Hi,
I have 8 different starch samples from stressed wheat (drought and heat). I want to plan an assay for define differences of amounts for amylose and amylopectin. I have found one assay kit for it but cannot afford the money (kit is for 100 tests and more than 550€ in Turkey). I was looking for some protocols online for wet lab and found 1 about it “iodine complex”. But couldn’t understand the quantification methods. Do you have any suggestions for it?
Thanks,
Deniz.
http://pubs.acs.org/doi/pdf/10.1021/ja01242a003
The reaction is pretty straight forward. A potassium iodide solution is used as iodine itself is not very soluble and in aqueous solution s the triiodide ion is formed which binds with the spiral structure of the starch molecule and turn to purple-blue color from brown. This blue-black color can easily be measured using a spectrophotometer usually at 600nm. You can easily construct a standard curve using pure starch samples.
As far as a distinction between amylose and amylopectin is concerned, there are some fairly old but tried and tested protocols available that can be used to separate amylose and amylopectin, or if you know the ratio of the two in wheat samples you can get away with measuring just amylose. Also you may need to do a multi-wavelength study to get more accurate results. It can be a bit time consuming but not at all difficult. And yes your concerns about the kits are not unfounded, their prices can be exorbitant but mostly because they are enzyme based.
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