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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
bath.
 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
Temperature
 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

pH
 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
acid)
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

Skills
 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