3ObjectivesDescribe the distinct features of three of four classes of organic moleculesDifferentiate between monomers and polymersIdentify reducing sugars, polysaccharides, lipids, amino acids and proteins using diagnostic reagentsConduct nutritional assessments from case studies
4OBJECTIVES Describe the functions of the major classes of biomolecules Explain how dehydration synthesis causes molecular bondsCompare the different bonds that help define the major classes of biomoleculesDescribe the reactive groups of the major biomolecules & explain how they relate to the basic chemical structure of each class of biomoleculesUse diagnostic reagents to identify biomolecules & explain the chemical basis for the testsDesign & present an experiment to test for the presence of biomolecules in food materialsExplain how the chemistry of biomolecules relates to FDA food labels, & how you can use this knowledge to make wise food choices
5Biomolecules: The Basics Organic molecules are necessary for biological processes.Organic molecules contain carbon.4 major types of organic molecules:CarbohydratesLipidsProteinsNucleic Acids (We’ll look at these in a future lab.)Similarities Between Biomolecules:All Contain CarbonMost Contain H, O, N, S and/or PALL Life on Earth is made up of Carbon
6Building Blocks of Life Each Macromolecule is comprised of building block molecules called monomers:Monosaccharides carbohydratesFatty Acids and Glycerol LipidsAmino Acids ProteinMacromolecule: A giant molecule formed by the joining of smaller molecules, usually by a condensation reaction.Monomer- The subunit that serves as the building block of a polymer.Polymer- A long molecule consisting of many similar or identical monomers linked together.
7Functional groupsThe components of organic molecules that are most commonly involved in chemical reactions are known as functional groupsEach functional group behaves consistently from one organic molecule to anotherProvide a way to differentiate between different biomolecules
8Hydroxyl Group Found in alcohols (ex. ethanol) and sugars Polar - increases the solubility of moleculesOH (not OH- hydroxide ion)Where are the Hydroxyl groups in the glucose and in the ethanol?ethanol
9Carbonyl Group Found in Carbohydrates: Two types Aldehyde Ketone terminalKetoneinternalOne or the other on every monosaccharide
10Carboxyl Group In fatty acids AND amino acids Acidic properties Double Bond between Carbon and OxygenSingle Bonds: Carbon to HydroxylWritten as COOH or C02H
11Amino Group In every amino acid Nitrogen atom attached to two hydrogen atoms
12Monomer – a single molecule that can combine to form polymers Polymer – long chains of monomers
13Dehydration Synthesis and Hydrolysis Monomers are connected by an enzymatic removal of waterOne monomer provides a hydroxyl group and the other provides a hydrogenResult is the release of a water moleculeHydrolysisCovalent Bonds connecting monomers into polymersA Hydrogen atom and hydroxyl group from a split water molecule attaches where the bond used to be1 H(+) Atom1 Hydroxyl Group (split from a water molecule)Attach where the bond used to be
14Examples of Dehydration and Hydrolysis Which one of these is also called a condensation reaction?
15Carbohydrates (Saccharides) Four major functions:Supply carbon for synthesis of other biomoleculesStructural components in cells and tissuesFuel (glucose)Store glucose (polysaccharides)Three types:MonosaccharidesDisaccharidesPolysaccharidesDefined by carbonyl groupAldehydeKetone
16Monosaccharides (Basic Unit of Carbohydrates) Simple sugarsClassified by the number of carbons they containTriosesPentosesHexosesMay Exist as Rings (Stable) or Linearly (Unstable)glucose
17Disaccharides Consist of 2 monosaccharides Linked via a condensation reaction (Dehydration Synthesis)Linked by a Glycosidic Bond
18Polysaccharides Chains of linked monomers Amylose Linear chain of several hundred glucose moleculesAmylopectinBranched chains of several thousand glucose moleculesStarch in plantsGlycogen in animals
19Reducing Sugars: Contain a free aldehyde or ketone group (carbonyl) Reducing sugars reduce something elseIncludes:All monosaccharidesDisaccharides where one aldehyde or ketone group is free
20Testing for Saccharides Benedicts TestTests for the presence or absence of reducing sugarA reducing sugar is a sugar with free aldehyde or ketone groupFree- not used in the bond between monosaccharides. All monosaccharides are reducing sugars.Positive test results red-orange-yellow color
21Testing for Saccharides Lugol’s TestIndicates the presence or absence of starchPositive test results black/purple colorAll our tests today will be qualitative not quantitative (specific numerical measurement)
22Lipids Nonpolar molecules Lipid molecules include: Composed of long chains of carbon and hydrogenLipid molecules include:Fats, Oils, Steroids, PhospholipidsMonomers (for triglycerides)glycerol and fatty acidsMajor functions include:Energy storageMaterial for membranes
23Lipids Saturated vs. Unsaturated Saturated lipids- Unsaturated lipids- All bonds between carbon & hydrogen are single bondsNo carbon-carbon double bondsStraight ChainsSolid at Room temperatureUnsaturated lipids-Some carbon-carbon bonds are double bondsKinked Chains where double bonds are presentKinks prevent molecules from packing tightly togetherPlants and Fish Fats (oils) are liquid at room temperature
