure the issue for now, but I can't understand how a leaky syringe could cause a high injection.

We're using a CombiPal autosampler and Agilent 7890.

[chromatographer1]

A partially plugged syringe when filling with multiple pumps doesn't purge out the sample each time but compresses it as it slowly leaves the syringe, causing a higher pressure within the syringe, condensing the solvents on the syringe walls. Then when injecting the sample the concentration in the last filling of the syringe was more concentrated than the headspace itself.

Rod

[christenseng]

If you are referring to multiple filling strokes from the same vial then your idea would not apply since I don't perform multiple pumps - one draw up and then right to the inlet. I always use one vial for one injection.

Interesting scenario though.

[chromatographer1]

Then there is something absorbing the analytes until a breakthrough point is reached in your flow path. With a syringe injection the only place would appear to be in the injector or at the head of the column.

The other possibility is that there was an error in making the vial with the higher than normal levels seen.

What other reason can you imagine?

best wishes,

Rod

OK, I just thought of one. If the higher vial was sealed more tightly than the others and the other leaked consistently a little (40% or so) then the results could be caused by this.

[Peter Apps]

Any ideas as to what could cause a static headspace injection to produce peak areas roughly 60-80% higher than the other injections in the series?

For example, in a 6-injection series of a std. solution containing 12 residual solvents, 5 out of the 6 injections will be right on top of each other with %RSDs <5 for all analytes. The odd injection will have peak areas up to 80% higher than the other five.

Replacing the syringe seemed to cure the issue for now, but I can't understand how a leaky syringe could cause a high injection but a leak inevitable causes low values - and an inconsistent leak can cause low values in the majority of runs, leaving the "true" leak-free values as apparent high outliers .

We're using a CombiPal autosampler and Agilent 7890.

If this is the same problem as you posted in your other thread then all the suggestions there also still apply.

Look for patterns in when the high values occur - first of a batch ?, last of a batch ? etc

Peter

[christenseng]

Thanks for all for the suggestions. I never like shooting down ideas unless I have something constructive to contribute but...

There is no pattern to the high injections - it can be anywhere within a run.

I'm using the same std. solution sub-sampled in all vials so even if the std. were not made properly, the areas for std. injections should still be the same. The error can show up using class A glass or a piston-type disposable pipet to transfer the solution to the vials.

If something in the inlet were adsorbing analytes then I would expect peak shapes to be affected. Chromatography is unremarkable. No carryover in blanks or empty vial injections.

I have leak-checked the inlet and am careful to replace the septum before use. If there is a constant leak somewhere, I can't locate it. And the number of "low" injections so far outweigh the high injections that I find it very difficult to believe that the "low" injections are all faulty.

Please keep the ideas coming!

[chromatographer1]

You said you replaced the syringe and that fixed it temporarily?

The problem seems to lie in the crimping of the vials.

You are not getting a consistent (good) seal. Perhaps, the vials are misshapen or the septa are not sealing well. Over tightening as well as under tightening can result in the vials leaking while the vials are heating.

The first step in doing HS is determining your sample prep is consistent. You do that by running a series of standards, just like what you are doing. If there is variation you have proven there is an issue in the vial preparation step.

Now you have the proof. Now find what is causing the problem.

Good luck,

Rod

[Peter Apps]

To check whether the vials are consistently sealed, prepare a batch of them as usual, then immerse them in hot water - if there is a leak you will see bubbles.

To take this any further we need full details of your operating conditions.

Peter

[christenseng]

Yes - replacing the syringe (actually only the plunger) seems to have alleviated the problem. I'm now keeping a careful count of the number of injections made with the new plunger to see if the problem appears again after so many injections. LEAP recommends replacing at approx. 500 injections regardless.

We are using screw-top vials so no crimping involved. I always check to make sure the septum is seated properly in the cap before screwing on the cap. As I'm sealing the vials by hand obviously I can't claim that the exact same torque was used to seal each and every vial. I'm careful to not over-tighten as I know that can deform the septum and cause leaks the same as an under-tightened cap.

I considered leaks in the vials early on but I always went back to the basic question - how can a leaky vial cause an increase in peak area?

Instrument conditions:

Column DB-624, 30 m x 0.32 mm i.d., 1.8 µm film
Column Flow Rate 1.5 mL/min
Carrier Gas Helium
FID Parameters
FID Detector Temperature 300C
Hydrogen Flow Rate 30 mL/min
Air flow rate 300 mL/min
Helium Flow Rate (Makeup) 28 mL/min

Inlet Parameters
Inlet Temperature 300C
Split Ratio 5:1

Headspace Parameters
Syringe Temperature 110 C
Syringe Flush Time 90 seconds
Injection Volume 1000 µL
GC Cycle Time 17.2 minutes
Plunger Fill Speed 100 µL/sec
Fill Strokes 0
Viscosity Delay 1 second
Pre Injection Delay 0.5 seconds
Plunger Injection Speed 500 µL/sec
Post Injection Delay 1 second
Sample Incubation Temperature 100C

Incubation Time 10 minutes
Incubation RPM 500 RPM
Agitator ON 5 seconds
Agitator OFF 2 seconds

Diluent - 5% ammonium hydroxide (30% soln) in NMP

[Peter Apps]

Questions:

what is NMP and what is its boiling point ?

do you really need the inlet to be at 300 C ?

What kind of GC is it ?

What inlet liner do you use ?

Have you checked the vials for leak tightness ? - faith is a poor guide in technical troubleshooting.

Peter

[Peter Apps]

Another question:

What is the sample volume ?

And an experiment: if, as I suggested, the high outliers are due to a small minority of vials being properly sealed (NB that this does NOT imply that a leak causes a high value) the puzzle is that the low values are so repeatable (it is unusual for something to go wrong repeatedly). Nonetheless, if the vials are leaking down to atmospheric pressure (which is perfectly possible) you will still get repeatable peak areas. So run a batch with the caps screwed only just tight enough to seat the septum but not to seal it, and see whether the peak areas match your low values, and how repeatable they are.

Peter