Processing raw data absorbances?

[mogs]

Processing raw data absorbances??

Hi, how do you process raw absorbances again, I know that you have to take the mean of the blank and take it away from all the other readings. However I have 9 blank readings? Which ones do I include in the average? The first two, or all of them?

 

[Yanick]

If this is schoolwork related, you need to post this in the HW section.

I'll start off by saying that it is impossible to know how to process your raw data without knowing what you did.

In general, a blank (or baseline) sample needs to include everything except the analyte. For example if you are assaying an enzyme in a buffer solution, your blank contains just buffer. The nonzero absorbance you get from the buffer must be subtracted from the absorbance you get from your sample (which is enzyme + buffer) thus the net absorbance should reflect the absorbance of the analyte (the enzyme in this case).

9 blanks doesn't sound right, but then again it could make sense in the context of your experiment. Are you sure you're not supposed to be making a calibration curve with different dilutions of a standard?

Once again its impossible to say what's right or wrong if we don't know what you did.

 

[mogs]

Yes we are doing a calibration curve for glucose oxidase, we took 10 absorbance readings over time, for 0.1, 0.01 and 0.00 concentration of glucose? I'm writing out the tables and you have to take the mean of the blank and minus it from every reading. I'm not sure which of the 10 abs readings for conc 0.00/blank I include in the average?

 

[Yanick]

I'm still not quite sure what exactly you are doing. Are you assaying concentrations or activities/kinetics? You need to be more specific as to what you did, then I or someone else may be able to help you.

Activity/kinetic studies make use of the change of absorbance over time (IE appearance of product or disappearance of reactant). Reactions still occur without catalysts present, but they are much slower. Therefore you can get a baseline value of the reaction rate by measuring the change in absorbance in a sample lacking the enzyme. Averaging the absorbances doesn't make sense, but averaging the rates of uncatalyzed reactions does make sense.

Once again you are being too vague with what you were trying to accomplish and how you went about it.

 

[mogs]

Hi yes I'm really sorry, we're not doing kinetics, the experiment we did is called of development time of the glucose oxidase chromophore, followed by standard curve for glucose. I'm doing the table for the first one (development time of the glucose oxidase chromophore) I think you average the first two abs readings for the 0.0 concentration, I wasn't sure though??