Chemical tests for lipids

Requirements:

Glassware: Test tube, test tube stand, beaker, wash bottle, burner,    filter paper, Chemicals: chloroform, benzene, concentrate sulphuric acid, formaldehyde sulphuric acid solution, acetic anhydride, sodium carbonate, sodium chloride, calcium chloride.

Theory for lipids

Lipids are a large group of naturally occurring compound, characterized only by their water repellant property hence called hydrophobic. Their solubility in organic solvent is variable. Some are soluble in polar organic solvent like alcohol and other in non-polar solvents. Like hexane, benzene etc although many proteins and polysaccharides are also not soluble in water, they do not repel water. They are easily wetted by water. by water may absorb and swell with water hence called hydrophilic unlike the protein or carbohydrate the lipid have no specific functional group or linkage characterizing all of them although a majority belong to the group of oils and fats. Lipids like carbohydrates provide energy for work and life processes. They also form part of some structure in the cell membrane, wall of tissue and organ. The lipids also provide a non-aqueous medium in the body which helps in regulation of transport of many chemicals, due to their partition between lipid and aqueous layers. Thus lipids play an important role in living organism. Lipids also occur in conjugation with protein and are termed lipoproteins. They may be classified into (1) simple lipid (2) phospholipids (3) complex lipid.
 

Procedure for Chemical tests for lipid

Solubility test

Take two mL sample solution in different three test tubes, followed by addition two mL chloroform, water, and benzene is added in respective test tube. Mix and allow to stand. Oil or lipid soluble in organic solvent and insoluble in inorganic solvent.

Spot test

Take three mL chloroform in test tube and add five drop of sample. Mix and take three drops on filter paper and allow drying till chloroform evaporated.

Salkowaski test (Sulphuric acid test)

Take few drop of sample solution in the test tube and dissolve in chloroform followed by addition of equal amount concentrated sulphuric acid. In chloroform bluish red to purple color observe and in acid green fluorosence indicates the presence of cholesterol.

Formaldehyde sulphuric acid test

Take two mL chloroform solution in test tube followed by addition of two mL formaldehyde sulphuric acid solution. Pour the chloroform layer into other test tube and add two-three drop acetic anhydride. Appearance of blue colour confirms the presence of cholesterol.

Acetic anhydride Sulphuric acid test or (Liebermann–Burchard test)

Take small amount of sample solution in test tube and dissolve in chloroform followed by addition of  ten drops of acetic anhydride and two- three drop of concentrate sulphuric acid. Solution become red finally bluish green indicate the presence of cholesterol.

Saponification test

Take four mL of 2 % sodium carbonate solution in test tube and add four drops sample solution. Shake well and boil, and cool and divided into three parts. 

(a) In first part add few drop of concentrate hydrochloric acid. Fat separate and float, confirms the presence of fat. 

(b) In second part add sufficient amount of sodium chloride. White precipitate appears and floats on surface that indicates the presence of fat. 

(c) In third part add few drop of calcium chloride solution. In soluble calcium precipitate form, confirms the presence of fat.

Result: In the given sample lipid is present. It is confirm by the different tests such as solubility test, spot test, salkowaski test, formaldehyde sulphuric acid test, acetic anhydride. sulphuric acid test, saponification test.