GC-MS: Peak Area and Hight increase over time

[Mr.Brown]

Same Standard Sample split to two vials and injected with roughly 20-25 injections (roughly 7-8 hours) in between
Same Method, same injector temp (275°C) same everything.
I get roughly 3 times higher response vor all the analytes of interest (tR 7 to 13).


(pickture is with base shift)
blue... empty organic solvent
black... Standard injection at the beginning
pink.... Standard injection at the end

The internal standard is doing his job and is compensating for this (more or less, still adds some deviation)
It should not be carryover as the empty solvent injection does not show any peaks for the analyts in question.

Any ideas?

If i would have the same respons at the beginning I could have much lower LOQ, LOD

[Peter Apps]

At what stage was the solvent blank injected ?

My crystal ball is at the cleaners - unless you post your analytical details before I get it back I would have to troubleshoot by guesswork.

Peter

[Mr.Brown]

Trying to analyse Beerepellents with GC-MS.
(Benzaldehyde, Phenol, p-Dichlorobenzene, Phenylacetaldehyde, Nitrobenzene, Naphthalene, Thymol)

blank organic solvent was injected right after Standard

Injection:
1 µL
Splitless
Sampling time: 0,75 min
HPI: 150 kPa for 0,75 min
Split 1:50
Injector:
Hot Splitless
Equil.-time: 5 sec
End time: 300 sec
Initial T: 275 °C
Final T: 275 °C
Carrier Gas
1 mL/min He
Oven:
40°C hold for 1 min
to 125°C at 10°C/min
to 170°C at 15°C/min
to 290°C at 30°C/min
hold for 4.5 min

MSD:
Interface: 290°C
Ione Source. 240°C
Solvent Cut 5 min
Voltage relative 0.2 kv
SCAN:
50 – 250 m/z (0.3 sec)
SIM: accordingly (0.2 sec)

[Peter Apps]

T

blank organic solvent was injected right after Standard

OK, but WHICH standard ? the first one or the second one ?

Peter

[Mr.Brown]

OK, but WHICH standard ? the first one or the second one ?Peter

after the second one at the end of the run

[Peter Apps]

OK

The peaks that might be solvent contaminants do not increase in area, but the analyte peaks do. Nonetheless they are a lot bigger than I would expect form good quality solvent.

If the increase was due to adsorptive sites getting progressively deactivated the analyte peaks would be tailed.

Evaporation of the solvent from the vial would increase both contaminant and analyte peaks.

Are these the compounds that you dissolved in methanol ? (from your other posts) ?, or are they in something different ? Is there a concentration step anywhere along the way ?

What happens if you eliminate solvent evaporation by doing the replicate injections from separate vials instead of all from one vial ?

How are you cleaning your syringe between injections, and when last did you replace (not top up) your cleaning solvent(s) ?

Peter

[Mr.Brown]

Hi Peter

I have split the sample in two separate vials so evaporation should not be an issue

only the first 2-3 peaks are from the solvent it selve the rest are some silanyls probably from the liner (for testing dirty samples I kept using it)
And on peak is dodecane I apperantly still have in the system.

The analyts are still the same Beerepellents just a differen, new problem i just recognized during the test runs.
The solvent in this case is t-BME (suprasolv vor GC).
Found a publication where they used for some of the beerepellents t-BME for extraction however they do not report any cleanup procedure
so i am testing SPE, dispersive SPE and so on.

The syringe is cleand with t-BME 2 times after injection, and one time bevore injection and 1 time with sample.
I replaced the cleaning solvent when i started with t-BME 2-3 weeks ago have not toped it up jet.

I looked at the internal standard of the samples meassured in between and it looks like the ISTD is increasing more or less linear with each injection.

[thohry]

Possibly, the vacuum gets better over time and the signals get higher.

[Mr.Brown]

hm
leak test and tuning look ok, no N2 no H2O, all parameters look ok

i do not see any fluctuation with the vacuum

by the way i am using a shimadzu QP2010 Ultra

In this method i am using splitless high pressure injection mode.
During method developement i recognized that with high pressure injection mode i get much nicer peaks and also increased hight and area
so it could also have something to do with the high pressure injection mode

[Peter Apps]

Hi Peter

I have split the sample in two separate vials so evaporation should not be an issue but your ares till doing multiple injections from each vial, with the last run several hours after the septum was first punctured.

only the first 2-3 peaks are from the solvent it selve the rest are some silanyls probably from the liner (for testing dirty samples I kept using it) The non-solvent contaminant peaks are remarkably repeatbale for inlet crud
And on peak is dodecane I apperantly still have in the system.

The analyts are still the same Beerepellents just a differen, new problem i just recognized during the test runs.
The solvent in this case is t-BME (suprasolv vor GC).
Found a publication where they used for some of the beerepellents t-BME for extraction however they do not report any cleanup procedure
so i am testing SPE, dispersive SPE and so on.

