Activity of Diastase On Starch

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                    ACTIVITY OF DIASTASE ON STARCH

                         

 Brief

                           

                       Starch is a polymer where the individual units in the polymer are glucose molecules. The most important enzyme that degrades starch is called diastase. This enzyme catalyses a split in the starch polymer so that eventually what is left over are short chains consisting of just a few glucose units (maltose). These can then be broken down by water so that the entire starch polymer is degraded to glucose. The glucose concentration that will be produced in a fixed time by the action of diastase on starch can be measured to find out the rate of hydrolysis of starch, as the equation below shows:

           STARCH              DIASTASE                      GLUCOSE  +  MALTOSE                

                              

                        The reaction between starch + water = glucose is called hydrolysis. In the chemical reaction that occurs, water is introduced into the bond between two glucose units with a -OH becoming part of one side of the bond and a hydrogen (H-) becoming attached to the other glucose molecule.

                        Rate of hydrolysis of starch    α   concentration of glucose produced

   

                                     (In the above relation time is kept constant)

                        Glucose concentration can be measured by titrating glucose solution with quantitative Benedict solution having sodium carbonate in it.

                         25ml of quantitative Benedict solution+10 grams of sodium carbonate can be titrated with a solution having glucose in it. This titration gives us the volume of glucose solution used to carry out the titration. Subsequently we can find out the rate of hydrolysis of starch by comparing the concentration of glucose produced in one minute.  

So, lesser the volume of glucose solution used greater the concentration of glucose and faster the rate of hydrolysis of starch.

 

        

Concentration of glucose    α                               1                                                                                          

                                                   volume of glucose solution used                                           

    So by comparing the equations we can say,

        Rate of hydrolysis of starch   α                     1    

     ,                                                  volume of glucose solution used 

                                                 

   

 I invesigated the

1) Effect of change in pH on the rate of hydrolysis. (Temperature and starch concentration is kept constant)

2) Effect of change in temperature on the rate of hydrolysis. ( ph and starch concentration kept constant)  

3) Effect of change in the concentration of starch on the rate of hydrolysis. ( ph and temperature kept constant)

           ROUGH TRIALS

                             The first experiment I carried out was by using 0.5% diastase and 0.5% starch solution. I wasn’t able to get any reasonable titration reading because the glucose concentration was very low. The experiment was then carried out using 0.5% starch but 2% diastase solution. This time the experiment went successful. A reasonable titration reading was obtained.

                      Buffers of different pHs were added in the starch and diastase solutions to adjust the pHs. Water baths were used to adjust the temperatures. Different concentrations of starch were also used in order to investigate the rate of hydrolysis of starch at different pHs temperatures and starch concentrations.                                                                  

                           The enzyme and starch solutions adjusted at different pH's temperatures and the starch adjusted at different concentrations were allowed to react separately for one minute. after one minute I added five drops of iodine in each of the reaction mixture in order to stop the action of diastase as iodine is a very good inhibitor of diastase. Iodine works by binding to the starch. When iodine binds to the starch it will more than likely interfere with diastase activity (this effect is called steric hindrance, literally the iodine would block the starch from binding at a molecular level) so you will not get a clear indication of how well the diastase is working.

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                     After this step I ended up with a solution containing glucose in different concentrations. In order to measure the glucose concentrations I titrated this solution with quantitative Benedict solution in which sodium carbonate was added this titration was carried out at continuous boiling and stirring of quantitative benedict’s solution the colour change in this titration was from blue to cream colour. I carried out three titrations mostly and their mean was taken in order to represent the final reading.

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