Biochemical Tests

Question 1

Explain the method for testing for a reducing sugar

Answer 1

1) add benedicts reagent (which is blue) to a sample and heat it in a water bath that’s been brought to the boil
2) if the test is positive it will form a coloured precipitate. The higher the concentrate of reducing sugar, the further the colour change goes, this ranges from green (not very much) to brick red (lots)

Question 2

Explain the method for testing for a non reducing sugar

Answer 2

1) if the result for the previous test is negative, and no coloured precipitate has formed, there could still be a non-reducing sugar present
2) take a new sample of the test solution and add dilute hydrochloric acid and carefully heat in a water bath that has been brought to the boil, then neutralise with sodium hydrogencarbonate
3) carry out the original benedicts test
4) if the test is positive it will form a coloured precipitate (as for the reducing sugar test), but if it is negative the solution will stay blue, meaning it doesn’t contain any sugar

Question 3

Explain the test used to detect starch in a sample

Answer 3

-add iodine solution to sample
-if starch is present it will change from a browny-orange to a blue-black colour

Question 4

Explain the test used to detect protein in a sample

Answer 4

-add biuret to the sample
-if positive sample with change from blue to a lilac-purple

Question 5

Explain the test used to detect lipids in a sample

Answer 5

Emulsion test:
-shake the test sample with ethanol for about a minute then pour it into water
-if lipid is present the solution will turn milky/ cloudy
-if no lipid the solution will stay clear

Question 6

Explain the steps for using a colorimeter and serial dilutions to determine the concentration of a glucose solution

Answer 6

To make the serial dilutions:
1) line up 5 test tubes in a rack
2) add 10cm cubed of the initial 40mM glucose solution to the first test tube, then 5cm cubed of distilled water to the other four test tubes
3) using a pipette, draw 5cm cubed of the solution from the first test tube and add it to the second, mixing thoroughly. You now have a 10cm cubed of solution which is half as concentrated as the first one
4) repeat this process 3 more times to create solutions of 10mM, 5mM and 2.5mM

Making a calibration curve:
1) do a benedicts test on each solution
2) remove any precipitate that was formed
3) use a colorimeter with a red filter to measure the absorbance of the benedicts solution remaining in each tube
4) use the results to make a calibration curve, showing absorbance against glucose concentration
5) this enables you to test the unknown solution in the same way as the known concentrations using the curve to find out its concentration