Chromatography Forum

Hello,
does anyone know an LC method for tri n-butyl phosphate quantification? is reverse phase an option?

is reverse phase an option?

Probably "yes", perhaps with the addition of some ion-pairing reagent.

While a RP column is a possibility, you will need a mobile phase containing high percentage of aqueous. Since the analyte of interst has no chromophore, you will most likely need an aerosol based detector, such as ELSD, CAD, or MS, which requires the mobile phase to be volatile. The sensitivity of these detectors is lower in high aqueous condition compared to high organic solvent condition. Considering all factors, a reversed-phase/anion-exchange mixed-mode column mag be a better choice for this application. The attached links lead to product information for two mixed-mode columns that can be used for analyzing three alkyl phosphate and many other anionic analytes. The mobile phase can be acetonitrile/ammonium acetate buffer, say in 60/40. The retention of the analyte can be controlled by buffer concentration. Good luck!

Acclaim Mixed-Mode WAX-1: http://www.dionex.com/en-us/webdocs/488 ... 021407.pdf
Acclaim Trinity P1: http://www.dionex.com/en-us/webdocs/707 ... 239-02.pdf

maybe this application would be good for you
i would not look at the fact that these are nucleotides that are polar anyway, but also the fact that they have the pospho group
http://www.sepax-tech.com/application_notes/OD1001.pdf
it is a polar embedded c-18 column, it should normally be a little cheaper then a mixed mode column
and also you might not need the ion pairing agent.

maybe this application would be good for you
i would not look at the fact that these are nucleotides that are polar anyway, but also the fact that they have the pospho group
http://www.sepax-tech.com/application_notes/OD1001.pdf
it is a polar embedded c-18 column, it should normally be a little cheaper then a mixed mode column
and also you might not need the ion pairing agent.

Thanks for this link, I will give it a try and hopefully it works with a UV detector!
Is that a phosphate buffer described for mobile phase? any recomendations if sodium or potassium phosphate? and buffer strength? should I start with 0.1M and then try lower strengths?

It is most likely you won't be able to detect tri-n-butyl phosphate using a UV detector due to lack of chromophore. Depending on the sensitivity requirement, a RI detector might be applicable. The more common detectors for this analytes will be ELSD, CAD or conductivity (if using IC). When an ELSD or CAD is used, the mobile phase must be volatile, such as ammonium acetate or formate.

How can you see phosphate by UV? If phosphate like this can be monitored by UV you would never use any of the phosphate buffers at low wavelength.
the only way you can use UV for your molecule is to use indirect detection. Add something UV active to your mobile phase (aromatic compound) and observe your compound as a negative peak. This is not a great approach but it works. You background noise might be significant.

How can you see phosphate by UV? If phosphate like this can be monitored by UV you would never use any of the phosphate buffers at low wavelength.
the only way you can use UV for your molecule is to use indirect detection. Add something UV active to your mobile phase (aromatic compound) and observe your compound as a negative peak. This is not a great approach but it works. You background noise might be significant.

Hello Vlad,
I understand this and we do use phosphate mobile phases at low wavelengths, I assumed maybe working at high wavelengths (254, 280 nm or higher) may be an option, to be honest I haven't yet read much about properties of this molecule and was screening first for some opinions with direct conclusions.
Thanks for the feedback

I was mistaken when I made my first response to the question. Tri n-butyl phosphate is a neutral (not a anionic)organophosphate. Sorry for the confusion.

GC is probably the best way to analyze it. If HPLC is the desirable method, a RP column is more appropriate. However, the detection allenging - RI, MS, and indrect UV suggested by Vlad. ELSD or CAD probably won't work because it is too volatile.

If you need to analyze the degradation products, such as di n-butylphosphate or mono n-butyl phosphate, what I suggested before such as using RP/anion-exchange mixed-mode column with ELSD or CAD detection would be valid.