Ethanol in wine - liner issues

Discussions about GC and other "gas phase" separation techniques.

12 posts Page 1 of 1
Hi all,

Apologies for the long message

We are determining ethanol in wine using GC.

We have done this lab for many years but I have never found the peak shapes to be great so decided to try to improve things.

The standards are made up in water to be between 0.1% to 0.5% (v/v) ethanol with 0.5% 1-propanol as internal standard.

I inject 0.2 ul (to avoid the well-known water expansions issues) using a 10 ul syringe. Would like to inject more but that's the joy of water. Also don't want to switch from water to another solvent.

Conditions are
Inlet 250C, split 100:1. pressure 14 psi
Column is HP-Innowax 30m x 320 um x 0.25 um.
Flow is 2.9 ml/min (46 cm/sec)
Oven is 75C isothermal
but is left overnight at 150C to flush column and possibly prevent build up of water on the column.
Detector is FID
Agilent Autosampler.
Analyte peaks elute within 2 minutes

A standard containing (0.3% ethanol and 0.5% 1-propanol) was used throughout these tests.

Results
Using a deactivated liner with glass wool such as the Agilent 5183-4647.
Precision of peak areas expressed (as %CV) around 1% for 10 repeats
Peak shapes show a lot of tailing with USP tailing factors (at 5%) > 1.5 and get worse as more injections are done.

Using a liner without glass wool
Precision of peak areas (expressed as %CV) around 25% for 10 repeats. Area ratio precision is good though - %CV is around 3% for 10 repeats
Peak shapes are excellent with USP tailing factor (at 5%) consistently close to 1.
- this was also the case when I used a Agilent 5183-4647 liner that I had removed the glass wool from.

In conclusion if I have a liner without glass wool I get great peak shape but low peak precision while if I have a liner with glass wool I get tailing peaks but much better peak precision.

Any suggestions what I might be able to try to obtain the Holly Grail of good peak shape and good peak area precision

Am I missing anything obvious?
To try to solve this issue, I replaced the gold seal, and split vent trap recently and also have trimmed the column. It seems though that the liner is the issue.

Thanks
Kevin
We validated our in-house procedure for ethanol for cGMP/FDA. We have used both ACN and 1-propanol as internal standards.

I'm not going to say you're doing anything "wrong", but will point out some differences you might want to try.

We used a 15m long x 0.53mm id column as can tolerate larger expansion volumes of water.
We used a 1 microliter syringe as 10 microliter can have larger hold-up volume in the needle.
We used either a wax/PEG column or G43 (USP designation).
When we do aqueous injections on a 0.32mm column we have to stay at or below 1ul injection volume to prevent problems with water expansion. At 0.3-0.5% (3000-5000ppm) you should be able to see those with even a 0.5ul injection, though may possibly have to reduce the split flow to 50:1 or 30:1, but the peak shapes may improve enough to negate any reduction in mass on column.

You may also want to try one of the cyclo liners like these https://www.restek.com/en/products/acce ... lies/9219/ to get more surface area without the holdup of the broken ends of the glass wool which cause the tailing with that liner.
The past is there to guide us into the future, not to dwell in.
Thanks Consumer Product Guy and James Ball for your replies.
I will definitely look into those Restek liners.

Consumer Products Guy - I have a question for you. How do you keep
1 microlitre syringes functioning. We have tried 5 microlitre syringes for this application so we could inject 0.5 microlitres rather than 0.2 ul but we find that the lifetime of a syringe is very short as the plunger gets stuck after 20-30 injections typically. We had use water as the rinse solvent and then tried methanol without success. Is there a solvent you would recommend here


Thanks
Kevin
I went back and checked: we injected 0.5 microliters from either a 1 microliter or a 5 microliter syringe. The Agilent autosampler was set to rinse the syringe with water between injections.

Our samples were hand sanitizer and other consumer products. For the stickiness issue, you could try diluting the samples and standards further with water once the internal standard has been added, since that ratio between the two has already been established then.
thank you for your response again Consumer Product Guy. I thought that using water as the rinse solvent for the syringe was what caused the sticking issues with the syringes. I guess not given your experiences.
Kevin.
I'm curious. Tossing aside your desire not to use another solvent for just a moment....

Why don't people use DMSO? Say a 10:1 dilution with DMSO and a 10:1 split?

Best regards,

AICMM
You might try CSR-LVSI as found at RESTEK. I've taken down my alcohol system for a few years now but I was using a 200um DB624 column with 5 meters of 0.53 inert column and a 1uL injection. These were oilfield brines. I was opening the purge valve pretty early to dump the water and I was using glass wool liners. I was trying to do methanol and glycols but ethanol and propanol were in my standard and seemed to have good enough peak shape. I was using dimethoxyether as the internal standard. for the alcohols.

There is a post over on the GC thread that gives a method for ethanol.
AICMM wrote:
Why don't people use DMSO?


1. Solvent cost

2. Solvent disposal cost

3. Environment concerns of above
RE the first post. It sounds like you are injecting the wine straight. There are plenty of sparingly soluble solids in wine that may be baking into the inlet. If ethanol is what you are after; I would try diluting your sample say 5X or 10X. then inject 0.2mL and try a lower inlet temperature say 200C to spare your column the higher temperature compounds. Also, you might try running a split. If wine has ~15% alcohol you have plenty of signal to play around. Water is probably the best solvent for cleaning up the syringe. I was injecting oilfield brines and I had trouble with my 5mL syringe as you did and went back to the 10uL Diluting the sample helped for me. Spill some wine and let it dry; water picks it up better than alcohol.

I think I was using the Tropez 4.0mm ID single taper with glass wool.
LALman wrote:
If wine has ~15% alcohol you have plenty of signal to play around.


Yes, dilute a lot, plenty of ethanol molecules.
Hi everyone
I wanted to update the group with developments. Thanks for all the advice which has helped enormously. I think I have come to a point where the analysis works well enough for the purpose (in a teaching lab).

To clarify, the samples are diluted wine samples. The wine is diluted by a factor of around 33x (3 ml in 100 ml). This gives us ethanol levels that fit within the calibration range of 0.1 to 0.5 % ethanol.

I have tried the cyclo type liner referred to in one of the replies and linked to (https://www.restek.com/en/products/acce ... lies/9219/). This is an improvement as explained below

Using this liner, the RSD for 10 injections is around 8% for both the ethanol and 1-propanol peak areas using 0.5ul injection from a 5 ul syringe. The RSD for the area ratio is less than 1% as expected
I can live with that as the peak shape is excellent and consistent.

I used a 100:1 split for this data. Interestingly I tried a 20:1 split and found that there was almost no increase in signal at all.

The other things that seem to have helped are 1) Use methanol to rinse the syringe 2) use a viscosity delay of 5 seconds when filling the syringe.

Take care
Kevin
12 posts Page 1 of 1

Who is online

In total there is 1 user online :: 0 registered, 0 hidden and 1 guest (based on users active over the past 5 minutes)
Most users ever online was 1117 on Mon Jan 31, 2022 2:50 pm

Users browsing this forum: No registered users and 1 guest

Latest Blog Posts from Separation Science

Separation Science offers free learning from the experts covering methods, applications, webinars, eSeminars, videos, tutorials for users of liquid chromatography, gas chromatography, mass spectrometry, sample preparation and related analytical techniques.

Subscribe to our eNewsletter with daily, weekly or monthly updates: Food & Beverage, Environmental, (Bio)Pharmaceutical, Bioclinical, Liquid Chromatography, Gas Chromatography and Mass Spectrometry.

Liquid Chromatography

Gas Chromatography

Mass Spectrometry