Carbohydrates and Proteins Lab Overview

Title: URL Source: blob://pdf/a5967803-cf5f-4e9e-a293-381fdfc5d131 Markdown Content: Chem 100, Spring 2025 Pre -lab 15 1 # Experiment 15 # Carbohydrates and Proteins Chem 100, Spring 2025 Pre -lab 15 2 ## Purpose To become acquainted with some of the properties of and standard chemical tests for carbohydrates and proteins. ## Materials Reagents and equipment you will need this week: Parchment paper or foil Spray oil (nonstick spray) Large cookie sheet saucepan cup sugar 4 tablespoons light corn syrup or honey 1 teaspoons baking soda Stove or microwave Pot or microwave -safe container to boil water Timer 1 scissors to cut the egg whites 1 mixer/hand blender or wire whisk to whip egg whites 1 bowl, 1 Large bowl Forks, Spoons, Knife (for cutting fruit) 6 small containers of about the same size. 2 eggs (split egg white into six parts); additional can be used (You can cut the egg white into pieces with your scissors.) 1/3 cup of rubbing alcohol (75 ml) 1/3 cup of white vinegar (75 ml) Gelatin flavored or unflavored 18 dixie cups (or a 12 spot muffin tray.) Plastic wrap Universal indicator strip or red cabbage strip indicator strip Table salt Ammonia (base) or Window cleaner (base) or Baking Soda (base) or Toilet cleanser (base) Vinegar (acid) Lemon Juice (acid) Meat Tenderizer Fruits and/or fruit juice: Pineapple fresh, frozen and canned, please try all 3. Papaya or Fig or Kiwi ( Choose one ). Apples or Grapes or Strawberries or Orange or other fruit (Choose 2) Chem 100, Spring 2025 Pre -lab 15 3 ## Quiz Preparation Vocabulary (terms you may see on the quiz) monosaccharide disaccharide polysaccharide carbohydrates triose tetrose pentose hexose ketose aldose reducing sugar glycosidic linkage ## Introduction/Background Carbohydrates: Carbohydrates serve as the primary chemical energy source for almost every form of life, and are therefore fundamental compounds in living organisms. Carbohydrates may be classified as monosaccharides, disaccharides, or polysaccharides. Classes of Carbohydrates > Monosaccharide A monosaccharide (or simple sugar) is a polyhydroxy (many hydroxyls) carbon compound that contains a carbonyl group. A monosaccharide may be classified as a triose, tetrose, pentose, or hexose, depending on the number of carbon atoms present. Each carbon, except for the carbonyl carbon, has a hydroxyl attached to it. It is these that give the physiological sensation of sweetness. Also, a carbohydrate may be classified as an aldose (monosaccharide with an aldehyde functional group) or a ketose (monosaccharid e with a ketone group) depending on the location of the carbonyl carbon. Three monosaccharides with which you will become familiar are glucose, galactose, and fructose. All three of these are hexoses which have the molecular formula C 6H12 O6 and Glucose are aldoses, while fructose is a ketose. > Disaccharides A disaccharide is a compound that is composed of two monosaccharides held together by a glycosidic linkage, which is actually an ether linkage. In the presence of acid or the appropriate enzyme, the glycosidic linkage may be hydrolyzed, producing the two monosaccharides. Three important disaccharides are maltose, lactose, and sucrose. Maltose is composed of two glucose; lactose is composed of glucose and galactose, and sucrose is composed of glucose and fructose. Sucrose is common table sugar. Chem 100, Spring 2025 Pre -lab 15 4 > Polysaccharides A polysaccharide is a polymer (a very long chain) of monosaccharides, each one held to the next one by a glycosidic linkage. Like disaccharides, polysaccharides may be hydrolyzed in the presence of acid or enzymes to produce monosaccharides. Three importan t polysaccharides are starch, glycogen, and cellulose. All three of these are polymers of the monosaccharide glucose. Starch is composed of two polysaccharides a branched polysaccharide called amylopectin, and a straight -chain polysaccharide called amylose . Reducing Sugars A reducing sugar is a monosaccharide or disaccharide which can be oxidized by Benedict's reagent. Any monosaccharide or disaccharide that has an available aldehyde functional group, or an -hydroxy (read as alpha -hydroxy) ketone functional group (a functional group in which a hydroxyl is on the carbon attached to the carbonyl carbon of a ketone), can be oxidized. It is called reducing sugar because it reduces (the opposite of oxidizes) the r eagent that causes the oxidation. All monosaccharides and most disaccharides are reducing sugars. Watch the video below to help understand the theory behind Benedict's Test for Sugars. In today's experiment, you will test several carbohydrates to determine which are reducing sugars. A carbohydrate that gives a positive test --the blue solution tums green and then a brick -red precipitate forms --is a reducing sugar. Iodine Test for Starch When an aqueous solution of iodine, I 2, is added to a starch solution, the iodine solution changes color from brownish orange to dark blue -black. This does not happen with any other carbohydrate. This is because one of the two polysaccharides of which starch is composed, amylose, is a long unbranched chain of glucose molecules which is twisted into a coil called a helix. When th e iodine solution Is added to the starch solution, the iodine gets trapped in the amylose coil and the blue -black color is observed. Chem 100, Spring 2025 Pre -lab 15 5 Hydrolysis In hydrolysis reactions of carbohydrates, the disaccharides and polysaccharides are broken down into monosaccharides. This changes the chemical reactivity. We will use sucrose and starch to model the effect of hydrolysis on the chemical properties of dis accharides and polysaccharides. The first step will be to hydrolyze the sample, acid hydrolysis will be used. Then the samples will be tested with the Benedict's and starch iodine tests. Afterward, you will answer some associated questions. Proteins: A polypeptide is a polymer (a very long chain) of amino acids. There are about twenty different amino acids. One amino acid in the polypeptide chain is held to the next one by a peptide bond, which is the name given to an amide linkage between two amino acids. The polypeptide chain is coiled and folded up in a manner that is dependent on the locations of the various amino acids in the polypeptide chain. A protein is composed of one or more polypeptide chains. Amino acids The twenty common amino acids all have the same basic structure: They differ only in their R groups (side chains). Color Tests for Proteins A color test for protein tests for a particular functional group or structural component present in a protein or in a particular amino acid found in proteins. > Biuret Test When the biuret test is performed on a compound that contains two or more peptide bonds, a violet color appears. Since all proteins contain peptide bonds, all proteins give positive Biuret tests. > Xanthroproteic Test Xanthroproteic test is specific for the side chains of three amino acids: tyrosine, tryptophan, and phenylalanine. Each of these has a side chain that contains a benzene ring with either an amino group or a hydroxyl group attached to it. When the Xanthropr oteic is positive, a yellow color appears. Chem 100, Spring 2025 Pre -lab 15 6 Protein Denaturation The polypeptide chain(s) of a protein needs to be folded in order for the protein to be able to function. To denature a protein means to cause the polypeptide chain(s) to unfold. A denatured protein cannot carry out its biological function, and often a den atured protein becomes less soluble (it coagulates). Certain reagents or conditions will denature proteins. The addition of acid, base, ethanol, tannic acid and heavy metal ions such as mercury, silver, and lead will denature a protein. Heating a protein w ill also cause it to denature. These cause denaturation by interfering with the forces that are keeping the polypeptide chain(s) folded. > The Effects of Denaturing on Egg Albumin: Here are some brief explanations for what you see: Control. Egg whites start out clear. They are almost 90% water, but the other 10% is packed with proteins. Egg whites contain more than 50% of the proteins found in the egg. The main protein in egg white is called albumin. The small, folded proteins in the egg whi te dont take up much space, and the gel -like egg white looks clear. The control egg showed us that, when left at room temperature, the egg whites stay clear, meaning the proteins maintain their original shape. These proteins were not denatured. Physical . When you beat raw egg whites to make a souffl or a meringue, you incorporate air bubbles into the water -protein solution. Adding air bubbles to egg whites unfolds those egg proteins just as certainly as heating them. To understand why introducing air bu bbles makes egg proteins uncurl, you need to know a basic fact about the amino acids that make up proteins. Some amino acids are attracted to water; theyre hydrophilic, or water -loving. Other amino acids are repelled by water; theyre hydrophobic, or wate r-fearing. Egg -white proteins contain both hydrophilic and hydrophobic amino acids. When the protein is curled up, the hydrophobic amino acids are packed in the center away from the water and the hydrophilic ones are on the outside closer to the water. Whe n an egg protein is up against an air bubble, part of that protein is exposed to air and part is still in the water. The protein uncurls so that its water -loving parts can be immersed in the water and its water -fearing parts can stick into the air. Once t he proteins uncurl, they bond with each other just as they did when heated creating a network that can hold the air bubbles in place. Room temperature water. Sometimes in this experiment, room temperature water has a small denaturing effect on some of the egg white. It acts in the same way, by breaking bonds, but its effect isnt nearly as strong as alcohol or hot water. Chem 100, Spring 2025 Pre -lab 15 7 Alcohol. Alcohol also denatures proteins. It does this the same way as heat, by breaking the bonds that hold parts of the protein in a folded shape. Sometimes the alcohol molecules bond directly to some of the parts of the protein, disrupting the normal way the pr otein would bond to itself. (So alcohol is called a bond disruptor.) The proteins unfolded, taking up more space and hardening in place next to one another. This process takes much longer with alcohol than it does with heat. The longer time for denaturat ion with alcohol is simply because it spreads more slowly than heat. The alcohol has to diffuse (or move through the fluid) into the egg in order to affect the proteins it touched. Changing pH (vinegar) . Changes in pH change the attractions between the groups in the side chains of the protein. The