Chromatography Forum

Hello, all!

I need advice in following problem:

I have to separate 3 positional isomers of Toluidine (o-,p-,m-), than I have to do their trace analysis, i.e. m- and p- isomers (at 0.1% level) in o-Toluidine (100%).

Up to this point I have succeded to resolve o-Toluidine from m-Toluidine (on C-18 stationary phase with subambient temperature), but p-Toluidine trace still eluted under the o-Toluidine main peak

Maybe there is some other technique (CE ?), I'll appreciate any suggestion.

You can play with buffer nature and try to adjust pH hoping that you can get separation. You can try to use phenyl column and add pi-pi interaction.

Another approach is to add ion-exchange interaction with mixed mode approach. The general scheme is described here:
http://www.sielc.com/Technology_2D_Properties.html
You are using 2-D approach on a single column. I will try to see if we can do this for you first week of January. We have similar application for the separation of aminobiphenyls:
http://www.sielc.com/compound_188.html
In your case I would go either with Primesep 100 (reverse phase and ion-exchange) or Primesep P (reverse phase, ion-exchange and pi-pi interactions) column.

You can also use GC.

Regards,

Vlad

>>Mixed mode
Thanx a lot for a hint, it might be interesting
>>GC
No separation on GC achived (yet)

This separation should be rather straightforward on a silica column with normal phase chromatography. This is where I would have started with this task.

Another excellent choice is to use a graphitized carbon column, these easily separate the named isomers.

Regards,
Mark

Uwe and Mark, thanks a lot for your advice, i'll update this topic with results...

Another excellent choice is to use a graphitized carbon column, these easily separate the named isomers.

Regards,
Mark

Mark, would you, please, provide me some reference or start-point, i've had no experience with this type of stationary phase

Thanx in advance

I would do it by GC

Avitan,

I ordered toluidines. If we get compouns this week you will have a method in couple of days.

Vlad

That would be wonderful, Vlad

Bill, thank you for your input, we didn't achived positive results on GC yet...[/b]

Hello

I think you'd get good results with pre-column derivatisation, using OPA and a C18. You need to use a gradient of course, but you will get baseline separation of o-,m- and p- isomers - within 20 minutes is my guess.

Usually OPA requires fluorescence detection, but in this case, a UV should do. Detection at around 340 nm.

If you try this out, I'd be very happy if you could share the results with me.

Best rgds,

...using OPA and a C18. ...

1. What OPA is?
2. I'll have to check recovery for derivatization process, am I?

Thanx for your input Srinivas SK

OPA = orthopthalaldehyde. Derivatising reagent used for primary amines. Usually meant for the HPLC analysis of, but not restricted to, amino acids. OPA-derivatives are fluorescent and can be detected at low levels.

Check out this link:

http://www.sigmaaldrich.com/img/assets/ ... 2_new_.pdf

This should answer some of your questions.

Best rgds,

Srinivas SK, thank you for this link, I'll consider this option

Hi Avitan,

My two messages to you email bounced back. We have a method for you and I will publish a link as soon as we place the method on our website. We achieved very good separation with 3-5 minutes between peaks (150 mm Primesep 200 column).

regards,

Vlad