Chromatography Forum

Need a bit of help/suggestions please. I am trying to determine p-toluenesulfonic acid at fairly low levels in drug substances (cannot elaborate further) and need some suggestions for a method. Have tried a gradient on C-18 ACN/water with 0.1% TFA and get very poor response for the p-TSA. Any help, including ioxn-exchange conditions would be helpful.

Thanks in advance for any help,
Mark

Hmm - what do you call a poor response?

I can see that you may not get a lot of retention in a gradient. If this is the case, I would run it in 100% water with a column that can handle such a mobile phase. I doubt that you need TFA. Anything will do that is compatible with your detector, even nothing (depending on the other ingredients in your sample).

If this faisl, a HILIC column will do just fine. Just do not get an amino HILIC column, otherwise you will never see your analyte again.

We use paired-on chromatography (a quaternary ammoinium compund in water, plus organic) on RP-18 for hydrotropes toluenesulfonic acid salt and xylenesulfonic acid salt, using UV detection on a RP-18 column.

Hi Mark,

If you have to use RP column for this application for any reason, as Uwe suggested, you need a column that is 100% aqueous compatible and stable under such conditions. You could try ammonium acetate or phosphate buffer with no organic modifer to elute p-toluenesulfonate isocratically first, then ramp up to elute the rest. Consider Atlantis C18 (Waters) or Acclaim PA (Dionex) for this.

If you have the flexibility to use something other than reversed-phase, I suggest you take a look at the Acclaim Surfactant column. This column gives 4.2 min retention for p-toluenesul on a 4.6 x 150 mm column under the following conditions:
mobile phase: ACN/0.1 M NH4OAc, pH5 v/v 50/50
flow rate: 1 mL/min
Temp.: 30 C
Detection: UV at 220 nm

This column also provides hydrophobic retention at the same time. The following link can give you more information

http://www1.dionex.com/en-us/webdocs/da ... et_V24.pdf

Thanks for all the suggestions and guidance. I think I will give the ion-pairing a try. I found an app on the Dionex site close to what I needed after I posted here. I appreciate the help and fast response.

Thanks again to all,
Mark

Mark,

Provide me with your email address and I will send you information on separation of p-toluenesulphonic and bensenesulphonic acid on Primesep column (no ion-pairing reagent, LC/MS compatible conditions).

Send you information to mail@sielc.com

We are going to post it on the website next week (in case you don't want to reveal your email address)

Regards,

VO

Why mess with ion-pairing. I used to run this on an anion exchanger (Dionex column) in suppressed mode and it worked nicely.

As Uwe points out HILIC may be an even more attractive route.

Einar,

Could you point me to the reference for the Dionex app please? Also HILIC sounds interesting, could someone give some general guidance like column and starting mobile pahse to try? Thanks in advance for all the help with this.

regards,
Mark

Mark,

Here is LC/MS compatible method for p-TSA, no ion-pairing reagent required. Mobile phase ACN-water-ammonium acetate. You can do it with 25 mmol of AmAc:

http://www.allsep.com/images/chr_113.gif

Contact us if you have any questions
Regards,

Vlad

Mark,

It's fairly easy to analyze toluene sulfonate on a wide range of Dionex columns. The closest I could find, though, is the test chromatogram for the OmniPac PAX-100 includes benzene sulfonate. Of course, toluene sulfonate elutes a bit after this but is easily resolved from benzene sulfonate. Frankly, though, I would recommend the AS20 column for this application. It provides excellent peak shape for aromatic sulfonates (more information can be found here: http://www1.dionex.com/en-us/webdocs/manuals/ic/065044- ). You'll see one example in the manual showing separation of including p-chlorobenzene sulfonate on page 16.