Sciencemadness Discussion Board

Pyridoxine to Pyridoxal

Om rana - 11-3-2011 at 09:57

hi....everyone..
I am an undergraduate student of Pharmaceutical Science.

I need to oxidise Pyridoxine to Pyridoxal....(reaction media also contain Hydrazine)

Reactiion must be quite specific..with minimum side product...any help would be greatly appreciated.

phlogiston - 11-3-2011 at 12:59

The enzyme pyridoxine oxidase does exactly this, with 100% specificity.

I don't know if the enzyme is/remains active in an environment with hydrazine.

Om rana - 11-3-2011 at 18:10

thanks man..
but we don't have this enzyme..I am looking for some selective oxidising agent or any other reaction which could introduce carbonyl group in pyridoxine ....
...anyway thanks for ur help:(

Chordate - 12-3-2011 at 00:53

Here is the problem: you have two alcohol groups there that have almost identical reactivity, and a third which is substantially similar. Things that attack one can attack the other.

Nature has gotten around this by making a huge molecule which can bind selectively bind the substrate in 1 position and squeeze into proximity with another reactant, forcing a very stereospecific reaction.

Here's that enzyme . (link to Protein Databank). It is comprised of hundreds of amino acids, and its molecular weight is in excess of 60000 grams/mole. It needs this in order to be able to bind to its substrate and flex into place when it is phosphorylated. Organic chemists dream of that sort of specificity.

Which isn't to say that it can't be synthesized, just that the starting material you gave is sort of synthetic dead end without an incredible catalyst system (like expensive enzymes). Can you maybe start with pyridoxamine? That will give you some better options.

jon - 12-3-2011 at 01:26

hydrazine is hardly an oxidizer.
chordate you have a good mind, impressive.

[Edited on 12-3-2011 by jon]

Om rana - 12-3-2011 at 11:39

thanks for your help..
...But actually I am following an analytical experiment for detection of Pyridoxine.

I want to..
Oxidise Pyridoxine to Pyridoxal

then

Pyridoxal + hydrazine would give Hydrazone, which is detected by Uv quantitatively.

So almost all of Pyridoxine should be oxidised to Pyridoxal and yes enzyme would do this but it can't be used in media.
....need help in oxidising step.


Hydrazine ia a reducing agent.:)

Chordate - 12-3-2011 at 14:56

If that is the case then I wouldn't about winding up at pyridoxal. Just oxidize both methylhydroxy groups on pyridoxine to carbaldehydes using pyridinium chlorochromate and then form a dihydrazone from there. In order to get quantitative analysis you'll have to figure out a new extinction coefficient of the dihydrazone, but that's do-able.

phlogiston - 12-3-2011 at 15:45

I routinely measure pyridoxine by HPLC-tandem MS (in biological samples), as well as pyrodoxamine, pyridoxal and pyridoxic acid and the phosphorylated forms.

But I guess you want a spectrophotometric method.

Om rana - 13-3-2011 at 05:35

thanks for the response.


hey Chordate....
Wiki says
" In practice the chromium byproduct deposits with pyridine as a sticky BLACK tar, which can complicate matters. Addition of an inert adsorbent such as crushed molecular sieves or silica gel allows the sticky byproduct to adsorb to the surface, and makes workup easier."

it will certainly complicate further Spectrophotometric determination.

and Phlogiston yes I want a Spectrophotometric Analysis...but could you tell me how you analyse Pyridoxine by Hplc...means how you detect it quantitatively.

:)


Chordate - 13-3-2011 at 10:38

It is more of a brown syrupy liquid substance which stains anything it touches. You can get white crystals of your final material out of it with an acid base extraction and several washes, but that is going to cause you to lose some material.

What sort of samples are you working with? Why can't you just run the UV on the original sample? According to the merck index (13th edition, page 1428) pyridoxine has a molar extinction coefficient of 8900 in 0.1N HCl at 290 nm, and 7100 at 325 nm in a pH 7 phosphate buffer.

Nicodem - 13-3-2011 at 11:08

Quote: Originally posted by Om rana  
I need to oxidise Pyridoxine to Pyridoxal....(reaction media also contain Hydrazine)

You might want to explain why does the hydrazine needs to be added before the oxidation, when what you need it for, is for the hydrazone formation?
It does not sound very logical to me, but I know not much about spectrophotometric determinations anyway. Though I do know enough to know that, unless you stick exactly to the original analytical method, your result will be totally unreliable. If you will use any other oxidant but the enzymatic reaction (I assume this is prescribed in the literature analytical method?), you will also have to develop the full method from the start (including trivial stuff like the calibration curve in your sample media, etc.). All this might cause you headaches when trying to validate the method.
Quote: Originally posted by Chordate  
If that is the case then I wouldn't about winding up at pyridoxal. Just oxidize both methylhydroxy groups on pyridoxine to carbaldehydes using pyridinium chlorochromate and then form a dihydrazone from there. In order to get quantitative analysis you'll have to figure out a new extinction coefficient of the dihydrazone, but that's do-able.

Reactions of 1,4-dicarbonyl compounds with hydrazine gives pyridazines rather than the corresponding bishydrazones. In this specific case, the product should be 7-methylpyrido[3,4-d]pyridazin-8-ol. In practice this should not be necessarily wrong as it should have a good absorbtion at discrete wavelengths, so in effect it might work as good as the hydrazone as far as spectrophotometric determination goes (with proper method development and all that).

Om rana - 13-3-2011 at 22:26

The sample contains both Pyridoxine (in small amount)and Hydrazine and both need to be determined Spectrophotometrically.

say Hydrazine(200mg) and Pyridoxine(10mg)
I was thinking of ...
oxidising Pyridoxine to Pyridoxal(10 mg) .....

1) reaction with hydrazine(200mg)......formation of Hydrazone(20mg)yellow colured(can be determined in visible region using simply colorimetrically).

2)now the hydrazine(190mg) is left in solution. now by adding excess of Pyridoxine externally hydrazine (190mg) left could be converted fully to hydrazone(yellow).

in the 2) step i want to add pyridoxine to remaining hydrazine ...so that complex is identical to that formed in1) step..it would result in additive absorption in a region.

and yes I want to bring complex to visible region so that it could be determined simple colorimetry.

and guys! forgive me if I ave made it a mess