You Are What You Eat

Transcription

1 An Investigation of Macromolecules Student Materials Introduction....2 Pre-Lab Questions.5 Lab Protocol..6 Post-Lab Questions and Analysis 9 Last updated: September 26 th,

2 Introduction When deciding what to eat for lunch, do you ever base your choices off of the amount of protein, carbohydrates, or fats (lipids) are in your foods? Or maybe you think back to the food pyramid: grains on the bottom, meat, dairy, fruits, and vegetables in between, with fats and sweets at the top? od-pyramid-1024x768.jpg?w=640 Now let s think about the food pyramid in terms of protein, carbohydrates, and lipids. You probably already know that grains like bread contain carbohydrates, but did you know that bread is also is a source of protein? Working up the pyramid, we all know that meat such as chicken or fish contain protein, but did you also know that they also contain fat? Even desserts such as chocolate aren t as simple as just containing fats, they are also made of carbohydrates and even proteins. On top of all of that, almost every time you re eating, you are eating DNA and RNA! You may not have realized it, but the protein, carbohydrates, and lipids foods contain comes down the food s biochemistry. Much of our food comes directly from plants and animals and we all know that plants and animals are composed of cells. All cells are composed of large, complex molecules called macromolecules; macromolecules are just big molecules composed of multiple subunits. The four main macromolecules are proteins, carbohydrates, lipids, and nucleic acids. Today s lab will be all about macromolecules and you already know so much about them! Now let s think 1000 times deeper into each of the four main macromolecules and their place in the foods we eat. Say you have a chicken breast and strawberries for lunch. Both foods cells are composed of the macromolecules that we have talked about: protein, carbohydrates, lipids, and nucleic acids. So why are they on different places of the pyramid? Let s think: Chicken breast and most meat is high in protein. The chicken breast is mostly muscle tissue. Muscle tissue is composed of muscle cells and is therefore packed with muscle proteins. Strawberries are not only sweet due to the sugars glucose and fructose, but are also a great source of fiber. The fiber in strawberries is primarily cellulose. What type of macromolecule is cellulose? 2

3 We know that macromolecules are made of atoms covalently bonded together. In biology, the most common atoms are carbon, hydrogen, nitrogen, oxygen, and phosphorus. Which of the macromolecules (protein, carbohydrate, lipid, and DNA) in your lunch: contain carbon? contain nitrogen? contain phosphorus? Most macromolecules are made of repeating subunits called monomers. Since macromolecules are made of several monomers they can be called polymers. When two monomers are joined together the chemical reaction is called dehydration synthesis. The reaction involves the removal of an OH from one monomer and an H + from another, which come together to make a water molecule. For every two monomers that are linked together, one water molecule is released. In addition to forming new polymers, cells also need to break down polymers; this reaction is called hydrolysis. Hydrolysis requires the addition of a water molecule and breaks the bond between the two monomers. Dairy products contain the carbohydrate lactose, which is made up of galactose and glucose (C 6H 12O 6). The overall formula for lactose is C 12H 22O 11. When lactose forms, a molecule of water (H 2O) is released. What is the chemical formula for galactose? Hint! Think of the overall reaction: glucose + galactose lactose + water. Fill in the formulas you know and balance the equation. What do you notice about the chemical formulas for glucose and galactose? Testing for macromolecules In this lab, you will use simple methods to test for the presence of proteins, carbohydrates, lipids, and nucleic acids. Proteins Proteins are used in cells in various ways; they are used as enzymes, as structural components, and for cell signaling. Some proteins are composed of a single long polymer of amino acids called a polypeptide. Other proteins are composed of two or more polypeptides. The amino acids in polypeptides are joined together in chains and these amino acid chains are extremely diverse because they are composed of any combination and number of the 20 different amino acids. The order of amino acids in a polypeptide dictates the structure and function of the protein. In this lab, you will use a chemical called Biuret to test foods for the presence of proteins. Biuret is a blue liquid that will turn purple when mixed with proteins. What do enzymes do? 3