interactions between the side chains of the amino acids determine the shape of a protein. When these interactions are disrupted, the protein unfolds. Cooking (hot water). Whenever eggs are cooked with heat, the egg whites turn from clear to white, and the gel becomes more rubbery. As heat -denatured the proteins in the egg white, it broke apart some of the bonds (mostly hydrogen bonds) that were holding the proteins in thei r original shape. The proteins unfolded, taking up more space (turning the gel white) and hardening them in place next to one another. > Denaturing and Enzyme Activity: The final portion of the experiment will look at how denaturing affects the activity of an enzyme, bromelain. Active bromelain will destroy other proteins, like the collagen found in gelatin. Enzymes designed to hydrolyze proteins are called proteases. Whe n the proteins in the pineapple are denatured, the enzyme is deactivated and has no effect on the gelatin. Some other fruits have a similar effect on gelatin because they also have enzymes that denature proteins. You will determine which fruits may have pr oteases in them and which do not. Video of Egg Albumin: This video gives an example and explanation of this portion of this experiment. Video of Pineapple and Gelatin: This video gives an example and explanation of this portion of this experiment. Chem 100, Spring 2025 Pre -lab 15 8 ## Station A: Benedict's Test for Sugars 1. Each of the sugars has been dissolved in water to make a 1% sugar solution. 2. Add a sugar solution to the labelled test tube as indicated. a. To tube A add ~ 1mL of glucose b. To tube B add ~ 1mL of fructose c. To tube C add ~ 1mL of sucrose d. To tube D add ~ 1mL of lactose e. To tube E add ~ 1mL of starch f. To tube F add ~ 1mL of hydrolyzed sucrose g. To tube G add ~ 1mL of hydrolyzed starch 3. Add ~ 1mL of Benedicts reagent to each test tube. 4. Place the test tubes in a hot water bath to speed up the reactions. 5. Allow the reaction to heat for about 5 minutes. 6. Observe the test reactions 7. Record the results in the table in your lab report. 8. Complete the table by indicating whether each compound is a reducing sugar or nonreducing sugar. 9. Take a photo of your test results and place it in your lab report. Results of Benedicts test Positive Test: If the color changes to muddy green/brown, the Benedicts test is positive, which means reducing sugar exists in the solution. Negative Test: If you find that no color change is visible, the result is negative and indicates that non -reducing sugar is present in the solution. Tube Sugar Name Benedict Color Reducing 1 glucose ***** Reducing or nonreducing 2 fructose ***** Reducing or nonreducing 3 sucrose ***** Reducing or nonreducing 4 lactose ***** Reducing or nonreducing 5 starch ***** Reducing or nonreducing 6 hydrolyzed sucrose ***** Reducing or nonreducing 7 hydrolyzed starch ***** Reducing or nonreducing Chem 100, Spring 2025 Pre -lab 15 9 Benedict's Test for Sugars Photo Insert photo here. Shrink photo to fit within this box. Benedict's Test for Sugars Questions 1. How do you know if you have a positive result in the Benedict's test? Write Your Answer Here. 2. What is the name of the functional group that must be available to give you a positive Benedicts test? (not the class of compound) Write Your Answer Here. 3. Do all monosaccharides give positive Benedict's tests? Choose one: Yes or No. 4. Do all disaccharides give positive Benedict's tests? Choose one: Yes or No . 5. Does hydrolyzed sucrose give positive Benedict's tests? Choose one: Yes or No . 6. What are the two monosaccharides produced when sucrose is hydrolyzed? Write Your Answer Here. 7. Did sucrose which was not hydrolyzed give a positive or negative Benedict's test? Choose one: positive or negative 8. Did hydrolyzed sucrose give a positive or negative Benedict's test? Choose one: Yes or No 9. Explain the difference in the results of these two Benedict's tests. Choose one: Yes or No 10. Does hydrolyzed starch give positive Benedict's tests? Choose one: Yes or No . 11. Do all polysaccharides give positive Benedict's tests? Choose one: Yes or No .Chem 100, Spring 2025 Pre -lab 15 10 ## Station B: Iodine Test for Starch These are the directions for the starch iodine experiment: Each of the sugars has been dissolved in water to make a 1% sugar solution. 1. Add a sugar solution to the labelled spot plate as indicated. a. To spot A add ~ 1mL of glucose b. To spot B add ~ 1mL of fructose c. To spot C add ~ 1mL of sucrose d. To spot D add ~ 1mL of lactose e. To spot E add ~ 1mL of starch f. To spot F add ~ 1mL of hydrolyzed sucrose g. To spot G add ~ 1mL of hydrolyzed starch 2. Add about 2 drops of iodine solution to each test tube. 3. Record your observations in the table below. 4. Record the results in the table in your lab report. 5. Indicate whether starch was present in each sample in the table in your lab report. 