The syringe is cleand with t-BME 2 times after injection, and one time bevore injection and 1 time with sample.
I replaced the cleaning solvent when i started with t-BME 2-3 weeks ago have not toped it up jet. Solvent boiling point is 55,2 C, so more volatile than hexane, you might be getting evaporation from the vials. Try the series of injections form separate vials. 2 + 1 + 1 syringe rinses is nowhere near enough - I would double it at least. You should not let the cleaning solution sit for 2 - 3 weeks. Change it every day - how much trouble would that be ?. Never just top it up - contaminants accumulate. I suspect that the contaminants are in your wash vial, and that they are being progressively transferred to the standards with each operation of the incompletely rinsed syringe

I looked at the internal standard of the samples meassured in between and it looks like the ISTD is increasing more or less linear with each injection. this fits either solvent evaporation, or progressive contamination by dirty wash solvent

Peter

[Mr.Brown]

but your ares till doing multiple injections from each vial, with the last run several hours after the septum was first punctured.
Nop, seperate vials seperate septas i never inject from the same vial more than once

The non-solvent contaminant peaks are remarkably repeatbale for inlet crud
You are right they are remarkable repeatable

Solvent boiling point is 55,2 C, Try the series of injections form separate vials. 2 + 1 + 1 syringe rinses is nowhere near enough - I would double it at least. You should not let the cleaning solution sit for 2 - 3 weeks. Change it every day - how much trouble would that be ?. Never just top it up - contaminants accumulate. I suspect that the contaminants are in your wash vial, and that they are being progressively transferred to the standards with each operation of the incompletely rinsed syringe
I always inject from seperate vials; doing a series of injections is a good idea i will try that; thank you for the advice i will adjust the syringe cleaning method accordingly;
when i chang i never top it up i allways discard the old cleaning solution, but apperently i should increase the changing frequency, changing it daily sounds like a good idea

this fits either solvent evaporation, or progressive contamination by dirty wash solvent
while injection of empty organic solvent does give the same contaminant peaks I do not see any analyte peaks
so no carryover is happening that would suggest an increase of ISTD or analyt peaks

This is the cleaning procedure vor the Syringe
maybe you have some other helpfull suggestions
Air Volume [µL] 0
Pre Clean with Solvent 1 1
Pre Clean with Solvent 2 0
Pre Clean with Sample 2
Filling Volume [µL] 5
Filling Speed [µL/s] 2
Filling Strokes 5
Pullup Delay [ms] 5000
Injection to OPTIC 4
Injection Speed [µL/s] 100
Pre Injection Delay [ms] 1000
Post Injection Delay [ms] 1000
Post Clean with Solvent 1 3
Post Clean with Solvent 2 0

[Peter Apps]

but your ares till doing multiple injections from each vial, with the last run several hours after the septum was first punctured.
Nop, seperate vials seperate septas i never inject from the same vial more than oncequoting from your first post "split to two vials and injected with roughly 20-25 injections" which is 10 - 12 injections from each vial. Depending which of these two you are doing, solvent evaporation is either a plausible explanation or it is not

The non-solvent contaminant peaks are remarkably repeatbale for inlet crud
You are right they are remarkable repeatable

Solvent boiling point is 55,2 C, Try the series of injections form separate vials. 2 + 1 + 1 syringe rinses is nowhere near enough - I would double it at least. You should not let the cleaning solution sit for 2 - 3 weeks. Change it every day - how much trouble would that be ?. Never just top it up - contaminants accumulate. I suspect that the contaminants are in your wash vial, and that they are being progressively transferred to the standards with each operation of the incompletely rinsed syringe
I always inject from seperate vials see above; doing a series of injections is a good idea apparently you have already done it - see above i will try that; thank you for the advice i will adjust the syringe cleaning method accordingly;
when i chang i never top it up i allways discard the old cleaning solution, but apperently i should increase the changing frequency, changing it daily sounds like a good idea

this fits either solvent evaporation, or progressive contamination by dirty wash solvent
while injection of empty organic solvent does give the same contaminant peaks I do not see any analyte peaks
so no carryover is happening that would suggest an increase of ISTD or analyt peaks if you have done a series of injections from separate vials, and you still get an increase in peak area then it cannot be due to solvent evaporation. That more or less leaves progressive contamination of the wash solution by analyte due to inadequate syringe cleaning. If there was an increase in detector sensitivity then the small contaminant peaks would be getting bigger through the series as well

This is the cleaning procedure vor the Syringe
maybe you have some other helpfull suggestions
Air Volume [µL] 0
Pre Clean with Solvent 1 1 increase to 3
Pre Clean with Solvent 2 0
Pre Clean with Sample 2 increase to at least 4 - I presume that it is simply sucking up sample and discarding to waste - it must not pump the syringe at the cleaning stage or it will transfer wash solvent to the sample
Filling Volume [µL] 5
Filling Speed [µL/s] 2
Filling Strokes 5
Pullup Delay [ms] 5000
Injection to OPTIC 4
Injection Speed [µL/s] 100
Pre Injection Delay [ms] 1000
Post Injection Delay [ms] 1000
Post Clean with Solvent 1 3 increase to 6
Post Clean with Solvent 2 0

you cannot clean the syringe too much - especially when troubleshooting. /color]

Peter

[Mr.Brown]

Dear Peter

First of all thanks again for all your help and time your are giving to my problem.

quoting from your first post "split to two vials and injected with roughly 20-25 injections" which is 10 - 12 injections from each vial.
i think i have to clear up a missunderstanding. the 20-25 injections in between were other test samples and befor the test samples i injected the standard and after all the test samples i injected the same standard from a different vial.

Pre Clean with Sample 2 increase to at least 4 - I presume that it is simply sucking up sample and discarding to waste - it must not pump the syringe at the cleaning stage or it will transfer wash solvent to the sample
i hope so but yes

[Peter Apps]

The second injection being from a sealed vial rules out solvent evaporation (which was not likely to explain such large rises anyway).

Watch the autoinjector in operation to see what it does - maybe it has some operational quirk that makes it susceptible to this kind of problem.

Peter