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Metropolitan Research Inc

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Testing for the macromolecules
(Carbohydrates, Proteins and Lipids)

Group Members
Sonovia Culmer
Welleana Ferguson
Ashley Morley
Jino Petit-Homme

Anatomy & Physiology I: Biol 131/ section 01
Lecturer: Mr. Woodrow Smith
Date: 11 February 2015

Introduction

Carbohydrates, proteins and lipids are all organic macromolecules composed of polymers which are made up of smaller simpler subunits (monomers). These macromolecules have unique characteristics that allow them to carry out complex functions.
In this lab, four (4) tests will be conducted: the Benedict test (for reducing sugar) BNT test; Iodine test (for starch) BIU test; Biuret test (for protein) IDN test; and Sudan III test (for lipids) SDN test.
The first test conducted would be the Benedict test. Carbohydrates include sugars, glycogen, starches and cellulose. Three substances will be tested for reducing sugars. These substances include sucrose, glucose and distilled water (H2O). If any of the substances are negative for non-reducing sugars it will have a blue color. If it positive for reducing sugar it will have a yellow to green to reddish orange color for weak to strong presence of the sugar respectively.
The Idoine test for starch would be the second test preformed. Staining by distinguishes starch from monosaccharides, diaaccharides and other polysaccharides. Two substances potatoes and distilled water will be tested for starch. If starch is present in any of the substances the iodine will turn it bluish-black. If the test is negative, the substance will have a yellowish brown color.
Peptide bonds in proteins are identified by a Biuret reagent which contains a 1% solution of copper sulphate. In the Biuret test, distilled water and Albumin solution will be tested for the present of protein. A violet color will reflect a positive presence of protein while a blue color gives a negative result.
The test for lipids is based on a lipid’s ability to selectively absorb pigments in fat-soluble dyes such as Sudan III. Sudan III stains fat droplets red.

Materials

General test tube rack micropipets distilled water
BNT test
3 test tubes thermometer 2ml of glucose test tube holder
2ml of sucrose beaker
2ml of Benedict solution bunsen burner
Timer
IDN test
2 test tubes iodine
2ml starch solution (potatoes) mortar & pestle
BIU test
2 test tubes
Albumin solution
Biurets solution
SDN test
1 test tube cooking oil
Sudan III

Methods
Benedict test (for reducing sugar) BNT test 1. Obtain 3 test tubes. 2. Add 2ml of glucose in one tube, 2ml of sucrose in one and distilled water in the other. 3. Add 2ml of Benedict’s solution the each tube. 4. Take temperature of boiling water. 5. Place both tubes in boiling water for 3mins. (note color change) 6. Let tubes cool and state which sugar is reducing or not. Why?
Iodine test (for starch) IDN test 1. Obtain 2 test tubes. 2. Place 2ml of distilled water in one and 2ml of starch solution in the other. 3. Add 3 drops of iodine to each tube and mix. Note the results.
Biuret test (for protein) BIU test 1. Obtain 2 test tubes. 2. Place 3ml of distilled water in one tube and 3ml of albumin solution in the other. 3. Add 3 drops of Biurets to both tubes and mix noting color change.
Sudan III (for lipids) SDN test 1. Obtain a test tube. 2. Add 3 drops of oil to 5 drops of water. 3. Mix contents of tube and let stand for a few minutes and note color separation.

Results
Benedicts test (for reducing sugars) BNT test
Sucrose is a non-reducing sugar. The test had a negative color reaction of blue.
Distilled water tested negative for reducing sugar. Color remains unchanged to blue.
Glucose tested positive for reducing sugar. After 2 minutes of being in the boiling water the color started to change to reddish orange. After 3 minutes the color changed to brick red.
Iodine test (for starch) IDN test
The starch solution (which was potatoes mashed in water) tested positive for starch. The color changed to a bluish black color confirming starch amylase is present.
The distilled water tested negative for starch. The color changed to yellowish brown.
Biuret test (for protein) BIU test The Albumin solution tested positive for protein resulting in a violet color change.
The distilled water tested negative for protein reflecting a blue color.
Sudan III test (for lipids) SDN tests
The Sudan III makes the oil more visible. The lipid layer clearly separates from the water layer. The water settles at the bottom and the fat droplets settle to the top forming a pinkish orange emulsion giving a positive test.

Conclusion
Many reducing sugars such as glucose and fructose have free aldehyde or ketone groups which can reduce weak oxidizing agents such as copper in Benedict’s reagents. The reason that sucrose is a non-reducing sugar is that it has no free aldehyde or keto group. Additionally, its anomeric carbon is not free and can’t easily open up its structure to react with other molecules.

Proteins are long chains of amino acids that are held together by peptide bonds. Free amino acids do not contain peptide bonds as they are the monomer building blocks of proteins. A peptide bond is link between two amino acids. Free amino acids are not like, therefore there is no bond.

Lipids are the most common class of hydrophobic molecules. Sudan III is not soluble in water but however it is soluble in lipids. Drops of lipid in emulsions are completely surrounded by water molecules. The detergents form a layer between the water and the lipids.
These tests (Benedict, Biuret, Iodine and Sudan III) can reveal hidden sugar, starch, protein and fats and their relative amount in various food items.

References

Tortora, Gerard, J. and Bryan, H. Derrickson. (2006). Principle of Anatomy and Physiology 11th edition. USA. John. Wiley & Sons, Inc. www.chacha.com www.brilliantbiologystudent.weebly.com www.ehow.com/facts_5882980_sucrose_non_reducing_sugar www.sciwebhfcc.net/biology/jababs/bio131/organic/organic.html

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