24Saturated Fats Unsaturated Fats have fatty acids with no carbon-carbon double bondsThere is a hydrogen at every possible locationThe molecule has straight chainsSaturated fats are solid at room temperatureMost animal fats are saturatedContributes to cardiovascular diseasehave fatty acids with one or more carbon-carbon double bondsThe molecule has kinked chains where there is a carbon-carbon double bondsPlants and fish fats, known as oils, are liquid at room temperatureThe kinks provided by the double bonds prevent the molecules from packing tightly together
25LipidsTriglycerides are solid lipids (fats) that are stored in human tissue OR liquid lipids (oils) that are stored in plant tissue.Store more energy than any other moleculeTriglycerides are formed by attaching 3 fatty acid molecules to a glycerol molecule through a condensation reaction.Functional group for glycerol (an alcohol) = hydroxyl group (OH)Functional group for fatty acid (a carboxylic acid) = carboxyl group (COOH)
26Phospholipids Make up the cell membrane Consist of: a polar head (hydrophilic)a nonpolar tail (hydrophobic)
27Testing for Lipids Sudan IV A nonpolar dye Bonds to other nonpolar moleculesPositive test resultsCloudy emulsion of dispersed lipid droplets which are red color due to the Sudan IV
28ProteinsStructural support, storage, transport of other substances, and intercellular signalingALL Enzymes are ProteinsAll Proteins have a complex 3D ShapeMonomers = amino acids (there are 20 amino acids used by biological organisms)Amino acids contain:An amino (NH3 ) groupA carboxyl (COOH) group.
29Proteins and Amino Acids When amino acids are linked together via a condensation reaction, they form a protein.The chemical bond between each pair of amino acids is called a peptide bond.Carboxyl Group of one Amino Acid joins the amino group of another amino acidPolypeptides: polymers of proteinsProteins consist of one or morePolypeptides folded and coiledinto a specific formation
30Proteins and Amino Acids When long chains of amino acids form proteins, the chemical properties of the amino acids cause the protein to take on more complex shapes:Proteins are used for:Structural supportTransport of substancesCell signalingEnzymesAll enzymes are made of protein. Enzymes regulare metabolism by selectively accelerating chemical reactions
31Testing for Proteins Biuret Test React with peptide bonds Positive test results purpleNinhydrinTests for single amino acidsReacts with free amino groupsPositive test resultsPurple- straight chain amino acidYellow- ring chain amino acid (proline)
32Lab Safety – Biology 111 Lab 3 - Biomolecules Goggles and closed-toed shoes are required!Gloves need to be worn when handling all reagents. Refer to the lab manual for safety notices. Some materials such as Ninydrin and Ninhydrin-collidine are corrosive and toxic. Other chemicals are flammable.Closely monitor all hot plates. Immediately turn off when not in use and use hand protectors when handling any beakers placed on hot plates. Keep all solvents away from heat sources.Chromatography solvent is highly flammable with toxic, volatile fumes. Keep tightly closed when not in use and keep away from all heat sources.Dispose of any solutions containing the following reagents in the appropriately labeled containers on the side counter:Dispose of chromatography solvent, ethanol, and Sudan IV in the solvent waste container.Dispose of Lugol’s solution, Benedicts reagent, and Biuret’s reagent in the base waste container.Dispose of ninhydrin filter papers in the ninhydrin waste bucket.Keep waste containers closed when not disposing of waste, and do not fill past the indicated line on waste containers. Inform your TA of any full waste containers.Do not eat or drink in lab.Ask your TA if there is doubt about any procedure.
33Lab Activity: Testing for the presence of organic molecules Each table will work as a single group.Within each group, students will perform different tests.When all tests are complete, group members will teach each other how to conduct all of the tests.ProcedureHow does the test work?What is a positive / negative result?
34Lab Activity: Testing for the presence of organic molecules Organization for each table (8 students):Benedict’s Test for Reducing Sugars- 2 studentsLugol’s Test for Starches- 2 studentsSudan IV Test for Lipids- 1 studentBiuret Test for Proteins- 2 studentsNinhydrin Test for Amino Acids- 1 student
35Benedict’s TestTests for the presence of reducing sugars (reacts with the free aldehyde or ketone groups)You will fill in the data for Table 3-1.You will need 8 test tubes to test the 8 solutions in Table 3-1.Make sure you gently shake the solution bottles before you use them.10 drops = 1 mLUse parafilm to mix your solutions with the Benedict’s reagent in the test tubes.WARNING! BENEDICT’S REAGENT IS A CORROSIVE BASE. DO NOT COME IN CONTACT WITH THIS CHEMICAL AND DISPOSE IN BASE WASTE CONTAINER.
36Benedict’s Test A positive test result is any color change. Color changes range from:Blue > Green > Yellow > Reddish OrangeOrange and Red indicate the strongest reducing sugars.
