Science Buddies: "Ask an Expert"

Hi, I am doing a post harvest biochemistry experiment that involves measuring ascorbic acid content in fruit juices. I have researched several inexpensive and relatively simple spectrophotometric/colorimetric methods (I don't want to use a titration). However, I don't know which one to pick nor have the resources/time to try and compare them all. I do want to pick the one that would give the most statistically accurate results with the least interference and was hoping you could give me an opinion. The methods are as follows:
A. Direct spectrophotometric method measuring absorbance at 266 nm. Interferences include: Iron(II), copper(II), magnesium(II), manganese(II), zinc(II), Nickel(II), citric acid, tartaric acid, benzoate. For details see (http://www.ijens.org/Vol%2011%20I%2002/ ... -IJENS.pdf)
B. DCIP- a blue dye, pink in acidic solution that when reduced by ascorbic acid becomes colorless. Reduction of DCIP is not limited to L-ascorbic acid, and any reducing substance present in the sample can reduce the dye. Substances that can interfere include cuprous, ferrous, and stannous ions, sulfite, thiosulfate, tannins, betanin, cysteine, glutathione.
c. Iodine/Starch method - oxidation reduction reaction so basically the same interferences as DCIP, though a little cheaper and possibly easier to detect
Its hard to compare these methods simply looking at different papers. With so much interference from metals in all methods I was thinking a chelating agent (EDTA) might also be useful (see http://www.mendeley.com/research/spectr ... e-to-iron/)
Thanks

Hi IxPowers,

This sounds like a fascinating project. Are you going to do any purification of the ascorbic acid before you assay it? Your assays will work better if do any kind of purification. Otherwise it sounds as though the two colorimetric methods are very similar. I would think that they would probably be equally effective and have equal amounts of error. So I would choose the one that is either easier t perform and control, or the one that has the largest dynamic range (largest color change from 0 to 100%). As for the spectrophotometric measurement it sounds like this one was designed to be used on a more pure sample. This may be the way to go if you are going to do any kind of purification on your juices. Would it be possible to use the spectrophotometric method (A) and one of the color methods (B or C)? If not I would say method A on a slightly purified sample and B or C on an un-purified sample.

I hope that helps,
Kierstyn