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Testing for Biomolecules

(Food Tests)
Benedict’s Test for Reducing Sugar
 Food needs to be crushed and added with water to make a suspension.
 Pour 2cm of FOOD into a test tube.
 Add 2cm of Benedict’s solution (blue) into the test tube.
 Shake the mixture.
 Heat the test tube for 5 mins in a boiling water bath.
 Record the colour changes that take place in the test tube while it is being heated in the boiling water
 An orange-red precipitate is observed.
 The mixture remains blue.
 Reducing Sugars: Glucose, Fructose, Galactose

Iodine Test for Starch

 Place a small quantity of food (if liquid, 1 drop) on a white tile.
 Add 2 drops of dilute iodine solution.
 Describe the change in colour observed.
 The mixture turns blue-black.
 The mixture remains brown.

Alcohol Emulsion Test for Fats

 Add 1 drop of food to a dry test tube.
 Add 3cm of alcohol.
 Shake the mixture thoroughly for about 5 minutes.
 Add the same amount of water to the test tube.
 Record the changes observed.
 An opaque white emulsion is observed (cloudy)
 No opaque white emulsion is observed (clear)

Biuret Test for Proteins

 Food needs to be crushed and added with water to make a suspension.
 Place about 2cm of food in a test tube.
 Add 2cm of dilute sodium hydroxide solution.
 Shake the mixture.
 Add about 3 drops of 1% copper sulphate solution, and shake after each drop.
 Describe the change in colour observed.
 The solution turns violet.
 The solution remains light blue.
Investigating the Effect of
Temperature and pH on
Enzyme Action
 Label the test tubes A1 and A2, B1 and B2 etc.
 Add 5cm of starch solution to test tubes A1 and B1.
 Add 3cm of distilled water to A2 and 3cm of amylase solution to B2.
 Place all the test tubes into a beaker of water at 37C.
 Leave the test tubes for 10 minutes to allow the solutions in the tubes to reach the temperatures of the
water bath.
 Pour the contents of A2 into A1 and B2 into B1. Keep A1 and B1 in their respective water baths
throughout the experiment and ensure that the temperatures are maintained.
 After 10 minutes, test the mixture in each tube with iodine solution, on a white tile.
o At 37 C: Optimum temperature, digests starch (eg amylase digests starch into maltose, so when
tested with iodine test turns brown)
o At 0 C: Inactive, unable to digest starch
o At 100 C: Denatured, unable to digest starch
o Water control test tube: no amylase present – serves as a control to prove that starch is digested
by amylase and not broken down spontaneously
o Next step: Benedict’s Test to ensure that it is maltose = reducing sugar

 Add 2cm of starch solution to test tubes W, X, Y, Z.
 Place them in a water bath at 37C.
 Make up the contents of the 4 test tubes as shown in the table below.
 Test the solution with iodine solution at the start of the experiment and after 10 minutes.

Test Tube Contents Colours observed in Iodine Test Conclusion

Before After
W 2cm starch solution Blue-black Blue-black Starch is present
2cm HCL (hydrochloric
2cm amylase
X 2cm starch solution Brown Brown Starch is absent
2cm distilled water
2cm amylase
Y 2cm starch solution Blue-black Blue-black Starch is present
2cm NH4OH
(ammonium hydroxide)
2cm amylase
Z 2cm starch solution Blue-black Blue-black Starch is present
4cm distilled water

o Acid: Denatures enzymes

o Alkali: Denatures encymes
o Neutral: Ideal pH for enzyme action
o Water: no amylase present

Extreme temperatures and pH break bonds and alter the 3D shape of enzymes, so the active site is unable
to access substrates and no enzyme action can occur

 Label all test tubes
 Label droppers for test tubes to avoid contamination of solutions in each test tube with solutions from
other test tubes
 Label white tiles to avoid contamination
 Wash white tile after every iodine test conducted
 Wear goggles during heating process and handling of acid/alkali
 Temperature of water baths maintained at stated temperature
 Do not use thermometer as stirrer, do not leave inside beaker during heating
 Measure temperature at eye level
 Rinse test tubes, knife and cutting surface before use
 Light Bunsen burner correctly
 Level of water in water bath slightly more than half
 Water bath heated over a tripod with a non-luminous flame
 Test tubes shaken before placing in the boiling water bath
 Level of water bath higher than level of mixture in test tube
 Test tube holder used to hold hot test tubes
 White tile washed and wiped dry before use
 Tables
o Vertical columns (independent variable) headed
o Horizontal rows (dependent variable) headed
o Units stated (eg C / min)
o Colours accurately recorded
o Results recorded in 1 table
o Appropriate title
o Use ruler to draw lines
o Overall layout clear