Hi guys,
I'm in biochemistry, but a lot of our problems deal with general chemistry concepts.
QUESTION
I have a mix of the following:
NAD+ = .04 ml of 5mM
G3P = .1 ml of .2M
phosphate = .1ml of .2 M
buffer = .46 ml
GAPDH = .2 ml, with a density of 9.7mg/ml
The absorbency is found to be at 340 nm.
Rate of increase of absorbency (delta(A)), is .17/min.
I need to calculate the rate of production of NADH in this reaction, in units of micromoles/min.
ANALYSIS:
The equation is as follows:
G3P + HPO4(2-) + NAD+ --------------> BPG + NADH + H+
So, there is a 1 mol : 1 mol ratio.
I converted all of the above reactants into moles:
G3P = 2e-5 mol
phosphate = 2e-5 mol
NAD+ = 2e-7 mol
Therefore, the limiting reagent is NAD+. I would then get 2e-7 mol of NADH, and thus, 2e-4 mmol of NADH. This is a part where I am stuck. There is a tip to consider the reaction volume.
There is a law that absorptivity is equivalent to the molar extinction coefficient times the concentration times the length, which is assumed to be 1, so it may be disregarded in this problem:
A = E x C
Since I have moles of NADH, I was considering placing that over the total volume of the mixture (this is where the volumes of buffer and GAPDH become quite useful) and then multiplying it by the molar extinction coefficient of 6,300 (which was given) to get a final absorption, but I feel like I'm just shooting in the dark with this.
Any direction is helpful. Again, the rate of absorption is .17/min.
Thank you!
EDIT: Actually, my problem said to "follow the absorbance at 340 nm" which is different from the absorbance being 340 nm. Absorbance does not have units.
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