Analyses of Lipids

Question 0

What is the difference between saturated and unsaturated fatty acids?

Answer 0

Saturated: fatty acids with single bonds between all carbon pairs

Unsaturated: fatty acids that contain double bonds between one or more pairs of carbon atoms.

Question 1

Name five lipids.

Answer 1

Fatty acyls (FA)
Glycerolipids (GL) 
Glycerophospholipids (GP)
Sterol lipids (ST) 
Sphingolipids (SP)

Question 2

What are the fatty acids esters of glycerol known as?

Answer 2

Glycerides.

Question 3

What does the prefix mono, di and tri relate to?

Answer 3

Indicates the number of fatty acids esterified to glycerol. Tricylglycerols are the most important class of lipids as they are the major components of fats and the predominant form found in plasma.

Question 4

What are steroid esters?

Answer 4

They exist in nature, a mixture of free sterol and esters with fatty acids.

Question 5

Sphingolipids are a complex group of lipids. True or False?

Answer 5

True.

Question 6

What is the structure of phospholipids?

Answer 6

Phosphate group and fatty acid is esterified to a glycerol.

Question 7

What are steroid lipids derived from?

Answer 7

Cholesterol.

Question 8

What is the structure of phosphosphingolipids?

Answer 8

Sphingolipids contain the long chain alcohol sphingosine, or one of its derivatives. The sphingolipids that contain a phosphate group, i.e. Phosphosphingolipids, are normally referred to as sphingomyelins which were first isolated from brain and nervous tissue where they are abundant.

Question 9

What are simple and complex lipids?

Answer 9

Simple 
Waxes
Acylglycerols 
Sterol esters 
Vitamins A, E and K

Complex
Phospholipids
Glycolipids

Question 10

Why are extraction of lipids necessary?

Answer 10

Lipids from liquid samples or cell suspensions may be extracted directly into organic solvents but solid samples will require prior treatment, such as homogenisation or ultrasonication.

Question 11

What is the method for extraction?

Answer 11

Lipids are hydrophobic, one phase extraction using methanol-chloroform-water mixture. The methanol disrupts the lipid-protein membrane complexes and inactivates the lipases, and the chloroform dissolves the lipids. A salt solution is often added and the non-lipid substances are taken up into this aqueous phase by vigorous shaking. They can be removed in the upper aqueous layer following centrifugation to separate the phases. Repeated extraction and washing will improve the isolation of uncontaminated lipids in the lower chloroform layer, which can then be analysed by the method of choice.

Question 12

Define hydrolysis.

Answer 12

Hydrolysis of acylglycerols splits the fatty acids from the glycerol.

Question 13

Define saponification.

Answer 13

The chemical hydrolysis of acylglycerols.

Question 14

What is thin layer chromatography?

Answer 14

It is based on the separation of a mixture of compounds as it migrates with the help of a suitable solvent through a thin layer of absorbent material which has been applied to an appropriate support. This can be looked upon as an “open column” system.

Question 15

What is the migration of a compound in a given TLC system?

Answer 15

By its RF value which is the distance travelled by compound.
RF= distance of the solvent front from the origin.

Question 16

What is the absorbent material used?

Answer 16

Silica gel (silicic acid combined with a small amount of gypsum as a binding agent) 
Alumina (for the separation of steroids).

Question 17

What is the solvent material?

Answer 17

Based on the nature of the compounds to be separated.

Question 18

What are the methods of visualisation?

Answer 18

Spray reagents; yields coloured derivatives on reaction with the separates spots.
Iodine vapour
Fluorescence and UV absorption
Autoradiography

Question 19

What is gas liquid chromatography?

Answer 19

Useful technique in lipid analysis, particularly for the separation of very similar compounds within classes. Because of the wide variations in structure and properties between classes it is not usually possible to resolve members of different classes on the same column. GLC is useful for both quantitative and qualitative analysis and also in the investigation of lipid structure.

Question 20

What lipids require GLC?

Answer 20

Fatty acids
Mono-triglycerides
Phospholipids

Question 21

What is the mobile phase for GLC?

Answer 21

Inert gas.

Question 22

What is the material for stationary phase in GLC?

Answer 22

Liquid.

Question 23

Why does temperature need to be above boiling point for GLC?

Answer 23

Compounds require boiling point to be separated. The boiling point of many compounds (Glycerides and free fatty acids) is too high. The compound must be chemically transformed into analogs that are more volatile.

Question 24

What does the salkowski and Liebermann-burchard test for?

Answer 24

Test for cholesterol.

Question 25

What is the salkowski test?

Answer 25

The presence of a double bond in one cholesterol rings is responsible for its ability to form colour products in the presence of concentrated inorganic acids.
Sulfuric acid; results in dehydration of cholesterol molecule with a formation of a red bicholestadien disulphonate.
Bluish colour; between the 1st layer (chloroform) and 2nd layer.

Question 26

What is the Liebermann-Burchard test?

Answer 26

A positive result is the formation of a deep green colour.
This is due to hydroxyl group of cholesterol reacting with the reagents and increasing the conjugation of the un-saturation in the adjacent fused ring.