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Lab #3 Organic Compounds (Taken from General Biology: Investigations into Life’s Phenomena) Name _______________________ Date ___________ Period ___________ Group Data, Individual answers Objectives 1. Distinguish among monosaccharides, disaccharides, and polysaccharides, giving examples of each. 2. Describe the functions and chemical structure of proteins. 3. Describe the chemical structure of nucleotides and nucleic acids and explain the importance of these compounds in living organisms. 4. Name the specific lab test used to identify (a) sugar, (b) starch, (c) fat, (d) protein, and (e) nucleic acid. Introduction The various molecules found in all life forms are called organic compounds because they are produced by organisms. Organic molecules are carbon based and range from small molecules to ones that are enormous in size. These larger molecules are not as complex as they first seem because they are composed of simpler, smaller molecules called monomers which link together into larger, longer chains called polymers. There are four major classes of biologically important organic compounds. Within a living cell, each of these four major classes has specific important functions. The table on the following page describes each of these classes, and lists examples. Study this table, and Section 2.3 of your text carefully. Most of the tests you will be performing today will lead to a color change as an indication of a chemical reaction. The chemical reaction will occur if the substance you are testing is present. Table 1 Four classes of biologically important organic compounds CARBOHYDRATES Carbohydrates are energy sources and provide structural support as in the cell wall of plants. _____________, ______________ and ________________ are the elements found in carbohydrates. Carbohydrates may be classified as monosaccharides, disaccharides, or polysaccharides. 1. Monosaccharides are _______________. 2. Disaccharides are _____ monosaccharides bonded together. 3. Polysaccharides are three or more monosaccharides bonded together. ______, ___________, and __________ are important polysaccharides. a. Starch: LIPIDS Lipids are a varied group of molecules that are mostly _______________ in water. Like carbohydrates, carbon, hydrogen, and oxygen are the principal elements of lipids, although the oxygen content is _____________. Lipids are essential components of ________________, a good means of ______________, and some lipids are _____________________. Neutral fats Phospholipids Steroids b. Glycogen: c. Cellulose: Waxes PROTEINS NUCLEIC ACID Proteins have several important roles in the living organism. The elements in proteins are carbon, hydrogen, oxygen, nitrogen, and sometimes sulfur. A protein is made of these elements arranged into subunits called ________________. Long chains of amino acids are linked together with ________ _____________ which make the chain coil and fold into complex shapes that determine the function of the resulting protein molecule. Proteins form many structural features such as _____________, ______________ and ______________. Many proteins called _______________ help control the chemical reactions that keep the cells alive. Nucleic acids are extremely large and complex molecules that have variety of important biological functions. The elements of nucleic acids are carbon, hydrogen, oxygen, nitrogen, and phosphorus. 1. One general type of nucleic acid, called ______________________________ is coded genetic information. 2. A second type, called ______________ is what turns the genetic code into a protein. In other words, it is involved in the expression of the genetic code. 3. Nucleotides are the units, or the monomers, of nucleic acid polymers 4. _____ is a nucleotide that stays in monomer form. It is responsible for supplying __________ for the chemical reactions of the cell Pre-Lab Questions These questions will help you prepare for the lab. The pre-lab questions are due before your group can start the lab. 1. Define organic compound ____________________________________________________________ ____________________________________________________________________________________ 2. Define monomer and polymer ______________________________________________________ ____________________________________________________________________________________ 3. Define reagent __________________________________________________________________ ____________________________________________________________________________________ 5. Define chemical reaction ____________________________________________________________ ____________________________________________________________________________________ 6. Define polar and non-polar ____________________________________________________________ ____________________________________________________________________________________ 7. Define soluble __________________________________________________________________ ____________________________________________________________________________________ 8. What elements make up carbohydrates? ________________________________________________ ____________________________________________________________________________________ 9. What are three categories of carbohydrates and what are their functions? ________________________ ____________________________________________________________________________________ ____________________________________________________________________________________ 10. Why do plants have starch? Why do they have cellulose? ______________________________ ____________________________________________________________________________________ 11. What does a positive test for simple sugar look like? Protein? Starch? Lipid? __________________ ____________________________________________________________________________________ ____________________________________________________________________________________ 12. What are two types of nucleic acids? What are their functions? ______________________________ ____________________________________________________________________________________ Part 1: Testing for simple sugar = Benedict’s Test Background Some simple sugars (monosaccharides) have an aldehyde group ( H–C=O) in their molecular structure. Aldehyde reacts with Benedict’s solution when heated. The reaction produces a color change in the Benedict’s reagent from an original blue color to an orange/red. A color change indicates a positive test for sugar. You will test three different types of carbohydrates to see which one is a simple sugar. You will also test water, as a control. You will need to see if/how Benedict’s solution changes when heated, so that you can compare the normal change to a change that is caused by the reaction with a simple sugar. Figure 1. Structural formula of ribose. Try to identify the aldehyde functional group. Materials 4 test tubes test tube rack test tube holder boiling water bath ruler starch solution Benedict’s Reagent glucose solution sucrose solution distilled water Procedure 1. Arrange your tables into stations, and obtain materials located in your station container. Note the location of the boiling water bath(s) that your station will be using. 2. Use a ruler and wax pencil to make 2 marks on each test tube. Make one mark at 1cm from the bottom, and another at 3cm from the bottom. Label the tubes 1-4. 3. Fill your test tubes according to Table 2.1. Table 1 Results of Benedict’s Test Tube Tube Contents Tube Contents # Add up to 1cm mark Add up to 3cm mark 1 water Benedict’s reagent 2 Glucose solution Benedict’s reagent 3 Sucrose solution Benedict’s reagent 4 Starch solution Benedict’s reagent Observations 4. Place all four test tubes in a boiling water bath and heat for three to five minutes. 5. Remove the test tubes from the hot water bath with the test tube holder, and place them in the test tube rack. Record your observations of the final results in Table 2.1 6. Thoroughly rinse and dry your test tubes in preparation for Lab 2.2 Results/Analysis 1. Which carbohydrate solution contains reducing sugars? _____________________________________ 2. What is the functional group in this particular carbohydrate molecule that produces a positive reaction with Benedict’s reagent? ____________________________________________________________ ____________________________________________________________________________________ 3. If onion juice mixed with Benedict’s reagent gives a positive test, what sugar would you conclude is present in onions? __________________________________________________________________ ____________________________________________________________________________________ 4. If you were given an unknown solution, what test would you do to determine the presence of simple sugars?______________________________________________________________________________ Part 2 Testing for Starch = Iodine Test Background Iodine reacts with starch resulting in a dark, blue/black color that is a positive test for starch. The reaction occurs because the starch binds with the iodine molecules and thus produces a color change. Starch is stored in the seeds, roots, and tubers of plants as an energy reserve. The starch is converted to simple sugars and is used by the plant during times of growth or when low levels of photosynthesis may not meet its energy requirements. Materials Ruler Wax Pencil 4 test tubes test tube rack potato solution sucrose solution water potato slice razor blade microscope Procedure 1. Use the ruler and wax pencil to mark 1cm from the bottom of each test tube. Label the test tubes 1-4. 2. Fill your test tubes according to Table 2.2 Table 2 Results of Iodine Test for Starch Tube Tube Contents Tube Contents # Add up to 1cm mark Add 3 drops, swirl to mix 1 Water Iodine solution 2 Carrot solution Iodine solution 3 Sucrose solution Iodine solution 4 Potato solution Iodine solution Observations 4. Record your observations in Table 2.2 5. Thoroughly rinse and dry your test tubes. Your instructor will demonstrate the following procedure. You will make observations. 6. Use a razor blade to cut a small, extremely thin slice of fresh potato. The potato slice should be thin enough to be nearly transparent. 7. Place the potato slice on a clean microscope slide. Add a drop of water to the potato, and cover with a cover slip. 8. Observe the potato slice under low power. Then switch to medium power. In detail, draw your field of view. 9. Carefully remove the cover slip, and add 2 drops of iodine solution. Replace the cover slip, and wait 2 minutes. Observe the potato under low power. Then switch to medium power. In detail, draw your field of view. Results/Analysis 1. Which solutions contained starch? _____________________________________________________ 2. Starch and cellulose are both polymers of glucose molecules. Make a prediction for how they would react in a Benedict’s test. ___________________________________________________________ ___________________________________________________________________________________ 3. Starch gives a positive iodine test, but cellulose doesn’t, even though they are both polymers of glucose. What is a plausible explanation for this difference? ____________________________________ ____________________________________________________________________________________ 4. What molecule is located in the dark bodies of the potato slice after the iodine was added? _________ 5. If you wanted to test if a mixture had starch in it, what test would you run? _____________________ Part 3: Testing for Proteins = Biuret Test Background Protein molecules consist of multiple amino acid molecules linked in long chains. The amino acids are linked by bonds called peptide bonds. Peptide bonds react with a copper-containing molecule in the Buiret reagent. This reaction causes a purple/violet color. Materials 4 test tubes test tube rack ruler Water Honey solution wax pencil Egg white solution Potato solution Biuret reagent Procedure 1. Mark four test tubes at 2cm and 4 cm from the bottom. Label the tubes 1-4. 