A2 Planning-effect of PH on digestion of starch by amylase

[harmony]

Hi, I'm doing my planning on the effect of PH on the rate of digestion of starch to maltose and dextrins by the enzyme amylase and I am a little unsure about what concentrations I should use.

I was thinking about using 10% amylase solution and 10% starch solution but I was wondering whether it would be better to use a greater concentration of starch solution.

Also, I was thinking about setting the thermostatically controlled water bath at 35 degrees C as my teacher said that the optimum temperature for amylase was 37 degrees C and so to be on the safe side, choose a temperature lower than this.

Any replies would be read very gratefully. Thank You.

[Happydadtoo]

Very very quickly.....
1% starch is as strong as you will get - ditto for the amylase, which is VERY quick-acting - you MIGHT even need 0.1%
Warm pH buffer separately, then add to starch solution
Warm this mixture AND the amylase SEPARATELY in the water-bath, then mix at time 0
Expect complete hydrolysis in 30s - 120s @ph 7.0 - no reaction at all below 5 and above 8, so use narrow range pH buffers
Note - stop the reaction by adding a few drops FROM the mixture in the w/b TO a spotting-tile with I2/KI soln in it
Repeat every 30s or so - determining the EXACT end-point is the major problem with thsi expt - comment in analysis.

ALTERNATIVE:
Use coplorimeter to determine end-point, BUT in that case you will need to use 1cm3 samples (so bigger vols needed at start) and mix with 0.5 or 1.0 cm3 of I2/KI solution.
Set filter to YELLOW and measure transmission v 'blank' of I2/KI soln and water (which should be 100%)
Better, more accurate, but needs more materials!!
Must dash - off to sydney.....
HD2