Jonathan Aivazi

2/4/15
Laboratory 3: Properties of Lipids

Objective
The goal of this laboratory is to study the chemical properties of lipids by testing
their solubility in various solvents, their ability to emulsify in emulsifying agents, their
nonvolatile nature compared to other nonpolar substances, and the effect of reacting the
different hydrocarbon bonds to Iodine molecules.
Theory
Lipids are a fundamental building block of the plasma membrane at the cellular
level. Lipids are made up of glycerol and fatty acid chains. Glycerol is a polar
hydrocarbon molecule with three alcohol functional groups. Fatty acids are double or
single bonded non-polar hydrocarbon with a carboxylic group at one end. Lipids are
formed when a Nonpolar bond between one of the glycerol alcohols and the HO group
of the fatty acid forms, resulting in the dehydration of H2O and the formation of an ester
bond. In this way, the lipid contains both a polar glycerol head and a nonpolar
hydrocarbon tail.
There are several classes of lipids. Triglycerides are those that are esterified to a
molecule of glycerol, as explained above, such as olive oil. Another type are
phospholipids, the main component of the phospholipid - bilayer, consist of a phosphate
group bonded to the third alcohol of glycerol in addition to two fatty acid chains.
The chemical composition of the fatty acid chains differs between the unsaturated
and saturated forms of a hydrocarbon. When the hydrocarbon contains double bonds,
then the fatty acid is thought to be lacking the possibility of all carbons containing 2
hydrogen atoms and thus is thought to be unsaturated. These kinks resulting from the
double bonds in turn results in a weaker aggregation and less tighter packing of the
hydrophobic fatty acid regions.
The individual components of lipids themselves exhibit different solubility, which
indicates their polarity. Glycerol, influenced by the alcohol side groups is thus more
soluble in a polar solvent such as water. Olive oil, containing the hydrocarbon chain will
readily dissolve in ether, a Nonpolar organic and volatile solvent, but will resist

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2/4/15

dissolution in a polar solvent. Similarly, solid fat will be soluble in ether, as well as
somewhat soluble in the partly polar methanol.
One key property of oil particles is that if they are allowed to remain stagnant,
they will slowly coalesce on the surface of the solution due to interfacial tension between
the oil and water layers. Once disrupted, they will undergo emulsion into droplets that
will, conditions permitting, then aggregate once more. An emulsifying agent prevents
coalescence by stabilizing the emulsion and reducing interfacial tension through
interactions with the hydrophobic fat and forming hydrogen bonds with the water. This
stabilized group is called a micelle and represents the effect that detergents, soaps, bile
salts, and other emulsifying agents exert on oily substances.
The presence of fats themselves are often quantified by applying a droplet of fat
onto filter paper and observing their absorbance into the paper, in contrast to a volatile
substance such as ether, which rapidly evaporates without settling into the pores of the
filter paper.
Fats are also important for the saponification process, in which soaps are made. In
saponification, an alkaline base is introduced into the neutral fat, which produces glycerol
and fatty acids; the fatty acids merge with the alkali salt to form the basis for soap and the
hydroxyl group is distributed across the glycerol.
Another distinguishing factor between unsaturated and saturated fats is the ability
of Iodine to add across the double bond in chloroform solution. Iodine, which is naturally
brown, loses its color once it reacts with the double bonded carbon atoms. If the
hydrocarbon is saturated however, no color will be lost since Iodine will have no double
bond with which to react with.
Materials and Reagents

Glycerol
Olive oil
Solid fat
Ether
Methanol
Distilled water
Soap solution
Detergent solution

Jonathan Aivazi

2/4/15

Bile salt solution

Filter paper
Chloroform
Iodine
Pipettes and rubber bulbs

Procedure and Methods

Solubility of Lipids in Various Solvents
1.
2.
3.
4.

Obtain seven test tubes for each lipid solvent test

Transfer 0.1g or 1 drop of lipid into the appropriate test tube
Add 1 ml or 20 drops of solvent into the test tube
Record the solubility of lipid in each test tube

Emulsification of Lipids
1.
2.
3.
4.

Prepare 4 test tubes

In the meantime, suspend bile salts in 30 ml of water
Pipette out 2 drops of olive oil into each test tube
Transfer 2.5 ml of water, soap, bile salt, and detergent solution to separate test

tubes
5. Shake tubes for 30 seconds and record solubility
6. Add 1 ml of ether to each tube, shake, and record solubility
Grease spot test for Fats
1. Dissolve a small amount of solid fat in 1 ml ether in a dry test tube
a. If solution is not clear, add another 1 ml of ether
2. Place 1 drop of solution in the center of a filter paper
3. Allow the paper to dry
4. Observe the grease spot
Iodine absorption test
1.
2.
3.
4.
5.

