Chromatography Forum

Hi,

I'm new here, and I have an issue with GC.

I'm working on an alcoholic beverage, and I need to have a standard curve to determine how much ethanol and methanol is in the beverage.

Well, I prepared standard as follow;

45% ethanol, 0.05% methanol and the rest is water.
40% ethanol, 0.1% methanol and the rest is water.
35% ethanol, 0.15% methanol and the rest is water
(ethanol is 100%, methanol is 99.99% pure)

Agilent 6890 GC with an autosampler.
HP-5 column, 30m.
Inlet temp: 175C
Oven temp: 40C for 3 mins, then to 80C by 10C increase, wait 3mins, then to 120C by 10C increase, wait 1min.
Split ratio: 40:1
Injection volume: 1 ul

I have irreproducible areas even from the same vial. Let's say, I have 4500 area, then 15000. and vice versa.

What would be the problem having such fluctuation?
Thank you.

Hi!

Have you ever test the linearity of this concentrations of methanol? Because some detectors might not answer so well if the concentrations are so high... Maybe you can put an split next to 1:100...

If I use this concentration of ethanol, I can't measure the area of the peak... because is too much for my detector... The detector can't even design the top of the peak correctly... there is just a horizontal line...

What kind of beverage do you have?

I hope this is helpful!

Hi Kaliane,

Thanks for the reply.

I have beautiful seperation, and peaks shape are great. Only problem is the area fluctuating especially for ethanol.

I checked the septum, liner, column, everything seems OK.

I do not know what to do else

Hi!

Have you ever test the linearity of this concentrations of methanol? Because some detectors might not answer so well if the concentrations are so high... Maybe you can put an split next to 1:100...

If I use this concentration of ethanol, I can't measure the area of the peak... because is too much for my detector... The detector can't even design the top of the peak correctly... there is just a horizontal line...

What kind of beverage do you have?

I hope this is helpful!

Did you change your autosampler syringe recently ? If not , try to replace it with a teflon tipped syringe.

I'd agree too much ethanol in that standard. Your dynamic range is also very tight for ethanol. I'd look for a range in the order of 100x. Try running 100ppm to 10000 ppm in water. Methanol would be a minor target I assume, so run that at maybe 5-500 ppm. Also I'd run a few extra points, five is my minimum, and many methods require it.

We had a similar problem and went the opposite way that uzman recommended, went from teflon to non-teflon tip on the syringe and it worked. So whatever you have on there, try the opposite I guess and see if it helps.

Another thing to check might be your syringe uptake speed and viscosity delay to make sure you are not dealing with air bubbles or inconsistencies that might be caused if the sample is not able to be consistent within the syringe under your current parameters

Water really expands a lot when injected and vaporizes.

We validated ethanol in our hand sanitizers after dissolving samples and standards in water, and we never injected more than 0.5µl.

Also, HP-5 is not very polar; you might want to consider a significantly more polar column for these polar molecules.

Hi,

Thank you everybody trying to help, I really appreciated.

I changed the column. Now, I have DB-17 ((we do not have any other polar(mid-polar) column), we only have HP-5, HP-88, and DB-17) the fluctuation problem is gone but I have a new problem

It does not separate ethanol and methanol. Any suggestions?

Thank you all.