Chromatography Forum

I am currently setting up the Agilent 5975 GC-MS to detect trace pharmaceuticals in waste water/soils- I can inject the same sample over and over and the consistency is HORRENDOUs-- carrying from several hundred thousand in abundance- this is the same vial of sample just injected 10x for 10 different chromatograms- I'm working with Agilent on it- I'm new to the system so I am just looking for an outside opinion- Agilent is confident it's a method optimization issue- and we've changed gain and threshold values to no avail...any thoughts?

Tell us about the rest of your system, column, gc, temp program, concentrator? etc and what analytes you are testing for and we will have a better shot at helping you.

Jon

30 m HP-5 5 %phenyl methlplysiloxane column
2 ul splitless 1:10 injection
Inject port 290 deg
Ms 230 deg
quafrpole 150 deg
Temp pgm 70 hold 2 min ramp 10/min to 250 hold for 5 min
contstnat flow 1.3 ml/min
looking for naproxen, ibuprofen, aspirin, clofibrate, caffeine, etc.....SPE extract with water derivitze 30ul MSTFA 35 min at 65 deg....

Any help?

First "1:10" is probably meant split 1:10 which is contradiction to Your statement - there is splitless injection without any split for certain time or split injection with some ratio.

So - check if Your method setup is correct - meaning You have really set up the splitless injection. Valve opening time is important- You do not mention it.

I think I found the article You are trying to reproduce. They had injector temperature 250C, You have 290C. Why?
High temperature will increase solvent vapour pressure and might have some unwanted effects.

As you say that the consistancy varies and discuss intensity. Do I take it that one run is very intense from end to end and another is not intense from end to end?

When the intensity varies from run to run, it is often a problem with being able to get a reproducable quantity of material on the column from run to run. So my first two questions are: have you checked for leaks, particularly around the inlet? And, when was the last time you replaced the septurm?

Leaks at the seal plate or the ferrule holding the column into the inlet can change from run to run as a result of the heating and cooling of the GC oven. An inlet septum that has had too many injections begins to leak as the needle wears a hole in the septum - again this is variable from run to run - for a while, then the worn spot in the septum becomes an open hole (yes, I've encountered this).

If the ratios of compunds are changing from run to run - this could be a derivitization issue. To separate sample from derivitization: If you inject a mixture that does not need to be derivatized - like a pesticides mixture - is the instrument consistant?

I have checked for leaks- and found nothing to indicate such. My auto tunes are all good.....I have changed the septum but I will put a new one in today again...The intensity varies so say run 1 is 400000 abundance run 2 might be 600000 run 3 300000 and run 4 500000 there is no pattern it's not like it's decreasing over the runs or anything it is a complete variable....Non derivitized samples do not give consistency...and the sample that is injected for runs 1-10 is out of the same vial anyway....I'm appreciating all the feed back- I will double check my method but I'm pretty sure is splitless mode.....And the paper I'm reproducing the method from is the Togola paper....

Thanks!

It sounds like an injection problem. It helps if you post detail splitless parameters. Make sure you check for leaks with leak detector. Try 1 ul to see if it helps. Inject a naphthalene standard and see if the responses are consistent.

Seems like a GC issue.

What kind of liner are you using and does it have sw? Your injection volume may be too high, try it with 0.5um and dropping the injection temp below 275

I'm also curious what solvent you are using. I assume that your carrier gas is helium?

What would happen if you loaded 10 vials with the same solution and injected once per vial?
Also depending on your solvent, liner, and inlet pressure you may be overloading the liner injecting 2ul.

So what is happening is I'm loosing my baseline more and more with each injection- I've put the injection volume down to 1 ul and threshold to 0 and am trying again- also adjusted over temperature up to 300 ..will let you know what I find out later today! Thanks!

SO now I"m not loosing my baseline- but my abundances are still varied...not consistant from same sample ....

I've see a 2 uL injection with MSTFA back flash into the plumbing and make a reall mess. And, if you have done enough of these injections, the purge vent line filter needs to be replaced.

I agree with the suggestion that you may have backflashed your injector. If you are using a 2mm ID liner and a 2 µL injection you almost certainly did. If you are not already, switch to a 4 mm liner if you are truely doing a 2 µL split-less injection.
We will be able to get more specific information if you do the following:

On your computer, Browse to your "method".M folder and open the text file acqmeth.txt. Copy the GC part and paste it here. Should look like this:
INSTRUMENT CONTROL PARAMETERS
-----------------------------

=============================================================================
6890 GC METHOD
=============================================================================

OVEN
Initial temp: 100 'C (On) Maximum temp: 350 'C
Initial time: 2.00 min Equilibration time: 0.50 min
Ramps:
# Rate Final temp Final time
1 15.00 150 0.00
2 2.50 270 7.00
3 0.0(Off)
Post temp: 0 'C
Post time: 0.00 min
Run time: 60.33 min

FRONT INLET (SPLIT/SPLITLESS) BACK INLET (SPLIT/SPLITLESS)
Mode: Pulsed Splitless Mode: Split
Initial temp: 275 'C (On) Initial temp: 250 'C (Off)
Pressure: 12.63 psi (On) Pressure: 0.00 psi (Off)
Pulse pressure: 25.0 psi Total flow: 45.0 mL/min
Pulse time: 1.50 min Gas saver: Off
Purge flow: 49.8 mL/min Gas type: Helium
Purge time: 1.50 min
Total flow: 53.5 mL/min
Gas saver: On
Saver flow: 20.0 mL/min
Saver time: 10.00 min
Gas type: Helium

COLUMN 1 COLUMN 2
Etc. etc... You get the point.

Ok...now I've removed the liner put in a new one without glass wool, removed 1 m of my column, new septum- injected and still this huge 91 mz mass spectra- which I know should be xylene/toluene solvent wash contamination- but the thing is - I've also removed my syringe- NO injections are going through the column- the spectra is from the carrier gas on the column only...PLEASE HELP!

Did you use xylene/toluene to clean the injection port? If so there may be some hanging in the split-vent (where it's cool and not heated). Is the 91 m/z as a background (through entire baseline) or is it as a large peak in the chromatogram?