6. Take a photo of your test results and place it in your lab report. Results of Iodine test: Positive Test: If the color changes to blue or black, the iodine test is positive, which means the starch amylose coil is present in the solution. Negative Test: If you find that no color change occurs (or the solution remains a pale yellow, the result is negative and indicates that no starch is present in the solution. Sample Table for Iodine Test Tube Sugar Name Iodine Color Starch present 1 glucose ***** Yes or No 2 fructose ***** Yes or No 3 sucrose ***** Yes or No 4 lactose ***** Yes or No 5 starch ***** Yes or No 6 hydrolyzed sucrose ***** Yes or No 7 hydrolyzed starch ***** Yes or No Chem 100, Spring 2025 Pre -lab 15 11 Image: Iodine Test: Insert photo here. Shrink photo to fit within this box. Iodine Test for Starch Questions 1. What color is produced when the iodine test is positive? Write your Answer Here. 2. What form of starch causes the reaction with iodine? Write your Answer Here. 3. What structural component makes this form of starch the only polypetide to react with iodine? Write your Answer Here. 4. Do all monosaccharides give positive iodine tests? Choose one : Yes or No. 5. Do all disaccharides give positive iodine tests? Choose one : Yes or No . 6. Does hydrolyzed sucrose give positive iodine tests? Choose one : Yes or No . 7. Do all polysaccharides give positive iodine tests? Choose one : Yes or No . 8. Does hydrolyzed starch give positive iodine tests? Choose one : Yes or No . 9. Does a hydrolyzed starch solution give a positive test? Choose one : Yes or No 10. What monosaccharide is produced when starch is completely hydrolyzed? Write Your Answer Here. 11. What disaccharide is produced when starch is partially hydrolyzed? Write Your Answer Here. 12. What color should the disaccharide that is produced when starch is partially hydrolyzed give in the Benedict test? Write Your Answer Here. 13. What color should the disaccharide that is produced when starch is partially hydrolyzed give in the iodine test? Write Your Answer Here. Chem 100, Spring 2025 Pre -lab 15 12 Table: Sugar Results Summary Tube Sugar Name Structure Mono, Di or Poly Aldose, Ketose, or neither Benedict Color Reducing or nonreducing Iodine Color Starch present? 1 glucose Mono Di or Poly A, K or neither ***** R or NR ***** Yes or No 2 fructose Mono Di or Poly A, K or neither ***** R or NR ***** Yes or No 3 sucrose Mono Di or Poly A, K or neither ***** R or NR ***** Yes or No 4 maltose Mono Di or Poly A, K or neither ***** R or NR ***** Yes or No 5 starch Mono Di or Poly A, K or neither ***** R or NR ***** Yes or No 6 hydrolyzed sucrose MonoDi or Poly A, K or neither ***** R or NR ***** Yes or No 7 hydrolyzed starch Mono,Di or Poly A, K or neither ***** R or NR ***** Yes or No Chem 100, Spring 2025 Pre -lab Experiment 15 13 ## Station C: Color Tests for Proteins These are the directions for the below experiment: 1. Number 4 clean, dry test tubes 1 through 4. 2. Add about 2 mL of albumin solution to each of tubes 1 & 2. 3. To tubes 3 & 4 add about 2 mL of gelatin. 4. Perform the following tests on the protein solutions as follows. 5. Dispose of all waste in the provided waste containers. > Biuret Test Perform the Biuret test on the protein samples in tubes 1 and 3 ONLY: 1. Add about 1 mL of the copper (II) sulfate solution to each of the two test tubes 2. Add a few drops of the sodium hydroxide solution to each of the two test tubes 3. Mix well by spanking. 4. Record the color(s) you observe in the table in your lab report. > Xanthoproteic Test Perform the Xanthoproteic test on the protein samples in tubes 2 & 4 only. 1. Add about 1 mL of the concentrated nitric acid solution to each of the two test tubes 2. Mix well by spanking. 3. Place the two test tubes in hot (not boiling) water for about 3 minutes. 4. Carefully add about 3 mL of ammonium hydroxide solution to each of the two tubes. 5. Mix well. 6. Record the color(s) observed in the table in your lab report. > Results of Tests 1. Record the results in the table in your lab report. 2. Take a photo of your test results and place it in your lab report. Chem 100, Spring 2025 Pre -lab Experiment 15 14 Table: Protein test > Test Test tube # > Albumin > Color > Albumin > Test tube # > Gelatin > Color > Gelatin > Biuret 1******* 3******* > Xanthoproteic 2******* 4******* Image: Protein Color Tests Insert photo here. Shrink photo to fit within this box. Protein Color Tests Questions Answer the following questions related to the above table. a. What functional group present in proteins gives a positive Biuret test? Write Answer Here b. Which proteins contain this functional group? Write Answer Here c. What is the side chain of this amino acid? Write Answer Here d. What are the names of the three amino acids which give a positive Xanthoproteic test? Write Answer Here Chem 100, Spring 2025 Pre -lab Experiment 15 15 ## Station D: Denaturing Egg Albumin and Physical Properties Procedure 1. In a spot plate, half fill 5 spots with egg albumin a. To Spot 1, do nothing to those egg whites. b. To Spot 2, add room temperature water. c. To Spot 3, add rubbing alcohol. d. To Spot 4, add vinegar e. To Spot 5, add boiling (or very hot) water. 2. Observe any immediate changes that occur in terms of egg white color and consistency. 3. Record your observations in the table in your Lab Report under the heading "Observations Initially". 4. If you try stirring the different treatments, be sure to use a clean stir stick for each sample. 5. Wait for ~5 minutes. 6. Use a toothpick to inspect the state of the egg whites in each treatment and note how they may have changed over time. 7. Record your observations in the table in your Lab Report under the heading "Observations After 30 Minutes". 8. Indicate whether each sample was denatured after 30 minutes. 9. Take a photo of your experiment and post it in your Lab Report. 