Measuring sugar content

[tttman]

I am doing a science fair project and need to measure the sugar content of a cellulose solution where cellulase has been added. I need an actual number, not just a chemical reaction to show the presence of sugar.

 

thanks

[hypervalent_iodine]

Is it pure cellulose? By sugar, I assume you mean free glucose monomers? Cellulose is a sugar polymer, so you should be a little clearer in what you want. Separating the glucose should be fairly easy on account of the distinct solubility profiles. I am also curious as to what exactly you mean when you say, 'cellulose solution'. Cellulose dissolves in very little of anything without chemical modifications.

[CaptainPanic]

I am doing a science fair project and need to measure the sugar content of a cellulose solution where cellulase has been added. I need an actual number, not just a chemical reaction to show the presence of sugar.

 

thanks

The best method is HPLC: High pressure liquid chromatography. It's also probably the most expensive, and you obviously need to have access to HPLC equipment.

 

But if you do not have that available, you can do a reasonably quantitative test using o-toluidine. You still need a spectrophotometer, because it only gets qualitative if you can measure how much color you have in your sample...

 

Google gave me some information (mostly about blood sugar level test, which uses the same o-toluidine), but I think you're gonna have to find an experimental description in a paper to get going.

 

Hopefully these keywords help you a little.

 

[edit] Much easier is to measure how much cellulose you have before and after the hydrolysis, and just assume that all cellulose that is gone became glucose. But that was not the question, still it may be a useful solution.

Edited January 6, 2012 by CaptainPanic

[tttman]

I want take newspaper, water, blender; add enzyme to to this paper pulp mixture to convert cellulose to sugar and measure that amount. I do want a qantitative result, not just a demonstration that there is sugar present.

[hypervalent_iodine]

The best method is HPLC: High pressure liquid chromatography. It's also probably the most expensive, and you obviously need to have access to HPLC equipment.

 

I don't think that someone doing a science fair project is going to have access to a HPLC.

 

I like the idea of using quantitative spectrometry, though I would still question whether or not the OP has a UV-Vis available to them. Another way to test for it with UV-Vis is by reacting the glucose with 3,5-dinitrosalicylic acid. This oxidises the glucose and reduces the DNSA to 3-amino-5-nitrosaliclic acid, the latter of which has a distinct red-brown colour. You would need to do a standards curve, of course, but that's no real issue. I tutored a first year prac where students tested the amount of sucrose in cereal and in soft drink by first hydrolysing the sucrose present and then reacting it with DNSA and it worked quite well (ignoring the obvious contributions to their readings by other sugars already present in their samples).

 

[edit] Much easier is to measure how much cellulose you have before and after the hydrolysis, and just assume that all cellulose that is gone became glucose. But that was not the question, still it may be a useful solution.

 

That would depend on what equipment they have access too. Separating the cellulose from the cellulase is a lot more involved than simply filtering off the cellulose/cellulase precipitate to give you a glucose solution; if you can't do a column or even a liquid-liquid extraction (which may also work), the former method becomes much less feasible (and would still be a bit more difficult than trying to simply isolate the glucose).

[CharonY]

With paper pulp I assume that the enzymatic reaction will not be terribly efficient. The easiest way is probably rough filtration and then to treat the supernatant of the mixture with Bendict's reagent (the quantitative version potassium thiocyanate). That should be readily available. And then quantification with either titrating or spectroscopy, depending on what is available (and with proper calibration curve as per hypervalent_iodine's suggestion).

Edited January 6, 2012 by CharonY

[tttman]

Could you use a glucometer?

[ewmon]

I am doing a science fair project and need to measure the sugar content of a cellulose solution where cellulase has been added. I need an actual number, not just a chemical reaction to show the presence of sugar.

Different suggestions:

1. I'm experienced performing an anthrone-sulfuric acid assay, but you'd need the anthrone and sulfuric acid as well as a (very) hot water bath, the nerve/dexterity of injecting the sulfuric acid reagent into aqueous solutions and then placing test tubes of these reacting chemicals into a hot water bath, and then a UV-Vis spectrophotometer (ie, one that reads in the ultraviolet through the visual spectrum) to obtain the absorbances resulting from the glucose reacting with the anthrone. The sulfuric acid should hydrolyze the cellulose, and so negate using cellulase. You'd obtain the quantitative concentrations of these solutions by back-calculating their absorbances against a standard curve made from known concentrations, which can be done using a spreadsheet app.
   
2. If you were confident that the vast majority of dissolved matter from the newspaper was cellulose, then you could try a dry weight assay. Add the newspaper to water, add the cellulase, allow it to react. Then filter out the non-dissolved particles — or centrifuge the solution and pour off the supernatant — and evaporate the water, weight the dried material, and compute the concentration based on the volume of the original solution.
   
3. If you contact food testing laboratories in your area, you might find one nice enough to test your samples (by whatever means) while you watch and/or participate in some (perhaps minor) way.
   
4. If you can use the glucometer (I don't know if it would work), its validity/accuracy may depend upon operating under biological conditions (37°C, pH≈7.4, etc).