37Lugol’s TestTests for the presence of starchesYou will fill in the data for table 3-1.Procedure begins on pg. 30You will need 8 test tubes to test the 8 solutions in Table 3-1 (these are the same solutions used in the Benedict’s test).Make sure you gently mix the solution bottles before you use them.10 drops = 1 mLUse parafilm to mix your solutions with the Lugol’s reagent.WARNING! LUGOL’S IS A TOXIN (ESPECIALLY FOR PEOPLE WITH IODINE ALLERGIES OR WILSON’S DISEASE). DO NOT COME IN CONTACT WITH THIS CHEMICAL AND DISPOSE IN BASE WASTE CONTAINER.
38Lugol’s TestA positive test result is a blue-black color, which indicates the presence of starch.Monosaccharides and Disaccharides will test negative for Lugol’s.
39Emulsification Test for lipids Use Ethanol + Sudan IVA non-polar dyeBonds to other non-polar moleculesPositive test results red colorPositiveNegative
40Sudan IV TestA positive test is a red stain left on the filter paper after rinsing.
41Thin Layer Chromatography Mixed lipids can be separatedYields a chromatogramMixed molecules interact with the solvent (mobile phase) and the silica gel (stationary phase) to separate as solvent moves upRelationship between distance moved by molecule to distance moved by solvent is specific for the matrix conditions & solventRelationship is given by RfOriginSolvent frontTriglycerideFatty acidsPhospholipidss
42Biuret TestTests for the presence of proteins by reacting with peptide bondsYou will fill in the data for TableProcedure begins on pg.You will need test tubes to test the solutionsMake sure you gently mix the solution bottles before you use them.10 drops = 1 mLUse parafilm to mix your solutions with the Biuret reagent.WARNING! BIURET IS A STRONG BASE. AVOID CONTACT AND DISPOSE OF IN BASE WASTE CONTAINER.
43Biuret Test A positive test is a violet color. Any other color change is a negative result.
44Ninhydrin TestTests for the presence of free amino acids by reacting with their amino groups.You will fill in the data for table.Make sure you gently shake the solution bottles before you use them.The test will be done on a piece of filter paper folded into eights.Use pencil to label each test area.Dry each solution on the filter paper before you test it with the ninhydrin solution.Dry the filter paper again after the ninhydrin is added.WARNING! NINHYDRIN IS POISONOUS. AVOID CONTACT AND DO NOT INHALE.
45Ninhydrin TestA positive test is either a purple/violet color OR a yellow color.Yellow indicates the presence of a specific amino acid, proline.Purple indicates the presence of any other free amino acid.
47Nutrition ActivityWhen the students at your table have shared all of their data and taught each other these different tests:Read the section on Nutritional AnalysisYou may do this individually or with others.You need to know:How to convert pounds (lbs) to kilograms (kg)How to calculate BMR (know the equation)Have a basic understanding of the Prudent Diet
48Clean upDispose of any solutions containing the following reagents in the appropriately labeled containers on the side counter:Dispose of chromatography solvent, ethanol, and Sudan IV in the solvent waste container.Dispose of Lugol’s solution, Benedicts reagent, and Biuret’s reagent in the base waste container.Dispose of ninhydrin filter papers in the ninhydrin waste bucket.Keep waste containers closed when not disposing of waste, and do not fill past the indicated line on waste containers. Inform your TA of any full waste containers.Rinse all used test tubes (after dumping contents in proper waste container) and mortar and pestles and return to the middle of the bench.Replace all bottles and other equipment to the middle of the bench in an orderly manner.Make sure all hot plates are offClean all pipettes and return them to the “clean pipette” container
49Learning ObjectivesBe able to identify the three classes of organic molecules based on their functional groups.Know the difference between a monomer and a polymer.Know the monomers for the organic molecules discussed.Know each of the diagnostic tests performed in classKnow the general procedureKnow how each test worksKnow what positive and negative results look like.Nutritional Analysis- You need to know:How to convert pounds (lbs) to kilograms (kg)How to calculate BMR (know the equation)Have a basic understanding of the Prudent Diet
50Chemical Tests: Test Test for Color before Color after Detects BenedictsLugolsSudan IVBiuretNinhydrin
51Biomolecules Summary Products Biomolecule Building Blocks Type of Bond Binding MechanismProductsCarbohydratesSimple SugarsGlycosidicDisaccaridesPolysaccharidesLipidsGylcerol Fatty AcidsDehydrationTriglyceridesPhospholipidsSteroidsProteinsAmino AcidsPeptidePolypeptides
52TestTest forColor beforeColor afterPositive control, negative controlBenedictsReducing sugarsPale bluePale blue – none, green – low, brown – intermediate, orange red - highGlucose solution, distilled waterLugolsStarchYellowBlue - blackPotato, distilled waterSudan IVLipidsLight redRed stained oil dropletsAny oil, distilled waterBiuretPeptide bondsVery pale blueVioletMilk, distilled waterNinhydrinFree amino acidsNo ColorOn drying, may be either purple or yellowAmino acid methionine, distilled water