10. Answer the questions in your Lab Report. 11. When you are done, filter the liquid through a paper towel and dispose of the filtrate (liquid) in a sink with plenty of water. The paper towel and solid can go into in the trash. Table: Denaturing Egg Albumin > Spot Conditions Observations > Initially > Observations > After 30 Minutes > Was it > denatured? > 1Nothing added ******* ******* Y or N > 2Water (room > temperature) ******* ******* ******* > 3Rubbing alcohol ******* ******* ******* > 4Vinegar ******* ******* ******* > 5Boiling water ******* ******* ******* Chem 100, Spring 2025 Pre -lab Experiment 15 16 Image: Denaturing of Egg Albumin: Insert photo here. Shrink photo to fit within this box. Denaturing Proteins Questions 1. What happens to the polypeptide chain(s) when a protein is denatured? Write Answer Here 2. What effects does denaturation have on the physical and chemical properties of a protein? Write Answer Here 3. When a protein is denatured, are the peptide bonds broken (hydrolyzed)? Write Answer Here Chem 100, Spring 2025 Pre -lab Experiment 15 17 ## Station E: Denaturing and the effect on Biological Activity 1. Treat each of the spots in the provided spot plate partially filled with gelatin as directed below: a. Do not place anything but gelatin in container 1. b. Sprinkle some salt/pineapple juice mixture in container 2. c. Pour some rubbing alcohol/pineapple juice mixture in container 3 d. Sprinkle some meat tenderizer/pineapple juice mixture in container 4. e. Pour the acid/pineapple juice mixture in container 5. f. Pour the base/pineapple juice mixture in container 6. g. Place the juice of some heated fresh pineapple in container 7. h. Place the juice of some fresh pineapple in container 8. i. Place the juice of some frozen pineapple in container 9. j. Place the juice of some canned pineapple in container 10. k. Place the juice of some papaya, fig or kiwi in container 11. l. Place the juice of another fruit in container 12. 2. Make an immediate observation. Record your observation under "Initial Observation" in the table in your Lab Report. 3. Test the pH of each sample. Record your observation under "pH" in the table in your lab report. 4. Observe the experiment after 5 minutes. Record your observation under "5 -Minute Observation" 5. Take a picture of your gelatin samples (1 picture with all the samples) put it in your lab report. 6. Indicate whether each of the samples seemed to affect the gelatin. 7. Choose from the following to write a brief explanation for your observation. 8. Record your explanation under "Did the sample affect the gelatin?" in the table below using the provided responses. For each sample in the table below, Choose 1: a. broke down the gelatin because an active protease was present b. did not affect the gelatin because the protease that is present in pineapple was denatured. c. did not affect the gelatin because there was no protease present. d. did nothing since nothing was added, so it is a control. 9. Answer the questions in your Lab Report: Chem 100, Spring 2025 Pre -lab Experiment 15 18 Image: Gelatin After Treatment with Various Substances. Insert photo here. Shrink photo to fit within this box. Chem 100, Spring 2025 Pre -lab Experiment 15 19 Table: The Effect of Denaturing on the Enzyme Activity of Bromelain in Pineapple. Conta iner Sample pH Initial Observation 30 -minute Observations Gelatin affected? Explanation for why there was/was not an effect 1 Control ** ******** ******* Yes or No Choose one: Heat was added, pH was changed, less polar solvent used, heavy metal added) 2 Salt+ pineapples ** ******** ******* Yes or No ******* 3 Alcohol + pineapple ** ******** ******* Yes or No ******* 4 tenderizer + pineapple ** ******** ******* Yes or No ******* 5 Acid + pineapple ** ******** ******* Yes or No ******* 6 Base + pineapple ** ******** ******* Yes or No ******* 7 heated pineapple ** ******** ******* Yes or No ******* 8 Fresh pineapple ** ******** ******* Yes or No ******* 9 Frozen pineapple ** ******** ******* Yes or No ******* 10 Canned pineapple ** ******** ******* Yes or No ******* 11 Kiwi/fig/papaya ** ******** ******* Yes or No ******* 12 Apple/orange/gra pe ** ******** ******* Yes or No ******* Chem 100, Spring 2025 Pre -lab Experiment 15 20 Denaturing and the effect on Biological Activity Questions 1. How could the canning process change bromelin? Write Answer Here 2. How could the cooking process change bromelin? Write Answer Here 3. What could you do to the fresh pineapple that would allow the gelatin to become firm? Write Answer Here 4. Why do you think it might make a difference if you use solid pieces of fruit instead of fruit juice? Write Answer Here 5. Changes in temperature or pH can change the shape of an enzyme. Why will an enzyme no longer work if it loses its shape? Write Answer Here An experiment was performed to test the effect of temperature and pH on the activity of Enzyme X. The following data was collected during the experiment: 6. What is the optimum pH of enzyme X? Write Answer Here 7. What is the optimum temperature of enzyme X? Write Answer Here 8. Why do you think enzyme X has low activity at a pH of 10? Write Answer Here 9. Enzyme X performs critical life functions. Use the data above to explain why a fever of 40 degrees Celsius may be dangerous. Write Answer Here ## You have completed this Pre -lab assignment .

