01 29 18 Structure and Functions of Macromolecules Lab PDF

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Destinee Logan Marie-Elaine Cousineau-Cote Grand Canyon University BIO-181L 29 January 2018 Structure and Functions of Macromolecules Lab Report Introduction to Experiment The Structure and Functions of Macromolecules Lab is intended to verify the presence of lipids, proteins, starches, carbohydrates, and monosaccharides/disaccharides in randomly selected liquids samples. The actual testing of the samples for the presence of these macromolecules consists of isolating the food and adding a particular solution/reactant to it. Each particular reactant should have a change of color when in the presence of the macromolecule that it was testing for, so it should be clear when there is a positive or negative reaction based on the color change. For example, the Benedict’s reagent and the Biuret reagent both have an obviously blue coloring to them, therefore the negative reaction would clearly be in occurance when the food that the reagent mixed with stays blue. There are also positive controls for each test that will cause a change of color of the solution when they come in contact with the reagent. Objectives The objectives of this lab are to determine/verify the presence of lipids, proteins, starches, carbohydrates, and monosaccharides/disaccharides in particular liquids. They do so using solutions/reactants whose individual purposes are to create a visual and chemical reaction when they come in contact with a particular macromolecule. Materials

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● Test tubes

● Unknown A

● Test tube racks

● Unknown B

● Potato extract

● Unknown C

● Glucose solution (+ control for Benedict’s test)

● Biuret reagent

● Starch solution (+ control for Lugol’s test)

● Benedict’s reagent

● Vegetable oil (+ control for grease spot test)

● Lugol's solution (IKI)

● Egg albumin solution (+ control for Biuret test)

● Filter paper

● A bottle of clear soda, such as lemon-lime

● Deionized water

● Corn syrup

● Beakers

● Sucrose solution

● Hot plate

● Nonfat milk

● Disposable pipettes

● Cream Procedures Benedict’s Test (carbohydrates/monosaccharides/disaccharides) 1. Record predictions on data table indicating either a positive or negative result. 2. Indicate the positive and negative controls. 3. Measure 100 mL of DI water out into a 250 mL beaker and heat the water to its boiling point using a hot plate. 4. Set aside 10 test tubes to use for the mixing of predetermined liquids with Benedict’s solution. 5. Fill each test tube with 1 mL of each predetermined liquid. 6. Add 1 mL of Benedict’s solution to each test tube. 7. Gently combine the solution and liquids together. 8. Using tongs, hold each tube (INDIVIDUALLY) in the beaker of boiling water to heat them, then allow them to cool. 9. Record any color changes in the test tubes and compare them to the negative and positive controls to verify if the result is positive or negative.

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Note: Blue (cool tones) = Negative result, Other colors (warm tones) = Positive result

Lugol's Test for Starch 1.

Record predictions on data table indicating either a positive or negative result.

2. Indicate the positive and negative controls. 3. Set aside 10 test tubes to use for the mixing of predetermined liquids with Lugol's solution. 4. Fill each test tube with 1 mL of each predetermined liquid. 5. Add 3-5 drops of Lugol's solution to each test tube. 6. Gently combine the solution and liquids together. 7. Record any color changes in the test tubes and compare them to the negative and positive controls to verify if the result is positive or negative. Note: Orange-brown = Negative result, Blue-black (w/visible precipitate) = Positive result

Biuret Test for Proteins 1.

Record predictions on data table indicating either a positive or negative result.

2. Indicate the positive and negative controls. 3. Set aside 8 test tubes to use for the mixing of predetermined liquids with Biuret solution. 4. Fill each test tube with 1 mL of each predetermined liquid. 5. Add 1 mL of Biuret solution to each test tube. 6. Gently combine the solution and liquids together. 7. Allow the contents to settle for 2 minutes. 8. Record any color changes in the test tubes and compare them to the negative and positive

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controls to verify if the result is positive or negative. Note: Light blue = Negative result, Violet-purple = Positive result.

Grease Spot Test 1. Divide 2 circular sheets of filter paper into 4 sections each and label each section with the name of the designated liquid that is going to reside in each section. There should be a total of 8 sections. 2. Indicate the positive and negative controls. 3. Using the droppers, distribute a drop of each sample into the corresponding sections on the papers. 4. Leave to sit for 30 minutes. 5. Wipe off any excess remaining that did not soak into the paper. 6. Leave to finish drying for another 30 minutes. 7. Hold the papers up to a light source and note any residues left behind in each section. Note the darkness of the residue and the diameter. 8. Record any residues on the papers and compare them to the negative and positive controls to verify if the result is positive or negative. Note: Residue = Positive Result, No residue = Negative result. Data Table 1 Results for Benedict’s Test Benedict’s Test

