Chromatography Forum

I get low response for methanol & ethanol in residual solvent test even when I switch to the highest FID sensitivity mode.when I have methanol std with conc 150 μg per vial the area is not more than 7151. There is some information may give you a clue.
thank you

instrument : Shimadzu 17 A ver 3 + CombiPal

headspace condition: vial 20 ml --- Incubation Temperature 80 (°C)
Incubation Time 3600 (s) ---- Syringe Temperature 85 (°C)
Agitator Speed (rpm) 500 ------- Fill Speed 200 (μl/s)
Injection Speed 500 (μl/s)

GC codition: column RTX 1301 0.53mm X 30 m df=3μm
Column Oven : 40(°C) Inj. Port:140(°C) Detector : 250(°C)
Temp. Program > Column Oven
1-Init. 40 (°C) for 20 min
2-10(C/min) 240 (°C) for 20 min
Total Time (min): 60
[ Flow ]
< Parameter > CAR1
(1) Control Mode : Split
(2) Column Pressure (kPa) : 25
(3) Column Flow (ml/min) : 5.09108
(4) Linear Velocity (cm/s) : 34.5295
(5) Total Flow (ml) : 16
(6) Split Ratio (1:X) : 2

I love to see info provided. THANK YOU.

However:

150 micrograms !!!!!!!!

WOW !

60 Minutes heating !!!!!!!!!

WOW !

Now the most important piece of information is missing.

Now if I only knew the volume of HS sample that was injected ! You are splitting whatever you are injecting 2:1, but of how much volume?

Or perhaps you are losing solvent with leaking vials? Easy to do. Hard to see it happen when it does except for low peak areas. Hmmm sounds familiar.

Good luck,

Rod

It's so nice that I can finally contribute something helpful to somebody.

The magic word when troubleshooting the combi-pal in headspace mode is displacement.

Two months of pulling my hair out can be boiled down to that one word. The other 2 styles of headspace injection do not have to worry about this. But the combi-pal takes a volume and puts it all in one place (injector) that is already occupied (by carrier gas).

What is your inlet liner? 1mm ID or 3.5mm ID? The syringe injection functions better with a larger ID and a gooseneck on the column inlet end.

Injection speed may also be something to consider. The 16ml/min total flow converts to 267µL/s. If you would happen to be injecting 1000µL of headspace at 500µL/s, my guess is that the dynamics inside the injector can become skewed.

Is there a septum purge flow? I believe in headspace situations that flow can be turned off.


Hope this helps.

As Charles has noted, performing HS injection with a split injector can be full of possible errors which can prove confusing if not confounding for the inexperienced.

I left that technique behind when doing my HS work. I found direct injection (not splitless, not split) was the best way. The only complication is getting a good column inlet seal from the injection source and making certain the injector volume was not overfilled (keeping the injection volume per min below the carrier gas rate flow). While syringe injection bypasses the patents and some other issues, the direct (timed) injection of PE and the sample loop injection (Tekmar and Agilent) made my life less complicated.

Unless one configures their injection port properly according to the method of injection bad results can occur.

Thoughtful planning is recommended, with an emphasis on thoughtful.

Remember, if it were easy, anyone could do it.

best wishes,

Rod

It's so nice that I can finally contribute something helpful to somebody.

The magic word when troubleshooting the combi-pal in headspace mode is displacement.

Two months of pulling my hair out can be boiled down to that one word. The other 2 styles of headspace injection do not have to worry about this. But the combi-pal takes a volume and puts it all in one place (injector) that is already occupied (by carrier gas).

What is your inlet liner? 1mm ID or 3.5mm ID? The syringe injection functions better with a larger ID and a gooseneck on the column inlet end.

Injection speed may also be something to consider. The 16ml/min total flow converts to 267µL/s. If you would happen to be injecting 1000µL of headspace at 500µL/s, my guess is that the dynamics inside the injector can become skewed.

Is there a septum purge flow? I believe in headspace situations that flow can be turned off.


Hope this helps.

Can you elaborate.be more specific on this? I have done a lot of headspace (actually mostly JUST headspace GC) but it has always been via loop style headspace sampler...

Although I have no experience with them, I am getting a Combi-Pal for a GC that I will need to do both liquid and Headspace on. I went that way as it was cheaper than separate liquid and headspace samplers and took less space (which was also a big consideration).

I really need to understand how i need to adjust headspace methods for a syringe based injection. And i will be working with a splt/splitless injector.

Thanks,
- Karen

Karen,

Just as with a liquid injection, you can overfill the space in the injector when you inject a gas sample so the sample can pass through the septum purge, backflash into the gas pneumatics, and condense the sample in cooler spots, or inject too much too quickly into too large a volume where you lose the 'plug' of sample. The sample can then become diffuse and 'smear'. This is why a narrow bore glass injection liner is used for SPME fiber injection and for HS transfer lines which terminate with a needle through a septum in a conventional injection port.