Obtain two test tubes

Place 0.2 ml of olive oil and 0.2 g of solid fat into separate test tubes
Add 1 ml of chloroform to each test tube
Add 3 or 4 drops of iodine to each test tube
Record color change, if any

Data and Analysis

Part A: Solubility of liquids in various solvents
Tube
1
2
3

Lipid
Glycerol
Glycerol
Olive oil

Solvent
Water
Ether
Water

Solubility
Soluble
Insoluble
Insoluble

Jonathan Aivazi
4
5
6
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2/4/15

Olive oil
Ether
Soluble
Solid lipid
Water
Insoluble
Solid lipid
Methanol
Somewhat soluble
Solid lipid
Ether
Soluble
This study sought to determine the solubility of different lipids in assorted

solvents. It was predicted that lipids, which as discussed above, are generally non polar in
nature, would resist dissolution in polar solvents. This was confirmed when olive oil and
solid lipid were observed to be insoluble in water, but soluble in ether. Furthermore, solid
lipid was also somewhat soluble in methanol since methanol (CH3OH) is partly non polar
because of the methane parent molecule, and contains less OH bonds than water. In
contrast, Glycerol is a lipid consisting of 3 OH groups per molecule will attract and
possess the electrons on the molecule, producing a dipole moment, and subjecting itself
to solubility in polar substances such as water, a trend confirmed by our data.
Part B: Emulsification of Lipids
Tube
1
2
3
4

Lipid
0.2 ml olive oil
0.2 ml olive oil
0.2 ml olive oil
0.2 ml olive oil

Solvent solution
2.5 ml water
2.5 ml soap solution
2.5 ml bile salt solution
2.5 ml detergent solution

Resulting Solubility
Insoluble
Soluble
Soluble
Soluble

After 1ml Ether

Insoluble
Insoluble
Insoluble
Insoluble

The emulsification of lipid into particles was accomplished by using a solvent that
acted as an emulsifying agent, which solvated the lipid particle by interacting with the
non-polar portion of the emulsifying agent as the agent interacted simultaneously with the
solution. Examples of agents implemented in the study were bile salts, a natural source
that aids in the digestive system, soaps, a synthetic emulsifier, and likewise, detergents. In
all cases solubility was displayed as micelles formed in solution upon addition of the
agents. However, when a 1ml aliquot of ether was introduced into the solution,
insolubility resulted due to the destruction of the micelles because of the preference of a
lipid to dissolve in a non-polar phase.
Part C: Grease Spot Test for Fats

Jonathan Aivazi

2/4/15

The grease test was a qualitative test carried out to visually identify fats by
placing a drop of fat on filter paper and observing the effects. Fat is nonvolatile and thus
was predicted to remain fixed onto the filter paper, which was visually confirmed as
per the image above. Because fat has a high boiling point, any energy or heat absorbed by
the fat is not sufficient enough to induce it to evaporate. As a contrast, ether was used;
ether is a volatile substance that rapidly vaporizes and escapes.
Part D: Iodine Absorption Test
Lipid
Olive oil
Solid fat

Color change upon addition of Iodine + Chloroform

Loss of brown color
No change in color, iodine lipid solution remains brown

The ramifications of molecules that are saturated versus unsaturated was evident
when both olive oil which is an unsaturated lipid and solid saturated fat were treated with
Iodine, which has the ability to add across the double bond in chloroform solution. Iodine
lost its brown color upon application into the olive oil test tube; this is consistent with our
expectations that Iodine would add across the double bond prevalent in olive oil. Thus it
would shed its color. However, Iodine retained its color when added to solid fat. This
again is a reliable result since fat is fully saturated at its C H bonds and Iodine would be
prevented from reacting with the lipids.
This experiment was not without shortcomings however. Sources of error include
improper pipetting and transferring of solutes to the test tube, and inconsistent mixing or
shaking. While our results are consistent with our expectations, in each part of the
experiment products could have been faultily prepared by not properly stirring the

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2/4/15

solution to form or not waiting the proper amount of time after the reaction to record
observations.
Conclusion
Our results confirm the literature that the chemical characteristics such as nonpolarity of lipids are responsible for the physical properties such as their solubility and
emulsification in various solvents. The difference between unsaturated and saturated fats
was also demonstrated in the iodine absorption, corroborating previous scientific
literature.