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  Section: Plant Lab Protocols
 
 
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Methodology for Carbohydrates

 
     
 

Determination of amylose
 
Starch is composed of two components, namely amylose and amylopectin. Amylose is a linear or non-breanched polymer of glucose. The glucose units are joined by a-1-4-glucosidic linkages. Amylose exist in coiled form and each coil contains six glucose residues.
 




Principle

The iodine is adsorbed within the helical coils of amylase to produce a blue colored complex which is measured colorimetrically.
 




Materials

Distilled water
1N NaOH
0.1% phenolphthalein
Iodine Reagent: Dissolve 1g iodine and 10g KI in water and make up to 500mL.
Standard: Dissolve 100mg amylase in 10mL 1 N NaOH; make up to 100mL with water.
 

Procedure

1.
Weigh 100mg of the powdered sample, and add 1mL of distilled ethanol. Then add 10ml of 1 N NaOH and leave it overnight.
2.
Make up the volume to 100ml.
3.
Take 2.5ml of the extract, add about 20ml distilled water and then three drops of phenolphthalein.
4.
Add 0.1 N HCl drop by drop until the pink color just disappears.
5.
Add 1ml of iodine reagent and make up the volume to 50ml and read the color at 590nm.
6.
Take 0.2, 0.4, 0.6, 0.8 and 1ml of the standard amylase solution and develop the color as in the case of sample.
7.
Calculate the amount of amylose present in the sample using the standard graph.
8.
Dilute 1ml of iodine reagent to 50ml with distilled water for a blank.
 
 




Calculation

Absorbance corresponds to 2.5ml of test solution = ‘x’ mg amylose
 

100 ml contains =
‘x’
X 100mg amylose
2.5
 
 
                               
                            = % amylose
 

Note
1. The sample suspension may be heated for 10min in a boiling water bath instead of overnight dissolution.
2. The amount of amylopectin is obtained by subtracting the amylose content from that of starch.

References
1. McCready, R M, Guggolz, J, Siliviera, V and Owens H S (1950) Anal Chem 22 1156.
2. Juliano, B O (1971) Cereal Sci Today 16 334
3. Thayumanavan, B and Sadasivam, S (1984) Plant Foods Hum Nutr 34 253.

 
     
 
 
     




     
 
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