Chromatography Forum

Hi everyone

if someone asked you to prepare a mobile phase using this: ACN: 1.0% TEAA 15:85 pH 4 This is reverse phase
I found this in a publication. I am assuming that aqueous (A) is 100% water and the above is for the organic(B). Also how do you interpret this :Methanol:ACN:HoAc:Tea 10:90:0.03:0.02 , this is for polar organic phase
thanks

It is a bit of slang.

No one weighs solvents or adjust to T and d. Normally people measure by volume and mix.

Say if you need A:B 15:85 take 15 mL of A and add 85 mL of B. For speed some people do 15 then add up to mark in a 100 mL cylinder or vol. fl.

Unless method is not robust a small change will not knock it off. The issue here is that you need to do it consistently, then your retention will be consistent too.

It is a bit of slang.

No one weighs solvents I do (or did when I worked in HPLC) for the simple reason that it a lot more accurate and precise than volumetric measurementsor adjust to T and d. Normally people measure by volume and mix.

Say if you need A:B 15:85 take 15 mL of A and add 85 mL of B. For speed some people do 15 then add up to mark in a 100 mL cylinder or vol. fl.which is a very good way of not getting a 15:85 volume ratio in the final mixture

Unless method is not robust a small change will not knock it off The inaacuracy of large volumetric cylinders is plenty big enough to cause significant changes in mobile phase composition. The issue here is that you need to do it consistently, then your retention will be consistent too.

Vladimir, the first recipe is for a mixture of 15 volumes of AcN mixed with 85 volumes of a 1% solution of TEAA in (presumably) water with its pH adjusted to 4 (with what is not specified). To make about a litre of the second mixture mix 100 ml of methanol with 900 ml of AcN, than add 0.3 ml HoAc and 0.2 ml Tea.

Peter

This is why it's a great skill to express your intentions exactly in writing, so no one can screw them up.

A year ago I changed a mobile phase preparation which used H3PO4 addition to a certain "apparent pH" in an inorganic salt-water-alcohol mix to just adding 6ml H3PO4 from a pipet. Our robustness studies showed that even a 10% difference in H3PO4 amount did not affect the assay.

Guess which way is more straightforward, and doesn't require apH meter or standardizing a pH meter? Obviously, we could have added that H3PO4 by weight as well.

For speed some people do 15 then add up to mark in a 100 mL cylinder or vol. fl.

::Throws (plastic) graduated cylinder at Alexandre (no injurious intent)::

That's a really bad habit to get into. Not all methods are robust enough to deal with the practice, especially if you are working w/ MeOH and aqueous components.

I emphatically second DR on this. How are you going to get repeatability into this? For instance one just pours things a bit differently and gets different mixing at the interface . . . . .

Thanks guys for all your help.