4 Carbohydrates Carbohydrates are primarily used as a source of energy for the cell but can also be used as structural components to give the cell shape. Carbohydrates, often called sugars, are macromolecules that come in different forms and sizes but all have the same ratio of carbon to hydrogen to oxygen; 1 carbon to 2 hydrogens to 1 oxygen, or CH 2O. Therefore, a sugar with 5 carbons would have the formula C 5H 10O 5. Monosaccharides are known as simple sugars, they only contain only 3-7 carbons. Disaccharides are molecules composed of two sugars linked together. Polysaccharides, known as complex sugars, are made up of long chains of monosaccharides. The varying number of sugars in different carbohydrates makes it possible to test foods for different carbohydrates. In this lab, you will use Benedict s and iodine to test foods for simple and complex sugars. Benedict s will change from blue to a red or rusty orange when monosaccharides are present. Lugol s s (iodine) will change from dark red to blue-black when it is mixed with the polysaccharide starch. Lipids Lipids are used for energy storage and as important components of membranes. Lipids are a large and diverse group of molecules that are primarily composed of long chains of hydrogen and carbon. They are nonpolar (hydrophobic) molecules, so they do not mix with water. Lipids are not composed of repeating subunits, so they are not actually polymers. Modified lipids called phospholipids are the major component of the plasma membrane and are critical to cells. In this lab, you will use Sudan IV to test for the presence of lipids. Like lipids, Sudan IV is hydrophobic, so if lipids are present, they will absorb the Sudan IV and form a noticeable bright-red layer. Given what you know about lipids, explain why some types of salad dressings separate. Nucleic acids Nucleic acids are primarily used for information storage and transmission; their information is needed for cells to make proteins. The nucleic acid you re most familiar with is DNA. In most eukaryotic cells, DNA is found as a long double-stranded molecule called a chromosome. Any region of a chromosome that is used to form a protein is called a gene. Nucleic acids are polymers of nucleotides. Each nucleotide is made up of sugar, a phosphate group, and a base. The sugar found in DNA nucleotides is deoxyribose, while the sugar in RNA nucleotides is ribose. The bases adenine, cytosine, guanine, and thymine (A, C, G, T) are found in DNA, while RNA contains adenine, cytosine, guanine, and uracil (A, C, G, U). Together, DNA and RNA contain the blueprints and instructions for all of life s processes. In this lab, you will use a DNA stain such as SYBR Safe or Gel Green to test for the presence of nucleic acids (or DNA). DNA stains contain a molecule that binds to DNA and causes it to glow green when viewed under blue light or UV light. 4

5 Pre-Lab Questions Directions: After reading through the introduction and protocol for the You Are What You Eat lab, answer the questions below. 1. What are the four primary types of macromolecules? 2. You read somewhere that having a protein-rich breakfast can help combat fatigue and improve test performance. Which of the following would provide a protein-rich breakfast on the morning of a big exam: an apple and toast OR a ham omelet? Explain why your choice has more protein. 3. Look at the diagram below. Count the number of carbons, hydrogens, and oxygens in the circled monomer. What type of molecule is this monomer? What type of macromolecule is formed by linking many of these monomers? Is the reaction below a dehydration reaction or a hydrolysis reaction? Adapted from 5. Match the following indicators with the type of macromolecule they will be used to test: 1. Benedict s a. Carbohydrates monosaccharides 2. Biuret b. Carbohydrate polysaccharides 3. Lugol s c. Lipids 4. Sudan IV d. Nucleic acids 5. SYBR Safe e. Proteins 5

6 Lab Protocol For each type of macromolecule, you will test a known sample as a standard. For example, you will test glucose, a monosaccharide, with Benedict s to see what a positive reaction looks like. Materials: Check your workstations to make sure all supplies are present before beginning the lab. Student Workstation: 1 p200 micropipette and pipette tips 2 microcentrifuge tubes microcentrifuge tube rack 8 test tubes test tube rack test tube holder 250 ml beaker 10 graduated transfer pipettes permanent marker 2 ml Biuret 3 ml Benedict s 2 ml Lugol s 60 μl of SYBR Safe 2 ml of Sudan IV 25 μl of Lambda DNA 2 ml of vegetable oil 2 ml of BSA 3 ml of glucose 3 ml of starch dh 20 Common Workstation: microwave or hot plate UV or blue light 600 ml beaker Procedure: In this procedure, you will set up a test sample and control sample for one macromolecule at a time. Then you will set up the samples for the next macromolecule. Table 1 provides an overview of the testing samples. Reminder: Safety goggles should be worn throughout this lab. Protein 1. Use a permanent marker to label two test tubes: P+ and P-. 6

