TESTS FOR LIPIDS Lipids constitute include fats, waxes, sterols, a broad group of (such naturally as occurring molecules

that A, D, E, and

fat-soluble vitamins

vitamins

K), monoglycerides, diglycerides, triglycerides, phospholipids, and others. The main biological functions of lipids include energy storage, as structural components of cell membranes, and as important signaling molecules. Lipids may be broadly defined as hydrophobic or amphiphilic small molecules; the amphiphilic nature of some lipids allows them to form structures such as vesicles, liposomes, or membranes in an aqueous environment. Biological lipids originate entirely or in part from two distinct types of biochemical subunits or "building-blocks": ketoacyl andisoprene groups. Using this approach, lipids may be divided into eight categories: fatty and

acids, glycerolipids,glycerophospholipids, sphingolipids, saccharolipids,

polyketides (derived from condensation of ketoacyl subunits); and sterol lipids and prenol lipids (derived from condensation of isoprene subunits). Although the term lipid is sometimes used as a synonym for fats, fats are a subgroup of lipids called triglycerides. Lipids also encompass molecules such as fatty acids and their derivatives (including tri-, di-, monoglycerides, andphospholipids), as well as other sterolcontaining metabolites such as cholesterol.[6] Although humans and other mammals use various biosynthetic pathways to both break down and synthesize lipids, some essential lipids cannot be made this way and must be obtained from the diet. The Ethanol Emulsion Test is a food test which determines the presence of a broad group of naturally occurring compounds known as lipids. Lipids consist of fats and oils. Other lipid tests include the Grease Spot Test and the Sudan Stain Test. The Grease spot test is performed on fats - lipids which are solid at room temperature. Sudan stain colours lipids red, but is a less common bench reagent than ethanol. The Ethanol Emulsion Test is the most common test amongst the three.

BROWN PAPER TEST Lipids can be tested with a brown paper. If it evaporates after being dropped on the paper, lipids are not present. If transparent spots appear on the paper, lipids are present. SUDAN IV TEST In the Sudan IV Test, dark red Sudan IV (a common dye) is added to a solution. If lipids are present, they will be dyed red.

Testing for the presence of lipids: Sudan red test

Demonstrate how to perform this test and refer to student hand-out. Add 2ml of any oil and 2ml of water to a test tube. Then add 2-5 drops of Sudan red to the mix. Shake. Then repeat with a test tube containing only water. Students will see that Sudan red with stain the fat molecules.

Allow students to test different samples for the presence of lipids. Interesting is to test milk with different fat content. The more fat it contains, the more particles the Sudan red will stain.

Be careful. Sudan red can stain clothes.

EMULSION TEST Lipids do not dissolve in water, but do dissolve in ethanol. This characteristic is used in the emulsion test. Do not start by dissolving the sample in water, but instead shake some of the test sample with about 4 cm of ethanol. Decant the liquid into a test tube of water, leaving any undissolved substances behind. If there are lipids dissolved in the ethanol, they will precipitate in the water, forming a cloudy white emulsion. The test can be improved by adding the dye Sudan III, which stains lipids red. ACROLEIN TEST

Because when a fat is heated strongly in the presence of a dehydrating agent such as KHSO4, the glycerol portion of the molecule is dehydrated to form the unsaturated aldehyde, acrolein (CH2=CH-CHO), which has the peculiar odor of burnt grease. A sample is heated with potassium bisulfate, and acrolein is released if the test is positive. That's why Acrolein test is a test for the presence of glycerin or fats. Acrolein is a compound formed by dehydration of glycerol, so its presence indicates the presence of a glyceride ester (usually a triglyceride) ie a fat or oil. The smell is a bit like a barbecue.

TESTS FOR CARBOHYDRATES THE MOLISCH TEST Shows positive test for: All carbohydrates. Monosaccharides give a rapid positive test. Disaccharides and polysaccharides react slower. Reactions: The test reagent dehydrates pentoses to form furfural (top reaction) and dehydrates hexoses to form 5-hydroxymethyl furfural (bottom reaction). The furfurals further react with -naphthol present in the test reagent to produce a purple product (reaction not shown).

How to perform the test: Two ml of a sample solution is placed in a test tube. Two drops of the Molisch reagent (a solution of -napthol in 95% ethanol) is added. The solution is then poured slowly into a tube containing two ml of concentrated sulfuric acid so that two layers form. A positive test is indicated by: The formation of a purple product at the interface of the two layers.

a negative test (left) and a positive test (right)

IODINE / POTASSIUM IODIDE TEST Shows positive test for: Starch Reactions: Iodine complexes with starch to form a blue-black product. Note that other polysaccharides may give other colors including blue or red. How to perform the test: Two ml of a sample solution is placed in a test tube. Two drops of iodine/potassium iodide solution and one ml of water are added. A positive test is indicated by: The formation of a blue-black complex.

a negative test (left) and a positive test (right) BENEDICTS TEST Shows positive test for: Reducing sugars Reactions: Reducing sugars are oxidized by the copper ion in solution to form a carboxylic acid and a reddish precipitate of copper (I) oxide.

How to perform the test: One ml of a sample solution is placed in a test tube. Two ml of Benedict's reagent (a solution of sodium citrate and sodium carbonate mixed with a solution of copper sulfate) is added. The solution is then heated in a boiling water bath for three minutes. A positive test is indicated by: The formation of a reddish precipitate within three minutes.

a negative test (left) and a positive test (right)

BARFOEDS TEST Shows positive test for: Reducing monosaccharides Reactions: Reducing monosaccharides are oxidized by the copper ion in solution to form a carboxylic acid and a reddish precipitate of copper (I) oxide within three minutes. Reducing disaccharides undergo the same reaction, but do so at a slower rate.

How to perform the test: One ml of a sample solution is placed in a test tube. Three ml of Barfoed's reagent (a solution of cupric acetate and acetic acid) is added. The solution is then heated in a boiling water bath for three minutes. A positive test is indicated by: The formation of a reddish precipitate within three minutes.

a negative test (left) and a positive test (right)

SELIWANOFFS TEST Shows positive test for: Ketoses Reactions: The test reagent dehydrates ketohexoses to form 5-hydroxymethylfurfural. 5hydroxymethylfurfural further reacts with resorcinol present in the test reagent to produce a red product within two minutes (reaction not shown). Aldohexoses react to form the same product, but do so more slowly.

How to perform the test: One half ml of a sample solution is placed in a test tube. Two ml of Seliwanoff's reagent (a solution of resorcinol and HCl) is added. The solution is then heated in a boiling water bath for two minutes. A positive test is indicated by: The formation of a red product.

a negative test (left) and a positive test (right) BIALS TEST Shows positive test for: Pentoses Reactions: The test reagent dehydrates pentoses to form furfural. Furfural further reacts with orcinol and the iron ion present in the test reagent to produce a bluish product (reaction not shown).

How to perform the test: Two ml of a sample solution is placed in a test tube. Two ml of Bial's reagent (a solution of orcinol, HCl and ferric chloride) is added. The solution is then heated gently in a Bunsen Burner or hot water bath. If the color is not obvious, more water can be added to the tube. A positive test is indicated by: The formation of a bluish product. All other colors indicate a negative result for pentoses. Note that hexoses generally react to form green, red, or brown products.

two negative tests (left, middle) and a positive test (right)