2. Fill your test tubes according to Table 2.3 and record observations Table 3 Results of Biuret Test for Protein Tube Tube Contents Tube Contents # Add up to 2cm mark Add up to 4cm mark 1 Water Biuret reagent 2 Honey solution Biuret reagent 3 Egg white solution Biuret reagent 4 Potato solution Biuret reagent Observations 3. Thoroughly rinse and dry your test tubes. Results/Analysis 1. Which solution contained protein? _____________________________________________________ 2. What type of chemical bond causes the biuret reagent to react and give a violet color? ___________ ___________________________________________________________________________________ 3. Would you expect a solution containing an enzyme to yield a positive biuret test? Why or why not? ___________________________________________________________________________________ ___________________________________________________________________________________ Part 4: Testing for Nucleic Acids = Dische Diphenylamine This portion of the lab will be omitted. The reagents are highly caustic. Part 5 Testing for Lipids = Sudan III Test Background Lipids are non-polar compounds composed of a glycerol molecule bonded to three fatty acid molecules. Because lipids are non-polar, they do not dissolve in water or other polar liquids. Lipids are soluble in non-polar substances such as ether. Therefore, solubility in Sudan III dye can be used as a positive test for lipids. Non-lipid substances will not dissolve in Sudan III dye and will form two distinct layers in a test tube. Materials 4 test tubes test tube rack wax pencil ruler water honey solution egg white solution vegetable oil Sudan III dye Procedure 1. Mark 4 test tubes at 2cm from the bottom. Label the tubes 1-4. 2. Fill your test tubes according to Table 2.5 and record your observations Table 2.5 Results of Sudan III Test for Lipids Tube Tube Contents Tube Contents # Add up to 2cm mark Add 10 drops and swirl to mix 1 Water Sudan III dye 2 Honey solution Sudan III dye 3 Egg white solution Sudan III dye 4 Vegetable oil Sudan III dye Observations 3. Thoroughly rinse and dry your test tubes. Please wipe off any wax pencil residue. Results/Analysis 1. Which compound(s) contained lipids? ________________________________________________ 2. Which tube served as the control? ______________________________________________________ 3. If you were given an unknown substance, which test would you run to see if the unknown substance contained lipids? __________________________________________________________________ Part 6 Testing an UNKNOWN SUBSTANCE Your instructor will provide you with a substance of unknown composition. 1. What test will you run to find out if the unknown has simple sugars? 2. What test will you run to find out if the unknown has proteins? 3. What test will you run to find out if the unknown has starch? 4. What test will you run to find out if the unknown has lipids? Results/Analysis Which organic molecule(s) did your unknown substance contain? ______________________________ ____________________________________________________________________________________ Post-lab questions 1. Using the pre-lab information and the information received from this investigation distinguish between monosaccharides, disaccharides, and polysaccharides. Give an example of each type. ____________________________________________________________________________________ ____________________________________________________________________________________ ____________________________________________________________________________________ ____________________________________________________________________________________ 2. Most of the most of the tests performed in this investigation lead to a color change in the presence of the compound being tested for. What caused these color changes? ______________________________ ____________________________________________________________________________________ 3. A positive result for lipids in the Sudan III test was not a color change. What was considered a positive result in this test? What does this tell us about the nature of lipids? _______________________ ____________________________________________________________________________________ ____________________________________________________________________________________ ____________________________________________________________________________________ 4. Contrary to popular belief the tests used in this investigation were not created to torture students. Name a real life situation where it might be important to test for a specific type of organic compound. ____________________________________________________________________________________ ____________________________________________________________________________________ ____________________________________________________________________________________ ____________________________________________________________________________________ 5. In your opinion is it important in the context of biology to understand organic compounds? Can studying organic compounds lead to a greater understanding of life? Explain. _____________________ ____________________________________________________________________________________ ____________________________________________________________________________________ ____________________________________________________________________________________ ____________________________________________________________________________________ ____________________________________________________________________________________