7 2. Using the water pipette put approximately 2 ml of distilled water into the tube labeled P-. 3. Using a clean transfer pipette, put approximately 2 ml of BSA into the P+ test tube. 4. Add 5 drops of Biuret reagent to the P+ and P- tubes. Hold the tubes over white paper and examine. Record the color in Table 1. Carbohydrates: glucose and starch 5. Use a permanent marker to label two test tubes: G+ and G-. 6. Use the permanent marker to label a transfer pipette water. You will use this pipette several times. 7. Using the water pipette, put approximately 2 ml of distilled water into the tube labeled G-. 8. Using a clean transfer pipette, put approximately 2 ml of glucose in the tube labeled G+. 9. Add 1 ml of Benedict s to the G+ and G- tubes. 10. Carefully place the test tubes in a hot water bath (at least 90 C). Wait 3-4 minutes. Using tongs or a test tube holder to protect your hand, remove the test tubes from the water bath and place them in a test tube rack. Record the color in Table 1. Caution! Be very careful when handling the test tubes, they will be hot. 11. Use a permanent marker to label two test tubes: S+ and S Using the water pipette put approximately 2 ml of distilled water into the tube labeled S Using a clean transfer pipette, put approximately 2 ml of starch into the S+ test tube. 14. Add 3 drops of Lugol s to the S+ and S- tubes. Record the color in Table 1. Lipids 15. Use a permanent marker to label two test tubes L+ and L Using the water transfer pipette, put approximately 2 ml of water into the L- test tube. 17. Using the water transfer pipette, put approximately 1 ml of water into the L+ test tube. 18. Using a clean transfer pipette, put approximately 1 ml of vegetable oil into the L+ test tube. 19. Add 3 drops of Sudan IV to both test tubes. Wait 8-10 minutes. Record the color in Table 1. Nucleic acids 20. Label two microcentrifuge tubes NA+ and NA Using a 200 μl micropipette, put approximately 25 μl of distilled water into the microcentrifuge tube NA Using a p200 micropipette and clean pipette tip, add 25 μl of Lambda DNA to the NA+ tube. 23. Using a clean pipette tip, add 25 μl of SYBR Safe to NA+ and NA- tubes. Important: Be sure to use a new tip for each test. 24. Gently mix the s and observe them under a UV light or blue LED light. Record the color in Table 1. Caution: Use the appropriate protective gear when using a UV light. 7

8 Nucleic acids Lipids Carbohydrates Protein Ps Table 1: Macromolecule Tests Tube Type Macromolecule Label Description Contents Test Reagent Color Change Observed P+ Protein BSA (protein) Biuret P- protein Biuret G+ Glucose Glucose Benedict s S- glucose and other monosaccharides Benedict s S+ Starch and other polysaccharides Starch Lugol s S- starch and other polysaccharides Lugol s L+ Lipids and vegetable oil Sudan IV L- lipids Sudan IV NA+ Nucleic acids (microcentrifuge tube) Lambda DNA SYBR Safe NA- nucleic acids (microcentrifuge tube) SYBR Safe Images: 8

9 Post-Lab Questions and Analysis Directions: After completing the You Are What You Eat lab, answer the questions below. 1. How did your predictions match up against your results? Did you get any unexpected results? 2. A food sample is tested for all four macromolecules with the following results: Benedict s no color change Iodine orange to black BSA blue to purple SYBR Safe no color change What macromolecules do these results suggest are in your food sample? Which macromolecules are likely not present? Explain your reasoning. 3. The nutritional information for fresh strawberries is below. Notice that there is no fat (Total Fat = 0g) in a serving of 8 medium berries. Your friend concludes that strawberries are fat free. Explain why that isn t true. 9