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Chem 100, Spring 2025 Pre -lab 15

# Experiment 15

# Carbohydrates  and Proteins Chem 100, Spring 2025 Pre -lab 15

## Purpose

To become acquainted with some of the properties of and standard chemical tests for

carbohydrates and proteins.

## Materials

Reagents and equipment you will need this week:

Parchment paper or foil

Spray oil (nonstick spray)

Large cookie sheet

saucepan

cup sugar

4 tablespoons light corn syrup or honey

1 teaspoons baking soda

Stove or microwave

Pot or microwave -safe container to boil water

Timer

1 scissors to cut the egg whites

1 mixer/hand blender or wire whisk to whip egg whites

1 bowl, 1 Large bowl

Forks, Spoons, Knife (for cutting fruit)

6 small containers of about the same size. 2 eggs (split egg white into six parts);

additional can be used (You can cut the egg white into pieces with your scissors.)

1/3 cup of rubbing alcohol (75 ml)

1/3 cup of white vinegar (75 ml)

Gelatin flavored or unflavored

18 dixie cups (or a 12 spot muffin tray.)

Plastic wrap

Universal indicator strip or red cabbage strip indicator strip

Table salt

Ammonia (base) or Window cleaner (base) or Baking Soda (base) or Toilet

cleanser (base)

Vinegar (acid) Lemon Juice (acid)

Meat Tenderizer

Fruits and/or fruit juice:

Pineapple fresh, frozen and canned, please try all 3.

Papaya or Fig or Kiwi ( Choose one ).

Apples or Grapes or Strawberries or Orange or other fruit (Choose 2) Chem 100, Spring 2025 Pre -lab 15

## Quiz Preparation

Vocabulary (terms you may see on the quiz)

monosaccharide

disaccharide

polysaccharide

carbohydrates

triose

tetrose

pentose

hexose

ketose

aldose

reducing sugar

glycosidic linkage

## Introduction/Background

Carbohydrates:

Carbohydrates serve as the primary chemical energy source for almost every form of

life, and are therefore fundamental compounds in living organisms. Carbohydrates may

be classified as monosaccharides, disaccharides, or polysaccharides.

Classes of Carbohydrates

> Monosaccharide

A monosaccharide (or simple sugar) is a polyhydroxy (many hydroxyls) carbon

compound that contains a carbonyl group. A monosaccharide may be classified as a

triose, tetrose, pentose, or hexose, depending on the number of carbon atoms present.

Each carbon, except for the carbonyl carbon, has a hydroxyl attached to it. It is these

that give the physiological sensation of sweetness. Also, a carbohydrate may be

classified as an aldose (monosaccharide with an aldehyde functional group) or a ketose

(monosaccharid e with a ketone group) depending on the location of the carbonyl

carbon. Three monosaccharides with which you will become familiar are glucose,

galactose, and fructose. All three of these are hexoses which have the molecular

formula C 6H12 O6 and Glucose are aldoses, while fructose is a ketose.

> Disaccharides

A disaccharide is a compound that is composed of two monosaccharides held together

by a glycosidic linkage, which is actually an ether linkage. In the presence of acid or the

appropriate enzyme, the glycosidic linkage may be hydrolyzed, producing the two

monosaccharides. Three important disaccharides are maltose, lactose, and sucrose.

Maltose is composed of two glucose; lactose is composed of glucose and galactose,

and sucrose is composed of glucose and fructose. Sucrose is common table sugar. Chem 100, Spring 2025 Pre -lab 15

> Polysaccharides

A polysaccharide is a polymer (a very long chain) of monosaccharides, each one held to

the next one by a glycosidic linkage. Like disaccharides, polysaccharides may be

hydrolyzed in the presence of acid or enzymes to produce monosaccharides. Three

importan t polysaccharides are starch, glycogen, and cellulose. All three of these are

polymers of the monosaccharide glucose. Starch is composed of two polysaccharides

a branched polysaccharide called amylopectin, and a straight -chain polysaccharide

called amylose .

Reducing Sugars

A reducing sugar is a monosaccharide or disaccharide which can be oxidized by

Benedict's reagent. Any monosaccharide or disaccharide that has an available aldehyde

functional group, or an  -hydroxy (read as alpha -hydroxy) ketone functional group (a

functional group in which a hydroxyl is on the carbon attached to the carbonyl carbon of

a ketone), can be oxidized. It is called reducing sugar because it reduces (the opposite

of oxidizes) the r eagent that causes the oxidation. All monosaccharides and most

disaccharides are reducing sugars.

Watch the video below to help understand the theory behind Benedict's Test for Sugars.

In today's experiment, you will test several carbohydrates to determine which are

reducing sugars. A carbohydrate that gives a positive test --the blue solution tums green

and then a brick -red precipitate forms --is a reducing sugar.

Iodine Test for Starch

When an aqueous solution of iodine, I 2, is added to a starch solution, the iodine solution

changes color from brownish orange to dark blue -black. This does not happen with any

other carbohydrate. This is because one of the two polysaccharides of which starch is

composed, amylose, is a long unbranched chain of glucose molecules which is twisted

into a coil called a helix. When th e iodine solution Is added to the starch solution, the

iodine gets trapped in the amylose coil and the blue -black color is observed. Chem 100, Spring 2025 Pre -lab 15

Hydrolysis

In hydrolysis reactions of carbohydrates, the disaccharides and polysaccharides are

broken down into monosaccharides. This changes the chemical reactivity. We will use

sucrose and starch to model the effect of hydrolysis on the chemical properties of

dis accharides and polysaccharides. The first step will be to hydrolyze the sample, acid

hydrolysis will be used. Then the samples will be tested with the Benedict's and starch

iodine tests. Afterward, you will answer some associated questions.

Proteins:

A polypeptide is a polymer (a very long chain) of amino acids. There are about twenty

different amino acids. One amino acid in the polypeptide chain is held to the next one by

a peptide bond, which is the name given to an amide linkage between two amino acids.

The polypeptide chain is coiled and folded up in a manner that is dependent on the

locations of the various amino acids in the polypeptide chain. A protein is composed of

one or more polypeptide chains.

Amino acids

The twenty common amino acids all have the same basic structure:

They differ only in their R groups (side chains).

Color Tests for Proteins

A color test for protein tests for a particular functional group or structural component

present in a protein or in a particular amino acid found in proteins.

> Biuret Test

When the biuret test is performed on a compound that contains two or more peptide

bonds, a violet color appears. Since all proteins contain peptide bonds, all proteins give

positive Biuret tests.

