[Schattenmaler]

Hey,

I am a biochemistry student and I am currently envolved in designing a coupled enzymatic reaction that results in release of carbonate (CO₃^2-). Usually I would add Calcium ions to the reaction in order to precipitate CaCO₃ and measure the decreasing Ca²⁺ concentration, but this is not possible in this context.

So I came up with the wild idea to use the carbonate in a follow-up organic synthesis reaction. The 3 main conditions are: Carbonate is converted in the reaction, water as a solvent, educt and product easily destinguishable and quantifyable by common spectroscopy methods.

(I got the enzymes coupled to beads so I could remove the carbonate-containing liquid from the column and perform the reaction without destroying the enzymes. So you don't have to take that into consideration => however it would be very nice if I would be abe to do it all in one pot)

Organic synthesis is not my field of expertise, so I would be glad if you could name or link reactions that could be interesting to me or give further advice on the subject. Also, correct me please if you find major flaws in my thinking.

Thank you in advance for your input,

Schattenmaler

[Arkcon]

I'm having a little bit of trouble understanding your plan.  I don't know everything about everything, so it may be possible.  But I'm afraid you may need to know everything about everything in order for this to work.  Let's try to break down your post into statements -- which fortunately are way simpler already than we sometimes get, so thank you for that.

Hey,

I am a biochemistry student and I am currently envolved in designing a coupled enzymatic reaction that results in release of carbonate (CO₃^2-).

Do you have an enzyme system that works for this reaction?  Please describe it for us?  I have an enzyme system that produces CO₂ gas, inside of each of my cells, the Krebb's cycle.  It is, if I recall university Biochemistry, a very complicated system.

Usually I would add Calcium ions to the reaction in order to precipitate CaCO₃ and measure the decreasing Ca²⁺ concentration, but this is not possible in this context.

Can you describe better why not?

So I came up with the wild idea to use the carbonate in a follow-up organic synthesis reaction.

Carbonate fixation in organic synthesis.  Hrm.  We usually consider carbonate reactions to be inorganic.  But I don't know everything.  Our experts may be able to help us here.

The 3 main conditions are: Carbonate is converted in the reaction, water as a solvent,

That might be asking for a lot.

educt and product easily destinguishable and quantifyable by common spectroscopy methods.

*Sigh* That might also be asking for a real lot.  We get this sort of request often.  And even in basic high school chemistry, I learned that often enough, you don't get a UV-Vis response for what you need, but instead need indicators.

(I got the enzymes coupled to beads so I could remove the carbonate-containing liquid from the column and perform the reaction without destroying the enzymes. So you don't have to take that into consideration => however it would be very nice if I would be abe to do it all in one pot)

Uh.  All done?  Great.  Or are you planning to do this?

Organic synthesis is not my field of expertise, so I would be glad if you could name or link reactions that could be interesting to me or give further advice on the subject. Also, correct me please if you find major flaws in my thinking.

Let's wait for our Organic experts.  Also, this may belong in Biochemistry.

Thank you in advance for your input,

Schattenmaler

Remember you wrote that if this turns out not to be feasible, when the urge to blame my shortsightedness swells within you.

[Schattenmaler]

Do you have an enzyme system that works for this reaction?  Please describe it for us?  I have an enzyme system that produces CO₂ gas, inside of each of my cells, the Krebb's cycle.  It is, if I recall university Biochemistry, a very complicated system.

I have an enzymatic system that works in principal. It utilizes Carboanhydrase and silicateine to convert solved CO₂ to HCO₃^- to CO₃^2-. On its own, each reaction works, but coupling both was not done yet.

Can you describe better why not?

Apparently because of pH condition (too alcaline), which of course is not impossible to fix, but finding a way to work around it would be great.

That might be asking for a lot.

Yes, I know...

Uh.  All done?  Great.  Or are you planning to do this?

Was done by my predecessor.

Also, this may belong in Biochemistry.

I put it in here, because the biochemistry part is not the problem. I just need input from someone more experienced in the field of organic chemistry.

[Borek]

Apparently because of pH condition (too alcaline), which of course is not impossible to fix, but finding a way to work around it would be great.

Not sure what you mean here - CaCO₃ precipitation requires alkaline conditions. Lower the pH and you have karst chemistry, lower it even further and you have fizzling.

[wildfyr]

If carbonate is being released, there should be a pH change. Use a pH indicator that undergoes color changes around the pH of interest and a UV-Vis. You will probably need to make carbonate stock solutions with the indicator and establish the absorbance curve yourself, since pH indicator color shifts are not just simple increasing absorbance with concentration curves. They tend to look like



If you poke around, perhaps you can find another metal salt that will work if calcium is having a problem (binds to enzyme?)

[wildfyr]

It occurs to me that youre probably already in a buffer. You should do the calibration curve in the buffer solution. I am a little worried that the pH changes my be small and difficult to measure.

[Babcock_Hall]

Are you certain that you cannot use some sort of coupled enzyme assay that involves bicarbonate as a reactant?  Many coupled enzyme assays are known, and adapting one of them might save you time.  Alternatively (and as implied by wildfyr), some enzymes that release or consume protons can be assayed via a pH-stat.