Food Item Prediction

Conclusion Result

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-

-

No change

2 – Glucose

+

+

Orange

3 – Corn syrup

+

+

Orange

4 – Sucrose

+

-

No change

5 – Soda

+

+

Orange

6 – Nonfat milk

+

+

Orange

7 – Cream

+

+

Yellow

8 – Unknown A

-

No change

9 – Unknown B

+

Green

10 – Unknown C

+

Green

1 – Deionized water

Table 2 Results for Lugol’s Test Lugol’s Test

Food Item

Conclusion

Prediction

Result

1 – Deionized water

-

-

Orange

2 – Starch

+

+

Blue

3 – Corn syrup

+

+

Blue

4 – Sucrose

-

-

Orange

5 – Potato extract

+

+

Black

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6 – Nonfat milk

+

-

Orange

7 – Cream

-

-

Orange

8 – Unknown A

+

Blue

9 – Unknown B

+

Blue

10 – Unknown C

-

Orange

Table 3 Results for Biuret Test Biuret Test

Food Item

Conclusion

Prediction

Result

1 – Deionized water

-

-

Blue

2 – Albumin

+

+

Purple

3 – Corn syrup

-

-

Blue

4 – Nonfat milk

+

+

Purple

5 – Cream

+

+

Purple

6 – Unknown A

-

Blue

7 – Unknown B

+

Purple

8 – Unknown C

+

Purple

Table 4 Results Grease Spot Test Grease Spot

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Food Item

Conclusion

Test Prediction

Result

-

-

No residue

2 – Vegetable oil

+

+

Lg/Dense Spot

3 – Soda

+

-

No Residue

4 – Nonfat milk

+

+

Light residue

5 – Cream

+

+

Faint residue

6 – Unknown A

+

“Oily” residue

7 – Unknown B

+

“Oily” residue

8 – Unknown C

-

No residue

1 – Deionized water

Table 5 Macromolecule Tests on the Unknowns Item

Benedict’ s Test

Lugol’s Test

Biuret Test

Grease Spot Test

Possible Foods

-

+

-

+

cornbread

+

+

+

+

chips

+

-

+

-

protein bar

Unknown A Unknown B Unknown C

Revealed Food

Analysis Benedict’s Test In the case of the Benedict’s Test, an indication for the presence of a reducing sugar

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would be a color change of the solution from a light blue to some type of orange or yellowishorange color. This test works at the molecular level in that the solution contains copper and the copper interacts with a particular sugar called aldose which contains an aldehyde group. This causes the reaction in the form of a color change. The obtained results do agree with the predictions for the most part with the exception of Sucrose. It was assumed that sucrose would cause a positive result because it is a type of sugar but because it contains no free aldehyde group, it does not interact with copper. It seems that sucrose was included in the test to display that there is a type of sugar out there that does not respond to the reactant. Although, when Sucrose is heated with hydrochloric acid it separates into glucose and fructose which are then detectable by the reactant since the aldehyde group then becomes free. Lugol’s Test In the case of the Lugol’s Test, an indication for the presence of starch would be the test turning blue/black (and precipitate can form). Lugol’s solution contains potassium iodide (KI) (orange/yellow), which turns to blue/black when it is introduced to starch due to the reaction of amylose portion with potassium iodide. The results obtained do agree with the predictions with the exception of non-fat milk as it was assumed that all milks contain some type of starch based addition and this is an incorrect assumption. Biuret Test In the case of the Biuret Test, an indication for the presence of protein would be the changing of color of the reactant from its’ initial light blue color to a shade of violet/purple. The test relies on the reagent, composed of potassium hydroxide and copper sulfate, to turns violet in the presence of peptide bonds which are the chemical bonds that hold amino acids together. The results of obtained do agree with the predictions as the presence of proteins in the selected foods

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are relatively easily assumed based on average prior knowledge of proteins. Grease Spot/Brown Paper Test In the case of the Grease Spot/Brown Paper Test, an indication for the presence of fat would be a remaining greasy/oily residue left behind a liquid after the evaporation of the water molecules. The way that the Grease Spot/Brown Paper Test works is that when samples are allowed to dry on filter paper or a brown paper bag, the water will evaporate, while the fats/lipids remain. The results obtained do agree with the predictions with the exception of soda as it was assumed that there would be some kind of lipid based product added into the soda. General Questions If a positive result were obtained for a negative control, possible conclusions could be that the test was completed incorrectly, the sample could have been contaminated, or the subjected liquids/reactants could have been confused. If a negative result were obtained for positive control, possible conclusions could be that the test was completed incorrectly, the sample could have been contaminated, or the subjected liquids/reactants could have been confused. The biological molecules in Unknown A were starches and lipids. This led to the idea that the food had to be some kind of corn or potato based product and the former seemed like the best option therefore the proposed unknown was cornbread. The biological molecules in Unknown B were sugars, proteins, starches and lipids. This led to the idea that the food had to be something considered either “unhealthy” and processed therefore coupled with the obvious look of the substance the proposed unknown was some kind of corn or potato based chip. The biological molecules in Unknown A were sugars and proteins. This led to the idea that the food had to be some kind of meat, nut, bean or some other heavily protein based product. The liquid itself

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looked like some kind of separated protein drink and because protein bars fit the positively indicated macromolecules the unknown was proposed to be some kind of protein power or ground up protein bar. The unknowns were not revealed prior to the completion of the experiment. Conclusion The Structure and Functions of Macromolecules Lab used Benedict’s reactant, Lugol’s reactant, Biuret reactant and filler paper to determine the presence of the macromolecules: sugar, starch, protein, and lipids in particular foods. Out of the tested foods, predictions for the presence of particular macromolecules were easily assumed with the exception of a couple items such as sucrose. This experiment helps display the real-life presence of macromolecules in commonly purchased and consumed foods.

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References Reece, J. B., Urry, L. A., Cain, M. L., Wasserman, S. A., & Minorsky, P. V. (2017). Campbell Biology (11th ed.). Boston, MA: Pearson. Areda, D., Boyles, R., Francis, G., & Hite, A. (n.d.). Academic Web Services - Grand Canyon University. Retrieved January 30, 2018, from https://lc.gcumedia.com/bio181l/laboratorymanual-for-general-biology-i/v2.1/#/chapter/1...