HPLC Method for sodium acetate anydrous determination

[alemaggot]

I'm sorry but I can't understand all that you say. Sorry for my bad english comprension

I write example.

I weigh, for example, 1 gram of my substance. I must filtrate it with about 10 ml of water + 2-3 drops of H2SO4 and I must inject the liquid phase without dilution. It's correct?

[tom jupille]

Probably best to approach the problem in small steps.

1. Begin with a solution of acetic acid in whatever your final solvent will be. Run that on the As I understand, your procedure looks approximately like this:
1. Extracting the sodium acetate from your formulation with water (most of your formulation is not soluble in water, but sodium acetate *is* soluble in water.
2. Acidify the resulting aqueous extract by adding sulfuric acid. That will shift the acetic acid - acetate equilibrium in the direction of acetic acid.
3. Carry out liquid-liquid extraction with DMF, to extract the acetic acid, leaving essentially sodium sulfate behind in your aqueous solution.
4. Inject the DMF solution into the GC

If this were my problem, I would work in small steps:

1. Start with a solution of acetic acid (at the expected concentration) in DMF and inject into the GC. That will confirm that the chromatography is working OK.

2. Dissolve some acetic acid in water, add the sulfuric acid, extract, and run GC. That will confirm that the liquid-liquid extraction is working OK.

3.Dissolve some sodium sulfate in water and re-run the acidification / extraction / chromatography. That will confirm that the acidification is working OK.

4. Finally, run through the entire procedure with a check sample of your formulation.

That will let you identify at what stage any problems occur.

[DSP007]

Hello, Regard Tom.
I do not think that DMF in this case is a good solvent Its pKa higher than of water. And from sulfuric acid DMF will as the base (Alkaline).
http://pubs.acs.org/doi/pdf/10.1021/ja00024a041
I think you first need to try to simply wash waters, and even better - with a model solution of sodium acetate.

[carls]

This is turning into a GC topic.

Liquid-liquid extraction is not possible using DMF and water since they are miscible. Perhaps hexane would be a better choice (acetic acid has reasonable solubility (miscible?) in hexane). Hexane's density is lower than water making the extraction easy from a vial (analyze the top layer). Example: add the sample to hexane, swirl to mix/dissolve(?), then add 0.1M H2SO4, sonicate 5min, vortex/mix 5min, allow phase separation, analyse top layer. Dibutyl ether could be substituted for hexane if another solvent is desired.

HILIC using an amino column should work. Phosphate is likely the best buffer due to its UV transparency but its solubility in 90% ACN is close to zero. Stay with phosphate buffer but start at 75% ACN with 5mM NH4+H2PO4- at pH 6, detect at 210nm.

[tom jupille]

DMF and water since they are miscible.

Yes, they are!

That's what happens when I post with an insufficient caffeine level in my blood!

However, the point was that working up the method one step at a time makes it easier to isolate and solve any problems (such as miscible solvents! ).

[alemaggot]

Thank you very much for your reply guys!

I've got some question... The injection with GC system that you've say they must be do with head space method or with direct injection?
However inject water in GC isn't the best way to conserve tha stationary phase of column. Same discourse for sulforic acid. It's vapour damage the GC system.

Now... What extraction I'll must be try? With hexane or DMF? If I use Hexane I can't use headspace method. If I use DMF the extraction dosn't work.

I've write error?

Carl before your reply I didn't Know the existence of HILIC. Now I can't try that you say (I don't have a HILIC column). In future maybe I'll can try it! For now thank you!

[carls]

add the sample to hexane, swirl to mix/dissolve(?), then add 0.1M H2SO4, sonicate 5min, vortex/mix 5min, allow phase separation, analyse top layer.

Pipet off aliquot of top liquid layer (hexane) and inject using split injection.

[alemaggot]

add the sample to hexane, swirl to mix/dissolve(?), then add 0.1M H2SO4, sonicate 5min, vortex/mix 5min, allow phase separation, analyse top layer.

Pipet off aliquot of top liquid layer (hexane) and inject using split injection.

Before the hexane addition I must wash my sample with water, correct? For filtrate water from my solution what is the better method?

If I understand when I inject the hexane solution sulforic acid is in down layer. He has reacted with sodium formed sodium sulfate. Than it is not in my hexane solution. Correct?

[tom jupille]

Just to review the overall procedure:

1. Wash ("extract") your sample with water. The sodium acetate will dissolve in ("be extracted into") the water. Filter the water to remove any remaining particles.

2. Add sulfuric acid to the water. This will shift the equilibrium from acetate to acetic acid. Sulfuric acid is stronger than acetic acid, so (sulfuric acid) + (sodium acetate) goes to (sodium sulfate) + (acetic acid).

3. Add some hexane and shake the mixture to get good contact. The acetic acid will end up in the hexane leaving the sodium sulfate in the water.

4. Pipette away some of the hexane layer and inject into the GC

[alemaggot]

Ok guys! I understand!

In the next week I'll try this and I'll write what I found.

Thank you

[alemaggot]

Today I try to do extraction!

During this In my mind was born one doubt. My compound isn't insoluble in water, but it's very slightly soluble in water. When I sonicate it some parts of it pass in acqueos solution. When I extract with hexane in the top layer solution (hexane + acetate) I found also some parts of compounds (I don't know if it is soluble in hexane, I must try)? If I inject it in Capillary GC column it's a problem. Can I resolve this problem?

[carls]

No problem to inject into GC. Thats what inlet liners are for

[Peter Apps]

Today I try to do extraction!

During this In my mind was born one doubt. My compound isn't insoluble in water, but it's very slightly soluble in water. When I sonicate it some parts of it pass in acqueos solution. When I extract with hexane in the top layer solution (hexane + acetate) I found also some parts of compounds (I don't know if it is soluble in hexane, I must try)? If I inject it in Capillary GC column it's a problem. Can I resolve this problem?

If you can see "parts of compounds" in the hexane layer then they are not in solution, they are particles in solution. Try filtering the hexane layer through a syringe=tip filter after you do the extraction and before you inject the hexane layer onto the GC. Despite what Carl says you can cause all sorts of problems by injecting dirty samples.

Peter

[DSP007]

You have headspace ? If yes - not need liquid/liquid extraction.
In headspase you have liquid/gase phase extraction , very effective by acetic acid water solution. Acetic acid is high boiled, but vary volature liquide.
The sulfuric acid (in water solutions) not volature liquid. Quicly dead of column by cross-link phase will not. Effectivity of column , tailing factor - in you quest is secondary question.

I'm recomendate. Make the some water solution of acetic acid by concentration 10, 5. 1, 0.5, 0.1 mg /ml and analise it (headspace 80-90 C, 10 ml , after add 1-2 drop of concentrated sulfuric acid and Na2SO4 ). For (from) determerminate calibration curve linearity and sensivity of analitical method.
After that we will discuss results.

[Greyhoundchrom]

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Good luck with your research