Lipids Experiment PDF

Lipids BIOCHEMISTRY Laboratory  Lipids There are four major families of biological molecules, including proteins, nucleic acids, carbohydrates, and lipids.  Lipids The lipids, unlike members of the other three families, members of...

Lipids BIOCHEMISTRY Laboratory  Lipids There are four major families of biological molecules, including proteins, nucleic acids, carbohydrates, and lipids.  Lipids The lipids, unlike members of the other three families, members of this families do not share a common structural feature, but rather share a common physical property; they are hydrophobic, nonpolar molecules. Lipids perform many essential functions in the human body, including energy storage, cushioning, and insulation. Cells store energy for long-term use in the form of fats. Lipids also provide insulation from the environment for plants and animals.  Lipids Lipids are also the building blocks of many hormones and an important constituent of all cellular membranes. Lipids include fats, waxes, phospholipids, and steroids. Dehydroepiandrosterone  Lipids The most abundant class of lipids and major components of depot or storage lipids in plant and animal cells are the triglycerides. Triglycerides that are generally liquid at room temperature are called oils and those that are generally solid are called fats. Lipids also include - waxes which are esters of fatty acids and long chain monohydric alcohols; - phosphorus containing compounds such as phosphatides, sphingomyelin, and cerebrosides which are readily isolated from the nervous tissues; - and steroids such as cholesterol and hormones.  Lipids Fats have a three carbon (3C) backbone called GLYCEROL and FATTY ACID CHAIN (which is a string of carbon and hydrogen atoms).  Lipids When an “OH” from the glycerol binds with an “H” from the fatty acid, it forms H2O that gets released. Then the two molecules link up.  Lipids The linking of glycerol and fatty acids produces monoglycerides, diglycerides, and triglycerides depending on the number of chain linkage. Lipids Lipids  Lipids Lipids: a heterogeneous class of naturally occurring organic compounds classified together on the basis of common solubility properties insoluble in water, but soluble in aprotic organic solvents including diethyl ether, chloroform, methylene chloride, and acetone Amphipathic in nature  Lipid Structure Open Chain forms fatty acids, triacylglycerols, sphingolipids, phosphoacylglycerols, glycolipids, lipid-soluble vitamins, prostaglandins, leukotrienes, and thromboxanes Cyclic forms cholesterol, steroid hormones, and bile acids  Lipids Structurally, fats and oils can contain double bonds in their structure, which causes the molecule to either pack closely together or not, and the presence of such bonds define their saturation Saturated Fat / Oil – Double bonds are absent; tight packing of molecules happen Unsaturated Fat / Oil – Double bonds are present; tight packing of molecules is impossible due to contortions caused by the bonds Lipids Lipids  TEST & REACTIONS  PHYSICAL PROPERTIES OF LIPIDS SOLUBILITY Solubility Solubility  Solubility cottonseed oil / Beeswax / Solvents cholesterol linseed oil paraffin wax Water Insoluble Insoluble Insoluble ethyl alcohol Insoluble Insoluble Insoluble Ether Soluble Soluble Soluble Pentane Soluble Soluble Soluble The test is based on the property of solubility of lipids in organic solvents and insolubility in water. PRINCIPLE: The oil will float on water because of lesser specific gravity. EMULSIFICATION  Emulsification Fats/Oils Observation Linseed oil - oil become dispersed into smaller globules when mixed with the soap solution - Soap solution is a good emulsifying agent - Emulsifiers are molecules that have both polar and nonpolar regions and thus capable of interacting with both lipids and water. When emulsifier mixes with lipids and water, it may act to suspend small droplets of lipid in water. The lipid does not dissolve in water, but is broken into smaller fragments that may remain suspended for longer periods of time. DRYING OILS  Drying Oils Fats/Oils Observation - a so-called drying oil, which means that it hardens upon exposure to air Unboiled linseed oil - raw/unboiled lined oil dries very slowly, taking weeks to fully cure - Dries faster - The actual boiling of some oils changes their drying characteristics. With linseed oil, Boiled linseed oil though, it is the addition of certain solvents that causes linseed oil to dry