> Xanthroproteic Test

Xanthroproteic test is specific for the side chains of three amino acids: tyrosine,

tryptophan, and phenylalanine. Each of these has a side chain that contains a benzene

ring with either an amino group or a hydroxyl group attached to it. When the

Xanthropr oteic is positive, a yellow color appears. Chem 100, Spring 2025 Pre -lab 15

Protein Denaturation

The polypeptide chain(s) of a protein needs to be folded in order for the protein to be

able to function. To denature a protein means to cause the polypeptide chain(s) to

unfold. A denatured protein cannot carry out its biological function, and often a

den atured protein becomes less soluble (it coagulates). Certain reagents or conditions

will denature proteins. The addition of acid, base, ethanol, tannic acid and heavy metal

ions such as mercury, silver, and lead will denature a protein. Heating a protein w ill also

cause it to denature. These cause denaturation by interfering with the forces that are

keeping the polypeptide chain(s) folded.

> The Effects of Denaturing on Egg Albumin:

Here are some brief explanations for what you see:

Control. Egg whites start out clear. They are almost 90% water, but the other 10% is

packed with proteins. Egg whites contain more than 50% of the proteins found in the

egg. The main protein in egg white is called albumin. The small, folded proteins in the

egg whi te dont take up much space, and the gel -like egg white looks clear. The control

egg showed us that, when left at room temperature, the egg whites stay clear, meaning

the proteins maintain their original shape. These proteins were not denatured.

Physical . When you beat raw egg whites to make a souffl or a meringue, you incorporate

air bubbles into the water -protein solution. Adding air bubbles to egg whites unfolds

those egg proteins just as certainly as heating them. To understand why introducing air

bu bbles makes egg proteins uncurl, you need to know a basic fact about the amino

acids that make up proteins. Some amino acids are attracted to water; theyre

hydrophilic, or water -loving. Other amino acids are repelled by water; theyre

hydrophobic, or wate r-fearing. Egg -white proteins contain both hydrophilic and

hydrophobic amino acids. When the protein is curled up, the hydrophobic amino acids

are packed in the center away from the water and the hydrophilic ones are on the

outside closer to the water. Whe n an egg protein is up against an air bubble, part of that

protein is exposed to air and part is still in the water. The protein uncurls so that its

water -loving parts can be immersed in the water  and its water -fearing parts can stick

into the air. Once t he proteins uncurl, they bond with each other just as they did when

heated creating a network that can hold the air bubbles in place.

Room temperature water. Sometimes in this experiment, room temperature water has a

small denaturing effect on some of the egg white. It acts in the same way, by breaking

bonds, but its effect isnt nearly as strong as alcohol or hot water. Chem 100, Spring 2025 Pre -lab 15

Alcohol. Alcohol also denatures proteins. It does this the same way as heat, by breaking

the bonds that hold parts of the protein in a folded shape. Sometimes the alcohol

molecules bond directly to some of the parts of the protein, disrupting the normal way

the pr otein would bond to itself. (So alcohol is called a bond disruptor.) The proteins

unfolded, taking up more space and hardening in place next to one another. This

process takes much longer with alcohol than it does with heat. The longer time for

denaturat ion with alcohol is simply because it spreads more slowly than heat. The

alcohol has to diffuse (or move through the fluid) into the egg in order to affect the

proteins it touched.

Changing pH (vinegar) . Changes in pH change the attractions between the groups in the

side chains of the protein. The interactions between the side chains of the amino acids

determine the shape of a protein. When these interactions are disrupted, the protein

unfolds.

Cooking (hot water). Whenever eggs are cooked with heat, the egg whites turn from clear to

white, and the gel becomes more rubbery. As heat -denatured the proteins in the egg

white, it broke apart some of the bonds (mostly hydrogen bonds) that were holding the

proteins in thei r original shape. The proteins unfolded, taking up more space (turning the

gel white) and hardening them in place next to one another.

> Denaturing and Enzyme Activity:

The final portion of the experiment will look at how denaturing affects the activity of an

enzyme, bromelain. Active bromelain will destroy other proteins, like the collagen found

in gelatin. Enzymes designed to hydrolyze proteins are called proteases. Whe n the

proteins in the pineapple are denatured, the enzyme is deactivated and has no effect on

the gelatin. Some other fruits have a similar effect on gelatin because they also have

enzymes that denature proteins. You will determine which fruits may have pr oteases in

them and which do not.

Video of Egg Albumin:  This video  gives an example and explanation of this portion of

this experiment.

Video of Pineapple and Gelatin:  This video  gives an example and explanation of this

portion of this experiment. Chem 100, Spring 2025 Pre -lab 15

## Station A:

Benedict's Test for Sugars

1. Each of the sugars has been dissolved in water to make a 1% sugar solution.

2. Add a sugar solution to the labelled test tube as indicated.

a. To tube A add ~ 1mL of glucose

b. To tube B add ~ 1mL of fructose

c. To tube C add ~ 1mL of sucrose

d. To tube D add ~ 1mL of lactose

e. To tube E add ~ 1mL of starch

f. To tube F add ~ 1mL of hydrolyzed sucrose

g. To tube G add ~ 1mL of hydrolyzed starch

3. Add ~ 1mL of Benedicts reagent to each test tube.

4. Place the test tubes in a hot water bath to speed up the reactions.

5. Allow the reaction to heat for about 5 minutes.

6. Observe the test reactions

7. Record the results in the table in your lab report.

8. Complete the table by indicating whether each compound is a reducing sugar or

nonreducing sugar.

