Exercise 8

LIPIDS
Bate AC, Enriquez IJ, Loberiano AK, Ochea C

OBJECTIVES
The objective/s of the experiment is as follows:
● To determine the different characteristics of different lipids through various qualitatitive
tests, and;
● To produce soap from a certain type of oil

RESULTS AND DISCUSSION

Lipids are esters which are long chains of fatty acids alcohols that occur in varieties of
structures. Common lipids are oils, phospholipids, fats, fatty acids, soaps, and waxes that also
have different particular purposes in nature. The characteristic of lipids being a non-polar
compound and soluble in organic solvents occurs with specific functions. Lipids are known to be
an energy reserve, being an efficient source as it can contain greater amounts in a reduced
space. It also facilitates the regulation of body conditions and signaling for brain activities.
Moreover, the presence of fat tissues serves as protection for delicate organs.​12 The previously
mentioned are some of the general functions of lipids that contribute to biological processes.
This experiment helps in recognizing the presence of lipids as well as its characteristics from
different materials that are commonly encountered to further assess how it can be useful for
certain processes.

I. Spot Test for Fat

Fats and oils have high boiling points so they cannot absorb enough heat to evaporate. Fat or
oil on paper diffracts light and produces a translucent spot as it penetrates on the paper that
causes a greasy texture.​5 On the other hand, water only wets the paper but the spot will tend to
completely disappear from the paper indicating that there is no presence of lipid.​11

Figure 1. ​Spot test results for different kinds of oils.

II. Color, Solubility, and Density

Table 1. ​Summary of data for the color, solubility and density of different oils.

Coconut Butter Palm Olive Corn

Water color colorless yellow faint yellow yellow yellow

solubility insoluble insoluble insoluble insoluble insoluble

density less dense less dense less dense less dense less dense

HCl color clear clear clear clear cloudy

solution yellow solution solution white
solution with a with a solution
yellow spot yellow spot

solubility insoluble insoluble insoluble insoluble insoluble

density more less dense less dense less dense less dense
dense

NaOH color faint yellow faint yellow faint yellow faint yellow clear
solution
with faint
yellow spot

solubility insoluble insoluble insoluble insoluble insoluble

density less dense less dense less dense less dense less dense

95% color cloudy clear cloudy cloudy cloudy

Ethanol white solution white solution solution
with a spot with a with a faint
of yellow yellow spot yellow spot

solubility insoluble insoluble insoluble insoluble insoluble

density more more more more more

dense dense dense dense dense

Hexane color colorless faint yellow colorless colorless colorless

solubility insoluble insoluble insoluble insoluble insoluble

density less dense more more more less dense

dense dense dense

Chloroform color cloudy faint yellow cloudy cloudy colorless

white white yellow
 solution

solubility insoluble insoluble insoluble insoluble insoluble

density less dense less dense less dense less dense less dense

Lipids contain large nonpolar regions that contribute to its solubility property. 13 ​ ​As indicated in
the table it is not soluble in polar solvents. Lipids are nonpolar thus it is soluble in non-polar and
in weak polar organic solvents, including ether, chloroform, benzene, and acetone, hexane.
These oils vary in their ability to dissolve in different organic solvents. They mostly have lesser
density compared to the solvent and usually retain their original color after the addition of the
solvents.

III. Acrolein Test

The acrolein test is performed to identify the presence of fats or glycerin. It rests on the
assumption that water molecules are removed from the glycerol by adding KHSO​4​. This
reaction, also referred to as the “dehydration reaction”​5​, forms the unsaturated aldehyde called
acrolein(CH​2​=CH​-​CHO) and is identifiable by its pungent odor similar to burnt cooking grease
and some black color precipitates settling at the bottom.

Figure 2. ​Reaction of glycerol in Acrolein Test.

