Chromatography Forum

Hello all,

I've to develop a method to separate Phenylalanine et Nitrophenylalanine
Someone know some conditions to separate them ?
For know in reverse phase (C18 CH3CN/H20 ph 6,5) these 2 compounds co-elute.
I surely have to optimise the pH of the eluent but i don't know if i have to work in acid or basic condition.

Tanks for your help

laurent

You can try Primesep 100 mixed mode column. Compounds retained by combination of reverse phase and ion-exchange. Both compounds have different characteristics and can be separated like other amino acids without IP reagent.
http://www.sielc.com/compound_002.html

Here are applications showing power of mixed mode approach:
http://www.sielc.com/compound_220.html (separation achieved on 50 mm column)
http://www.sielc.com/compound_215.html
http://www.sielc.com/compound_287.html

What is your detection technique?
Send us samples and we will develop a method for you free of charge.

Vlad

Laurent,

Although a Mixed-Mode column might be able to separate these two compounds, I feel simply a RP column should solve the problem. I would use the same C18 column at a lower pH, say 25 mM phosphate buffer at pH3 if allowed. You might need to adjust mobile phase organic content.

If no luck, try one of the RP polar-embedded (PE) columns using the same condition. Usually PE columns provide additional interactions such as hydrogen bonding and better shape selectivity. There are a collection of PE columns available, including sulfonamide-embedded (Acclaim PA from Dionex), carbamate-embedded (Waters Symmetry Shield RP18), and amide-embedded (Dionex Acclaim PA2, Supelco Discovery Amide-RP, Agient Zorbax Bonus RP, ...).

I have Phenylalanine, and I will check to see if I have Nitrophenylalanine in the lab. If I have both, I might try it myself.

It was a few years ago now, and I had a separation problem with nitro compounds. I ended up using a fluorinated reversed phase column to separate some structurally related camptophecin analogue nitro compounds. Worth a shot?