Split Injector Misbehaving

[Jana P]

Hi All

I am having some issues with too much of my sample being injected onto the column.

I am using an Agilent 6890N GC system with G1888 Headspace Sampler. I was running my method with no problems until I last set up the method.

The most recent analysis I performed resulted in too much of my sample being injected onto the column giving me broader peaks (fronting) and subsequently a loss of resolution (between acetonitrile and dichloromethane).

My GC is controlled via Empower software and my instrument method had not changed. I use a split ratio of 5:1.

After investigating the problem I found that if I increase the split ratio to 10:1 my peaks appear to be better shaped and more resolved (resolution is still not as good as it originally was, but it getting there).

Note: during the investigation of this problem I discovered that there had previously been a leak in the helium line (possible contamination??). Also I have changed H2O/Oxy traps etc. and used two different columns as well as fresh and old solutions etc.

Can anyone tell me what would cause my chromatography to go down hill and require me to adjust my split ratio to achieve resolution on a method I have run many times before without having to change my split ratio?

If anyone has any ideas your thoughts would all be appreciated

Thanks in advance…

My method parameters are as follows:

GC Operating Parameters
Carrier Gas Helium
Flow or Linear Velocity 2.2 mL/min or 36 cm/sec
Split Ratio 1:5
Initial Temperature (1) 40°C
Initial Hold Time (1) 20 min
Temperature Ramp Rate 10°C/min
Maximum Temperature (2) 240°C
Hold Time (2) 20 min
Injection Temperature 140°C
Detector Temperature 250°C
Sampling Rate 5
Data Sensitivity HIGH
Pneumatics Mode Constant Flow
Fuel Flow 30 mL/min
Oxidizer Flow 450 mL/min
Make-up Mode Constant Makeup + Column
Make up/Combo Flow 30 mL/min
Gas Saver Flow Rate 15.0
Gas Saver On Time 1.50
Inlet Back
Detector FID

Headspace Parameters
Equilibration Temperature 80°C
Vial Equilibration Time 45 min
Transfer Line Temperature 105°C
Loop Temperature 105°C
Carrier Gas Helium
Pressurization Time 0.2 min
Injection Time or Volume 1.00 min or 1.0 mL
Loop Fill Time 0.2 min
Loop Equilibration Time 0.2 min
GC Cycle Time 60 min
Shake HIGH

[Jana P]

whoops...thought i should add that i am using the Volatiles inlet

[krickos]

Hi

What do you dissolve your samples and standards in?

If dissolved in anything else than water, how does the sample solvent peak look like, between "good" and "bad" runs?

If this is a validated method that has worked for some time, you should not have to change split ratio, potentially it is some contamination and/or build up.

How does the RSD % look like?

[Jana P]

Hi

We disolve our standards in water.

Would the contamination most likely be in the volatiles inlet itself??

RSD does not appear to have been affected.

[bhuvfe]

Maybe a restriction/obstruction in the split vent line?

[krickos]

Hi

I am a bit puzzelled as your peak areas seems to have increased. Normally when you have a contamination issue in the headsampler of this type (needle/valve/loop), peak shape may deterioate and peak areas rather decrease (interaction with residues), not increase.

On the other hand if that leak (less sample injected?!) has been there for a while it might not be surprisning that the areas increase.

If it is contamination is is not likely the volatiles inlet rather the needle/valve/loop, for instance if you previoulsy have injected samples dissolved in DMAA/DMSO/DMF/DMI with needle/valve/loop/transfer line temperatures far below these sample solvents boiling points you may have a build up of residues of these in your system.

[Jana P]

Hi all

Thanks for your input to date.

I ran a few injections yesterday and i have noted that the precision (%RSD Peak Area) is now very poor for any peaks which elute after approximately 7.4 minutes (this is about 8 of my peaks, all with resonable peak areas). Any of the early eluting peaks have a satisfactory %RSD Peak Area (worst being 3.7%).

Retention times are good and more than satisfactory in terms of %RSD across injections.

Does this indicate something else may be happening?

[Peter Apps]

Can you post a good and bad chromatogram (instructions are in a sticky at the top of the LC page) - doubling a split ratio is not likely to be enough of a difference to solve concentration overloading, which causes fronting, and it could be that you have volume overloading - the peaks are wider but no higher.

Are the headspacer-GC connection, and volatiles interface to column connection permanent, or do you re-make the connections each time you set up the method ?

Where was the leak ?, upstream of the instrument or between headspacer and GC ?

What are the analytes, and what column (phase, length and diameter) are you running them on.