9. Take a photo of your test results and place it in your lab report.

Results of Benedicts test

Positive Test: If the color changes to muddy green/brown, the Benedicts

test is positive, which means reducing sugar exists in the solution.

Negative Test: If you find that no color change is visible, the result is

negative and indicates that non -reducing sugar is present in the solution.

Tube  Sugar Name  Benedict Color  Reducing

1 glucose  *****  Reducing or nonreducing

2 fructose  *****  Reducing or nonreducing

3 sucrose  *****  Reducing or nonreducing

4 lactose  *****  Reducing or nonreducing

5 starch  *****  Reducing or nonreducing

6 hydrolyzed sucrose  *****  Reducing or nonreducing

7 hydrolyzed starch  *****  Reducing or nonreducing Chem 100, Spring 2025 Pre -lab 15

Benedict's Test for Sugars Photo

Insert photo here. Shrink photo to fit within this box.

Benedict's Test for Sugars Questions

1. How do you know if you have a positive result in the Benedict's test? Write Your

Answer Here.

2. What is the name of the functional group that must be available to give you a

positive Benedicts test? (not the class of compound) Write Your Answer Here.

3. Do all monosaccharides give positive Benedict's tests? Choose one: Yes or No.

4. Do all disaccharides give positive Benedict's tests? Choose one: Yes or No .

5. Does hydrolyzed sucrose give positive Benedict's tests? Choose one: Yes or No .

6. What are the two monosaccharides produced when sucrose is hydrolyzed? Write

Your Answer Here.

7. Did sucrose which was not hydrolyzed give a positive or negative Benedict's

test? Choose one: positive or negative

8. Did hydrolyzed sucrose give a positive or negative Benedict's test? Choose one:

Yes or No

9. Explain the difference in the results of these two Benedict's tests. Choose one:

Yes or No

10. Does hydrolyzed starch give positive Benedict's tests? Choose one: Yes or No .

11. Do all polysaccharides give positive Benedict's tests? Choose one: Yes or No .Chem 100, Spring 2025 Pre -lab 15

## Station B:

Iodine Test for Starch

These are the directions for the starch iodine experiment: Each of the sugars has been

dissolved in water to make a 1% sugar solution.

1. Add a sugar solution to the labelled spot plate as indicated.

a. To spot A add ~ 1mL of glucose

b. To spot B add ~ 1mL of fructose

c. To spot C add ~ 1mL of sucrose

d. To spot D add ~ 1mL of lactose

e. To spot E add ~ 1mL of starch

f. To spot F add ~ 1mL of hydrolyzed sucrose

g. To spot G add ~ 1mL of hydrolyzed starch

2. Add about 2 drops of iodine solution to each test tube.

3. Record your observations in the table below.

4. Record the results in the table in your lab report.

5. Indicate whether starch was present in each sample in the table in your lab

report.

6. Take a photo of your test results and place it in your lab report.

Results of Iodine test:

Positive Test: If the color changes to blue or black, the iodine test is

positive, which means the starch amylose coil is present in the solution.

Negative Test: If you find that no color change occurs (or the solution

remains a pale yellow, the result is negative and indicates that no starch is

present in the solution.

Sample Table for Iodine Test

Tube  Sugar Name  Iodine Color  Starch present

1 glucose  *****  Yes or No

2 fructose  *****  Yes or No

3 sucrose  *****  Yes or No

4 lactose  *****  Yes or No

5 starch  *****  Yes or No

6 hydrolyzed sucrose  *****  Yes or No

7 hydrolyzed starch  *****  Yes or No Chem 100, Spring 2025 Pre -lab 15

Image: Iodine Test:

Insert photo here. Shrink photo to fit within this box.

Iodine Test for Starch Questions

1. What color is produced when the iodine test is positive? Write your Answer

Here.

2. What form of starch causes the reaction with iodine? Write your Answer Here.

3. What structural component makes this form of starch the only polypetide to react

with iodine? Write your Answer Here.

4. Do all monosaccharides give positive iodine tests? Choose one : Yes or No.

5. Do all disaccharides give positive iodine tests? Choose one : Yes or No .

6. Does hydrolyzed sucrose give positive iodine tests? Choose one : Yes or No .

7. Do all polysaccharides give positive iodine tests? Choose one : Yes or No .

8. Does hydrolyzed starch give positive iodine tests? Choose one : Yes or No .

9. Does a hydrolyzed starch solution give a positive test? Choose one : Yes or No

10. What monosaccharide is produced when starch is completely hydrolyzed? Write

Your Answer Here.

11. What disaccharide is produced when starch is partially hydrolyzed? Write Your

Answer Here.

12. What color should the disaccharide that is produced when starch is partially

hydrolyzed give in the Benedict test? Write Your Answer Here.

13. What color should the disaccharide that is produced when starch is partially

hydrolyzed give in the iodine test? Write Your Answer Here. Chem 100, Spring 2025 Pre -lab 15

Table: Sugar Results Summary

Tube  Sugar

Name

Structure  Mono, Di or

Poly

Aldose,

Ketose, or

neither

Benedict

Color

Reducing or

nonreducing

Iodine

Color

Starch

present?

1 glucose  Mono

Di or

Poly

A, K or

neither