If you inject 1cc in 2 seconds you can overwhelm the injection liner with sample. This can smear peaks or split them. In any case your peak sizes will be small and often non-reproducible. If you inject too slowly then your peaks can be broad and poorly formed.

Discuss this problem with your vendor and let them guide you as far as syringe volume and injection speed so you will not waste time learning the 'hard' way. Combi-Pal autosampler should be able to perform routine HS analysis but different parameters than those used with a fixed loop or timed injection HS analyzer may be required to achieve the desired results.

Charles may wish to add his well earned experience to this discussion. I would welcome his comments to my sparse contribution here.

best wishes,

Rod

I love to see info provided. THANK YOU.

However:

150 micrograms !!!!!!!!

WOW !

60 Minutes heating !!!!!!!!!

WOW !

Now the most important piece of information is missing.

Now if I only knew the volume of HS sample that was injected ! You are splitting whatever you are injecting 2:1, but of how much volume?

Or perhaps you are losing solvent with leaking vials? Easy to do. Hard to see it happen when it does except for low peak areas. Hmmm sounds familiar.

Good luck,

Rod

Do not be surprised. Uncritically stolen screening methods "to residual solvents" from your USP. Brain no it

Although I have no experience with them, I am getting a Combi-Pal for a GC that I will need to do both liquid and Headspace on. I went that way as it was cheaper than separate liquid and headspace samplers and took less space (which was also a big consideration).

I really need to understand how i need to adjust headspace methods for a syringe based injection. And i will be working with a splt/splitless injector.

Thanks,
- Karen

This reminds me of my experiences buying a house for the first time. You know exactly what questions you should have asked 6 months after you've moved in.

What is the make up of your yearly sample load? Since you've already stated that most of it is headspace, were you changing loop sizes often on your fixed loop apparatus?

I'd be cautious regarding the savings in money. The combi-pal is like an inkjet printer. The basic models are essentially given away because they know they'll make a killing on the ink cartridges you'll need to buy.

If the majority of your injection sizes are between 500 and 2000 µl, you are in good shape, if you need to go outside that range then it gets expensive. Usually the system comes stock with 1 syringe heater meant for a 2.5 ml headspace syringes. However, there are 1ml and 5ml syringe sizes that need their own block heater. Those are pricey.

Do you change temperatureof your vial heater and consequently your transfer line? Ideally, you would have dedicated syringes for different temperatures. For instance, USP <467> calls for 80°C vials. The syringe would need to be at 85°C. But if you have a vendor/in-house method that required a 105°C vial temp, then 110°C would be the recommended syringe temp. In this range of temperature, the teflon is significantly changed because it's malleable and would expand when heated. The chances for a bad seal would be highly likely if you went back to the original temps first mentioned.

Be sure to have extra syringes available especially for the honeymoon phase with the instrument. There are no second chances with a zee'd needle.

Screw cap vials seem to be preferred over the standard crimp caps. May be more expensive.

Other questions to ask yourself when choosing this system, how active are the compounds you intend analyze? I could tell you just a fraction of what makes this an important question, but Rodney's the man to ask about that subject. Mainly because he's where I got the fraction to begin with.

Each style of HS analyzer comes with good and bad issues. Personally, I'm more comfortable with fixed loops and pressure balanced systems. I had to do a trial by fire with the combi-pal and it was very trying, but I did gain a lot of knowledge.

Anything else, just let me know.

Chuck

This reminds me of my experiences buying a house for the first time. You know exactly what questions you should have asked 6 months after you've moved in.

Well let's just say if I knew then what I do now about my house I would have not made an offer on it!

What is the make up of your yearly sample load?

This will be for an R&D instrument to help figure out oddball stuff connected to an APGC interface to a QToF, as well as as a backup to the fixed loop HS-GC for FID assays (it also has an FID). I think it will be primarily for liquid injections without too much headspace workm (unless the fixed loop instrument goes down or we get a lot more samples than usual for that assay)

The fixed looped instrument does about 75-300 injections/week and occasionally more, with about 150/week being typical.

Since you've already stated that most of it is headspace, were you changing loop sizes often on your fixed loop apparatus?

The fixed loop instrument is always 1 mL and I'm doing a split injection with all the carrier flow routed through the headspace transfer line and split ratio of 5:1.

BTW this is a relatively new job. At my last company the GC work was exclusively headspace (various residual solvents from a lot of different matrices) and as i had multiple instruments I rarely has to change loops.

In this job I'm not doing residual solvents.

I'd be cautious regarding the savings in money. The combi-pal is like an inkjet printer. The basic models are essentially given away because they know they'll make a killing on the ink cartridges you'll need to buy.

Yikes!

