Hi guys

In an experiment we did, the reaction was Ethanol + NAD <=> Acetaldehyde +NADH+ +H+

to the reaction at equilibrium, we added sodium sulphite, HCl and then NaOH

and measured abosrbance at 340nm.

For the sodium sulphite increased indicating and increase in NADH

for the hcl absorption decereased and for NaOH absorption increased.

My question is how does the addition of these chemicals affect the amount of NADH produced.

Thanks for in advance for any help explaining this for me

I can't answer your question very well. I'm not a biochemist. Here is some information on NAD+ and NADH. Maybe you already have this information:

Nicotinamide adenine dinucleotide - Wikipedia, the free encyclopedia (http://en.wikipedia.org/wiki/Nicotinamide_adenine_dinucleotide)

I suspect that the sodium sulfite, HCl, and NaOH merely forced the NADH into a form where it was more or less easily detected (at 340 nm). Sodium sulfite is a reducing agent, so it might be able to reduce NAD+ to NADH. The acid and base probably put the enzyme into an acid or salt form -- the salt being the greater absorber.

Maybe some other wise soul can answer this better than I.

Hi

Thanks for your response. I have looked at that website and many more. Like you I thought that since sodium sulphite is a reducing agent it was reducing more NAD to NADH hence more absorption. However, I read that NAD reduces to NADH by gaining hydrigen ions which NA2SO3 doesn't have. I am new to this and still learning as I go so I might be wrong. As for the acid and base I can't figure how they could affect the reaction, mind you, the NaOH is put in the mixture after HCL to bring it back to its original equilibrium.

Like you said, I hope some guru can come and explain more. I have spent the past two days trying to figure this one out. I need some HELP!

While Na_2SO_3 doesn't have hydrogen ions in its structure, it's still a reducing agent:

2Na_2SO_3\, +\, O_2\, =\, 2Na_2SO_4

Here Oxygen is reduced and Sodium Sulfite is oxidized to sodium sulfate.

Note also that there's always hydrolysis involved with any of these salts of weak acids:

2Na_2SO_3\, +\, H_2O\, =\, 2H_2SO_3 2+ NaOH

and, of course

H_2O = H^+\, + \, OH^-

So in aqueous solution, there is always hydroxide and hydrogen ions present.

Oxidation-reduction always involves something gaining electrons (reduction) and something losing electrons (oxidation), so it's not simply gaining a proton that causes the reduction. NAD may gain a hydrogen ion, but it also must gain one or more electrons to be reduced.