Settings for Trace analysis in GC?

[Tremenda]

Dear all,

I have a small peak whose intensity I want to increase.
There is a small problem though:
There is a peak a 100 times larger that elutes a few seconds earlier.
They do not overlap, but they are close.

I can think of the following options:
-Increase injection volume
-Modify Split ratio
-make a splitless injection

I could also preconcentrate the sample...

Any idea that could avoid the overlapping?

Thanks

[GasMan]

Tremenda,

Doing what you suggest will only make the situation worse. You must first separate the two peaks more than they are at the moment. To help you, people will need much more information. Please post the following

1. The names of the two compounds you are tring to separate.

2. The column you are using, a full description.

3. The GC conditions you are using, mainly oven program, column flow rates, carrier gas type, and inlet conditions.

Ideally, it is better to have the small peak elute before the large peak, but this is not always possible.

Are there other peaks in your mixture, as giving advice on separating the two peaks that you mention may make matters worse for other peaks in your sample.

I am more of a hardware person, but maybe somebody with more column knowledge will be able to help, but they will need the information that I mention above.

Once you have better separation, then you can think about injecting more sample.

Gasman

[aldehyde]

Yes, please give us more information about the method parameters and what you are trying to separate.

Your best bet is going to be modifying oven program, but please give us more info.

[Tremenda]

GC: Varian 3400

The column is a Chrompack CP-Sil 5 CB (100% Dimethylpolysiloxane)

Length: 15 m

ID: 0.25 mm

Film thickness: 0.25 um

Headpressure: 6 psi

Linear velocity: approx. 20 cm/s (This figure seems quite high for such a small flow, this might be because we are using a second-hand column and we are not sure about the length)

Column Flow: 0.72 ml/min

Gas: Nitrogen

Split ratio: 30:1

Sample size: 0.2 ul

Temperature program:

5 min @ 40 ºC

30 ºC/min up to 200 ºC


The compound is a derivatised carboxylate. TMS-carboxylate.


Is the change of column the only possibility then?

[chromatographer1]

Your solution as implied by others is a longer column or a column with a different selectivity.

You can also use a heartcut valve or Deans Switch to reduce the first peak in size while keeping the second peak at the same size.

best wishes,

Rodney George

[Tremenda]

Right so...what is the minimum time between a large peak and a trace peak in order to be able to improve its intensity?

If I manage to separate enough the peaks, what can I do then to improve the intensity of a trace peak?

Regards

[chromatographer1]

Time between peaks?

That is a difficult question and is dependent upon the peak widths of the peaks involved. I can't give you a specific answer. It must be determined experimentally.

I can tell you that it is possible to have a small peak hidden on either side of a peak and still be able to separate it clearly from the heartcut ramp of the primary peak.

The easiest solution is to have a better selectivity. This again depends upon the analytes involved. Double the length of a column and the # of plates increase by a factor of square root of 2, ie 1.4 or so, the column length has its limits for separation improvement.

A heartcut valve is the best solution unless you can find a different selectivity which works for your situation.

best wishes,

Rod

[Peter Apps]

Hi Tremenda

All of your suggestions are quick and easy to do and do not involve any irrevocable hardware changes - so try them. You only have a problem if they do not work.

I would also try a slower temperature programme - 10C/min is closer to the optimum for your column length. Why so long at 40C ?

Peter

[aldehyde]

Even though column length gets to be diminishing returns a 30 m column could help. Decrease the oven ramp speed a little as well.

By the way, I plugged your numbers into Agilent's flow calculator and am getting essentially the same numbers:

15 x 0.25 x 0.25 @ 0.73 mL/min N2 gives me a velocity of 20 though the P calculated is 3.9 PSI.

[Tremenda]

Thanks for your replies.

I followed Chromatographer1's advice and I used a 50 m 0.32 mm column.
The difference between the solvent peak (eluting first) and the trace peak I want to magnify is approximately 4 min.
Is this a great separation?

I get such separation by running an isothermal cycle at 80 ºC (The solvent elutes at 10 min).

However, the lower the temperature, the better the separation but the less sharp peaks are. Could I compensate such effect by increasing the head pressure?

Is it really important that the solvent elutes first or as early as possible?


I think now I could concentrate on trying to increase the intensity of the trace peak. What are your thoughts about this?

Thanks

[aldehyde]

The order in which the elute doesn't matter, what matters is that they are completely resolved (separated). The solvent peak probably tails a little, is the tail gone by the time the peak of interest elutes? How much time after the tail is gone does your peak elute?

You could start from a low temperature and then ramp the oven temperature in order to sharpen the peaks a bit, try 10 degrees a minute and play with it depending on your results.

[JTM]

Now that they are seperated, you want a lower detection limit?

Anything that can more analyte through the detector... splitless (or lowering the split ratio) injection would be #1 for me.

[Tremenda]

Thanks

How is the LOD determined?
What is minimum the peak-to-noise ratio?

My target peak has not a sharp gaussian shape. Instead it is like the mirror image of a fronting peak (when the column is overloaded).

I found this equivalent question in the LC forum, but I do not know how much it applies to my case

Thanks

I am doing a separation of TFA and Acetate on a reversed phase column, and I notice that as I increase the amount injected of either compound the peak becomes increasingly right-triangular. At lower concentrations/injection volumes, the peaks are fairly gaussian, but as I increase, the front end becomes more and more vertical, and the back end becomes more and more sloped (a linear slope, not really a typical tail). Also, retention seems to decrease as the peak shape becomes more right-triangular. However, despite the changes, linearity is still very good.

Does anyone know what causes such a change in peak shape? Can anyone recommend a good resource that might give some explainations for various changes in peak shape and how they could be interpreted?

Thanks.

[chromatographer1]

You then have a tailing peak and this indicates that the peak is seeing active sites which affect its shape and possibly you may be 'losing' some of it on the column or injection port.

Increasing the oven temperature will sharpen the peak but make it elute more closely to the solvent peak.

Increasing the sample size may help or may hurt your detection limit for the analyte whose limit of detection you should determine experimentally.

best wishes,

Rodney George