Chromatography Forum

Dear Experts,

I am trying to performs BP Adrenaline Injection method to separate noradrenaline acid tartrate and adrenaline acid tartrate. One of the modifier used in the mobile phase was tetramethylammonium hydrogen sulphate. However, due to the high cost of the material, we tried to swith it over to tetrabutylammoium hydrogen sulphate. But unfortunately, there was no resolution observed for the two compounds. Has anybody out there ever done this before ? Any other suggestions on replacing the tetramethyammonium hydrogen sulphate?

Thanks a lot in advanced.

Louise

If you are looking for a "plug and play" direct substitution, you probably won't find it.

By switching from a short-chain (methyl) ion-pair reagent to a longer-chain (butyl) reagent, you are essentially shifting the ion-pair distribution in the direction of the stationary phase. You can compensate to a large extent by decreasing the concentration, and you will probably have to tweak the organic solvent concentration as well.

Hi Tom,

Thanks for you advise. But i dont quite understand what you meant by longer chain ion paired reagent tends to shift the ion pair distribution in the direction of the stationary phase. Can you furthur explain on this phenomenon?

Thanks.

Louise

This is my assay method;

Carry out the method for liquid chromatography, Appendix III D, using the following solutions. Solution (1) contains 0.02% w/v of adrenaline acid tartrate BPCRS in the mobile phase. For solution (2) dilute 1 volume of the injection to 10 volumes with the mobile phase. Solution (3) contains 0.02% w/v of adrenaline acid tartrate BPCRS and 0.02% w/v of noradrenaline acid tartrate in the mobile phase.

The chromatographic procedure may be carried out using (a) a stainless steel column (10 cm × 4.6 mm) packed with stationary phase C (5 µm) (Nucleosil C18 is suitable), (b) as the mobile phase with a flow rate of 2 ml per minute a solution prepared by adding 4.0 g of tetramethylammonium hydrogen sulphate, 1.1 g of sodium heptanesulphonate and 2 ml of 0.1M disodium edetate to a mixture of 950 ml of water and 50 ml of methanol and adjusting the pH of the mixture to 3.5 with 1M sodium hydroxide and (c) a detection wavelength of 205 nm.

Louise

The ion-pairing is actually between your basic analytes and the heptanesulfonate. The quaternary ammonium salt is actually a competitive agent; a rather kludgy approach in my opinion. Trying to substitute TMA with TBA will be a rather tricky proposition, not to mention the revalidation headache. You could try replacing TMAHSO4 with equimolar amounts of TMA hydroxide and sulfuric acid; I've done that before.

For an alternative mobile phase that we use for our electrochemical methods, see Figure three in http://www1.dionex.com/en-us/webdocs/41 ... 20_V19.pdf

Look for a cheaper supplier of the correct reagent. It just has to be chromatographic grade - with transmittance of not less than 50% at 200 nm and 90% at 220 nm using a 0.005M solution.

In my limited experience, There's usually not much difference in price between tetrabutyl and tetramethyl salts for ion-pair from each supplier, but there can be large differences in prices of ion-pair reagents from different suppliers.

On this side of the planet, Acros and Fisher can be noticably cheaper ( about 1/3 the price ) than Merck and Romil, but the quality of the latter may be superior. However as long as the reagent complies with the BP....

Please keep having fun,

Bruce Hamilton

Dear Chromatoghers,

Thanks guys, i will try out your options. Hope it will work enough to make my boss smile.