more quickly, acting as if it were boiled. CHEMICAL REACTIONS OF LIPIDS USED AS IDENTIFICATION TEST  TEST FOR DEGREE OF UNSATURATION Test for Degree of Unsaturation Test for Degree of Unsaturation  Test for Degree of Unsaturation The degree of unsaturation in a lipid is measured by its iodine number defined as the number of grams of iodine that would add to the double bonds present in 100 grams of the lipid. Animal fats have low iodine numbers while vegetable oils have higher values. The iodine value is a measure of the relative degree of unsaturation in oil components. The greater the iodine value, the more unsaturation and the higher the susceptibility to oxidation.  Test for Degree of Unsaturation Substance Tested Iodine Value oleic acid 90.1 cooking oil linseed oil 155-205  Test for Degree of Unsaturation Substance Tested Number of drops of iodine absorbed oleic acid - unsaturated so will decolorize the iodine solution upon addition of the solution - More iodine will be absorbed cooking oil - depending on its type, certain oils like palm oil is unsaturated which may absorb iodine at all linseed oil - unsaturated so will decolorize the iodine solution upon addition of the solution - More iodine will be absorbed ACROLEIN TEST  Acrolein Test Acrolein test is used to detect the presence of glycerol or fat. When fat is treated strongly in the presence of a dehydrating agent like potassium bisulphate (KHSO4), the glycerol portion of the molecule is dehydrated to form an unsaturated aldehyde, acrolein that has a pungent irritating odor.  Acrolein Test Acrolein (prop-2-enal) is a compound formed by dehydration of glycerol, so its presence indicates the presence of glycerides ester (usually triglyceride) ie. a fat or oil. The smell is a bit like a barbeque or burnt grease. Reaction with AgNO3 Substance Odor (Tollen’s Test) Test for aldehydes, which are oxidized to carboxylic Tested + (burnt grease) acids. + formation of silver mirror + due to presence of glycerol + formation of silver mirror glycerol which forms acrolein acrolein is an aldehyde paraffin no silver mirror due to absence of - due to the lack of glycerol wax acrolein + due to presence of glycerol + formation of silver mirror linseed oil which forms acrolein acrolein is an aldehyde SALKOWSKI TEST  Salkowski Test Used to detect cholesterol in a solution Cholesterol undergoes dehydration in the presence of H2SO4 to give 3,5-cholestadiene which dimerizes to bis-cholestadiene.  Salkowski Test When cholesterol reacts with concentrated sulfuric acid, it creates different chemical products called sulfonated products. These products give specific colors: Mono-sulfonated products (only one sulfur group added) give a green color. Di-sulfonated products (two sulfur groups added) give a red color.  Salkowski Test In the test tube shown in the picture: There are two layers because the solution contains two substances that don't mix well (like oil and water). The top layer (chloroform) contains the cholesterol and shows the red and/or green color. The bottom layer (acid) is the sulfuric acid.  Salkowski Test Sample Observation positive reaction Reddish violet color in the upper Cholesterol chloroform layer while lower layer is yellow with green fluorescence  Did you know? Sterols are a type of fat found in plants and animals. Their main job is to help build and maintain cell membranes, the outer layer of cells. They keep the membrane strong but flexible, so it can protect the cell and let things in and out as needed. In animals, the most well-known sterol is cholesterol, which also helps the body make important substances like hormones (e.g., estrogen, testosterone), vitamin D, and bile acids (used for digesting fats). In plants, sterols (called phytosterols) help protect and strengthen their cells. When humans eat plant sterols, they can help lower bad cholesterol in the blood, which is good for heart health.  POST-LAB QUESTIONS  Post-Lab Question 1. What happens chemically when a drying oil “dries”? What structural characteristics are necessary in good drying oil?  Post-Lab Question 2. Give 2 examples of lipids that do not give a positive acrolein test? Explain.  Post-Lab Question 3. Using data and results in your experiment, tell how you would distinguish between the following pairs of substances. Give names of the tests and results expected with each substance. a. stearic acid and cholesterol b. linseed oil and mineral oil c. lard and paraffin wax PRAYER