The ketone method determines soluble sugars.

I, principle
sugar in the action of thick sulfuric acid, can be dehydrated reaction to produce acetaldehyde or hydroxymethyl acetaldehyde, the resulting acetaldehyde or hydroxymethyl formaldehyde can be produced with acetaldehyde reaction blue-green acetaldehyde derivatives, in a certain range, the depth of color is directly related to the content of sugar, so it can be used for quantitative determination of sugar.
This method is characterized by the determination of almost all carbon water
compounds
, not only can determine the sugar and hexose, but also can measure all oligosaccharides and polysaccharides, including starch, cellulose, etc. (because the strong sulfuric acid in the reaction fluid can put polysaccharides Hydrolysing into monosaccharides and reacting), so the carbohydrate content measured by the ketone method is actually the total amount of all soluble carbohydrates in the solution, in the absence of the need to carefully divide a variety of carbohydrates, the use of ketones can measure the total amount at once, saving a lot of trouble, therefore, there is a special application value.
, however, when measuring water-soluble carbohydrates, the measurement error
increased due to
reaction to the reagent of ketones. In addition, different sugars and ketone reagents color depth is different, fructose color is the deepest, glucose second, semi-lactose, sugar is lighter, five carbon sugar color is lighter, so the determination of sugar mixtures, often due to different sugar proportions caused errors, but when measuring monosaccharides, can avoid such errors.
of colored substances produced by the reaction of sugars and ketones in the visible light region is 630nM, so they are compared at this wavelength.
, instruments and appliances,
electronic
balance
;
capacity bottles
:100ml 4, 50ml 2;
funnel
; small
test tubes
several; electric furnaces; scale straws 0.5ml 1, 2.0ml 3, 5ml 4; light-splitter; marker pen.
III, reagents
ethyl acetate reagents: take analysis of pure ketone 1g, dissolved in 50 ml ethyl acetate, stored in brown bottles, can be stored in the dark for several weeks, if crystallized, can be slightly dissolved by heat;
fourth, method
. 1. The production of the standard curve according to the method of a standard curve of the production method to add the standard sucrose solution, and then in sequence to the test tube to add 0.5ml of ethyl acetate reagents and 5ml of thick sulphuric acid, fully oscillating, immediately put the test tube into the boiling water bath To ensure the temperature of 1Min, after removal, naturally cooled to room temperature, with blank as a control, at 630nM wavelength to measure its light density, light density as the ordinate, sugar content as the horizontal coordinates, draw the standard curve, and find out the standard linear equation.
2. Soluble sugar extract fresh plant leaves, wipe the surface dirt, cut and mix well, weigh 0.10 to 0.30g, a total of 3 servings, respectively, into 3 scale test tubes, add 5 To 10 ml distilled water, plastic film seal, in boiling water to extract 30min (extract 2 times), extract
filter
into a 25 ml capacity bottle, repeatedly rinse test tube and residue, fixed to scale.
3. The color determination absorbs 0.5 ml of sample extract in a 20 ml scale test tube (repeat 2 times), plus 1.5 ml of distilled water, the following steps are the same as the standard curve determination, to determine the light density of the sample.
4. The content of soluble sugar is calculated, the amount of sugar is calculated by the standard linear equation, and the sugar content in the test sample is calculated by formula (1-1).
soluble sugar content (%) (C×V / A×n) / (W×106) (1-1)
medium C: standard equation to find the amount of sugar
(g);
W:
organization
weight (g).
.