If the majority of your injection sizes are between 500 and 2000 µl, you are in good shape, if you need to go outside that range then it gets expensive. Usually the system comes stock with 1 syringe heater meant for a 2.5 ml headspace syringes. However, there are 1ml and 5ml syringe sizes that need their own block heater. Those are pricey.

In the routine assay I'm doing I'm looking at high concentrations (.1% to 20% ) so I will be able to inject in that range as long as I can do a split.

For the other stuff I don't know yet.

Do you change temperatureof your vial heater and consequently your transfer line? Ideally, you would have dedicated syringes for different temperatures. For instance, USP <467> calls for 80°C vials. The syringe would need to be at 85°C. But if you have a vendor/in-house method that required a 105°C vial temp, then 110°C would be the recommended syringe temp. In this range of temperature, the teflon is significantly changed because it's malleable and would expand when heated. The chances for a bad seal would be highly likely if you went back to the original temps first mentioned.

How 'different' is different? is 10°C too different?

Sigh... looks like I made the wrong decision, as I don't know what I'll need other than for the routine assay. With the loop instrument one does not need to worry about such things. I suppose I can get and keep a few syringes for specific temperature ranges.

Be sure to have extra syringes available especially for the honeymoon phase with the instrument. There are no second chances with a zee'd needle.

OK.

Screw cap vials seem to be preferred over the standard crimp caps. May be more expensive.

Do you know why? Seems to me, it should not matter and that screw-cap would be more likely to leak as they could be sealed less reproducibly. For my fixed loop instruments, I aways use crimped vials because of that.

Other questions to ask yourself when choosing this system, how active are the compounds you intend analyze?

Although not the main targets, in the routine assay i do quantitate some short chain fatty acids... the levels are high enough that they are not a problem with the loop sampler.

Mainly because he's where I got the fraction to begin with.. For unknowns ... I don't know!

I had to do a trial by fire with the combi-pal and it was very trying, but I did gain a lot of knowledge.

Are you still using it?

BTW too I am very comfortable with the loop sampler as I have been using them for 16 years with relatively few problems and little maintenance beyond yearly PMs.

As I said I chose the Combi-PAL for flexibility, space savings and being able to use a single sampler.

Anything else, just let me know.

Thanks I appreciate it.

It will likely be several weeks before I start using it as it has not been installed yet and I am going on vacation soon.... but I'm sure I will have more questions once I get started using it!

- Karen

Hi,
1.Examine inject splitless.
2.Change your vial crimp headspace.
3.Change your septum.
4.Change your liner and use of a goseneck liner(little space).
5.Change speed injection to 1000.
6.Cut 10cm of your column and backwash with carriar gas.
7.Reinstall column and check again distance from injector.
8.Incrase first temp. to 50 and incrase flow to 8ml/min and see changes.
9.Spike your peak with MeOH and EtOH and see,See have a change in your chromatogrph?
if do this work and don't any change your detector is dirty...
Good Luck .

Reply to Karen's posts:

We use an A200S for headspace work - we have modified it to heat the syringe, but depending on the analyte that probably is not necessary. A CombiPal is the Lexus equivalent of my Toyota Corolla A200S.

MS presents certain challenges using headspace that you don't find with other detectors. You have pumping capacity issues, and you have vacuum chromatography. You have to weigh both issues when setting up methods. We use an iterative approach to method creation, guided by some horse sense and a health dose of pigheadedness.

In general, the lower the column flow the better off you are, so we use small diameter (0.18) columns with flows around 0.5 ml/min. This dictates the use of split injections, which in my opinion are the only kind of injections you should ever use with gas phase samples. You need to try to maintain laminar flow - this is very, very contraintuitive! You will find that increasing the split ratio will actually decrease the detection limit - at first it blows your mind. I must have run 100 experiments trying to determine where I screwed up before I learned that this is just the way gas phase works. In your case you have a LOT of analyte, so DL is not critical (it is in my work). That means that you can work with truly high split ratios, which will make your headspace work extremely reproducible.

You also need to use internal standards - matrix has a huge impact on headspace (partial pressures are a bear to calculate in the presence of unknown analytes). Salting out is a good option - it not only increases the concentration of the analyte in the headspace but it also damps the matrix effect of the sample significantly. Additionally, you want to work in a polar solvent - water is ideal - and you want to work at relatively low concentrations (mid-ppb to mid-ppm), so you'll probably end up diluting your specific samples. This is good - makes matrix matching much easier. We use 200 uL of a saturated salt solution with 800 uL of aqueous sample in a 2 mL screw-cap autosampler vial, and then inject 50 uL of the headspace. We're getting sub-ppb detection limits for benzene, with RSDs of 20-30% at 2 ppb.

Happy to provide you with more "don'ts" if you need more. From your posts you're too good of a chromatographer to make some of the boneheaded mistakes we made, so I think you'll figure it out pretty quickly.

Mark Krause
Austin Analytical, LLC
Austin, TX
mckrause@austinanalytical.com