Chromatography Forum

A little background. I developed a 5 analyte separation. Selectivity, resolution, precision are awesome when just testing the standard solution (no sample) containing the solvent DMSO and MeOH, EtOH,ACN, Chloroform and Pyridine. RSD% for all 5 compounds <1%.

Unfortunately I noticed some matrix effects when recovery with pyridine was consistently ~75% of expected when actually testing samples. The other 4 peaks are all +- 5% of expected.

I changed sample prep to standard addition. I thought this would be the ticket, but the linearity isn't good enough. I can't get better than r= 0.95

First question is is 0.95 a typically good enough linearity for a standard addition method?


I was considering trying to eliminate the matrix effect by switching to a total vaporization technique, when I encountered a question that seems trivial, but I can't find the answer for which is the purpose of this post.

Simply put for a given analyte amount at a given sealed volume (In this case it's 10ml vials), what's the minimum temperature at which the analyte will be totally in the vapor phase? It seems like this is a dumb question and a reasonable answer should be ~boiling point, but it seems like the building pressure in a sealed container will have an effect on vaporization. Is there an equation where this can be estimated?

"First question is is 0.95 a typically good enough linearity for a standard addition method?"

It would not meet my expectations, Mike.

Your problem probably lies in the acidic nature of your sample matrix.

Adding a base or a buffer to change the pH to improve the recovery of the pyridine can be problematic, especially with DMSO, but I would give it a try.

With 10mL vials you won't be able to add much DMSO for complete evaporation.

good luck,

Rod

I checked and we have some pKa data for this compound. In water a 1mg/ml solution has a pH of 0.9. Is there some established ideas to deal with this problem? You mentioned adding a base to sample matrix. How would this work ie. what base could I add?.

Have you ever dealt with this problem and how did you solve it?

Much appreciation
Mike. H.

A pH of 0.9 ! Wow, you are NOT kidding around. That is ACIDIC !

You can try bases like potassium (sodium) hydroxide, tetra methyl (tetra butyl) ammonium hydroxide (TMAH or TBAH), and carbonate salts.

Try to use aqueous based solutions of your matrix and get that drug at a neutral or slightly basic pH. You might have issues where you have to do a separate determination of pyridine by means of headspace. It is always nice to do all your solvents with one vial and dissolution solvent, but sometimes you have to do what you have to do.

I have used carbonate salts and metal hydroxides to achieve pH modification and dissolution. NaOH will dissolve almost everything to a degree, but I would not mix it with DMSO.

Remember to check solubility at your equilibration temperature, not ambient.

good luck,

ROd

Thank you.