Peter

[gpronger]

Did anything specific occur between the last previous "good" run and the later where it appears too much sample is being delivered?

In reading the symptoms, it sounds as if a leak which was causing you to effectively be operating at a higher split ratio than you thought was corrected somehow.

[Jana P]

Hi all

I have issues trying to post chromatograms the security on our computers doesn't like it very much

In regards to Peter's questions:

The headspace and GC connection have to be moved each time I use the instrument, the Headspace is always connected but I am required to move it from being connected to the split/splitless inlet to the volatiles inlet. So the connections are remade/moved each time I use the system.

The leak was of Helium and occurred at the gas bottle and a little further down the line at a safety valve, both well before any connection with the instrument. i could not identify or detect any leaks else where.

I am using a Restek column, RTx-624 column, 30m x 0.32, 1.8mm film thickness. I am separating methanol, acetonitrile, dichloromethane, trans-1,2-dichloroethene, cis-1,2-dichloroethene, THF, cyclohexane, methylcyclohexane, 1,4-dioxane, toluene, chlorobenzene, ethyl benzene, m-xylene/p-xylene and o-xylene.

I have used two separate columns now and the problem is the same on both.

I have been able to resolve the issue as in I can now obtain resolution between acetonitrile and dichloromethane if I increase the split ratio to 15 (from 5) however my peaks are getting smaller and this does not explain why a split ratio of 5 was satisfactory for so long and all of a sudden is no longer acceptable when nothing obvious has been altered with the method. Leaving me to ponder why our Agilent instruments are doing this…

[krickos]

In regards to Peter's questions:

The headspace and GC connection have to be moved each time I use the instrument, the Headspace is always connected but I am required to move it from being connected to the split/splitless inlet to the volatiles inlet. So the connections are remade/moved each time I use the system.

Excatly which connection are we talking about? The whole thing? I am kind of used that you permanently have the volatiles inlet installed at the back injector (plug it if not in use) and use the front injector for split/splitless unless you have a standalone headspace. Meaning that you do not switch connenction (apart from maybe gas supply if you do not control that via EPCs) between headspace/autoinjector applications.

Also one have to handle/protect the voilatile inlet from "dirt" the volume is only like 30-40ul if remebering right so if you get anything in there....


I see a couple of issues with your methodology, not sure all are related to your problems, but here it is:

A. When you use a pressured loop headspace like Agilents one got to learn that you MUST keep needle/valve/loop/transferline temperatures ABOVE the highest boiling point of the solvents (including sample solvent). You have several solvents with boiling points above your settings, it is like asking for a contamination/interaction with surfaceas issues in the long run.

A possible reason as it seems like most of your higher boiliers has poorer precision than your more voilatile solvents.

B. Water as sample solvent. It is great and cheap but have some drawbacks and considerations.
1. You try to avoid setting the headspace oven temperature above the solvents boilingpoints, pratically the solvents may start to boil if to high and cause instable gasphases. In your case I would not go above 60°. But hard to judge as you have a large mix of solvents and several interactions can occour.
2. Generally I do not favour preparing non polar solvent standards in water, it is like asking for reproducibility problems. When having solvents like toluene, xylens etc it is better to do it like in some pharmacopieas with regard to water solouble samples ie prepare standards in DMF (better stability as bonus), add 0,5-1,0ml DMF based standard to standard vials then water, to samples/blanks 0,5-1,0ml pure DMF then water/samplesolution (just make sure the DMF/water ratio is same in all vials).
3. Also at 80-90° the water vapour in the gasphase tends to impact precision negatively.



Lastly:

Loop Equilibration Time 0.2 min

Thats 12s for a loop of 1ml(?). For normal 1ml residualsolvents that is to long, just increases interaction risk with loop surface. I always keep that short (0,05min) for standard residual solvent analysis.

[Peter Apps]

Hi Jana

If you are making and breaking connections each time you run the method I suspect that previously you had a leak that was giving you an effective split ratio of 10 - 15 to 1. This time you have no leak, and you have to adjust the split settings to get the desired peak size.

With the Agilent volatiles interface in default configuration the split ratio is potentially vulnerable to the position of the column tip, because the internal volume is very small there is a potential for incomplete mixing of gas streams.

Have a careful look at the peak heights as well as the areas - do they correlate with the set split ratio ?

Peter

[larkl]

One thought for longer term consideration. We too have a headspace sampler that we need to use on both inlets/columns on a GC. We added a 6 port valve (a 4 port would have worked, but didn't have one) so we can use the headspace on either side without having to do any plumbing modifications.