Yet, in the experiment, this expected result was not obtained. Burnt grease odor with black
precipitates was not observed when 3 drops of the sample with 0.2 g KHSO​4 reagent in a test
tube was heated. Instead, the odor of the oil sample remains in its original state. Coconut oil still
smells like coconut oil and glycerol has still no odor. Theoretically, coconut oil and glycerol
should exhibit a positive result because both contain fat and glycerin in its structure. Both had
the capability to undergo dehydration process and produce acrolein as a product. But a factor
that might affect the result of the experiment was that the reagent used might not effectively let
the samples undergo a dehydration reaction yielding improper results.

Table 2. ​Summary of data for Acrolein Test for Coconut Oil and Glycerol.

Oil sample Observations

Coconut oil Had a coconut-like odor

Glycerol Had no odor

 IV. Test for Unsaturation

The Unsaturation Test is used to determine the amount of unsaturated fatty acids in a sample.
The principle behind this test is that if a lipid sample is more unsaturated, the more they are
likely to absorb iodine​5​. For this experiment, all samples but for butter resulted in a
reddish-brown color. This means that all the other oil samples are unsaturated​14​. Butter yielded
a lighter color compared to the other samples because it is a saturated fat​15​.

[PLQ 1]​
An oil’s iodine number indicates the amount of iodine required to saturate the fatty acids
present in the oil or fat. Coconut oil has an iodine number that ranges from 7.5-10.5 while olive
oil has an iodine number from 74-94.​1​ Therefore, olive oil absorbs more iodine than coconut oil,
and is more unsaturated than coconut oil.

Table 3. ​Summary of data for the Test of Unsaturation for different oils.

Oil samples Observations

Coconut oil Resulted in a reddish-brown color

Butter Resulted in an orange-brown color

Palm oil Resulted in a reddish-brown color

Olive oil Resulted in a reddish-brown color

Corn oil Resulted in a reddish-brown color

V. Liebermann-Burchard Test

Liebermann-Burchard Test is a chemical estimation used to detect the presence of cholesterol.

It follows the principle that the sample is reacted with acetic anhydride, the solvent and
dehydrating agents, then sulfuric acid used as the dehydrating and oxidizing agent to produce
colored substances which indicates the presence of cholesterol. A positive result is obtained
when the solution becomes red , then blue, and finally bluish –green color after a few minutes.​8
This gradual color change occurs due to the ˆOH group of cholesterol and the unsaturation
found in the adjacent that fused to form ring.

In the experiment, 10-20 drops of acetic anhydride and 2 drops conc. sulfuric acid was
cautiously added to the test tube of the mixture of 1-2 crystal grains of cholesterol and 3 mL
chloroform. It was gently mixed and the formation of color from lilac then blue to emerald green
was observed.
 Figure 3. ​Emerald green result for Liebermann-Burchard Test.

After complying the provided procedures, the solution did not achieve the expected color at first
which was the emerald green color and instead exhibited a light brown solution. It has achieved
the color in the image below when a solution of excess sulfuric acid was added to the first
solution. Hence, it can be inferred that certain errors have been encountered in this particular
test. For one, the precise amount of cholesterol was not provided so the solution was only
conducted with an estimated amount.​9 The amount of Sulfuric acid in the solution was also
suspected to be a source of error based on the observation of the solution that it had exhibited
the anticipated result when a solution of excess Sulfuric acid was added.​10 This has suggested
to perform another test with an increased amount of the acid but it was still difficult to achieve.

VI. Saponification

Corn oil, like any other vegetable oil, contains triacylglycerol (TAG), which can be broken down
into fatty acids and alcohols through hydrolysis. [PLQ
​ 3]​
This reaction is called saponification and it
usually occurs in the presence of a strong base​6​. In this experiment, 10% NaOH was used. [PLQ ​
3]​
The products formed are glycerol and 3 fatty acid salts. These fatty acid salts are called soaps,
and they have both polar and nonpolar ends.

Figure 2. ​Corn oil after the saponification process.

For this experiment, 25 mL of 10% NaOH was added to 15 mL corn oil and was heated and
stirred constantly until the solution became thick enough to stick to the stirring rod. The resulting
mixture was then transferred in a plastic mold and was left to cool overnight.

Corn oil contains various fatty acids, including oleic acid, linoleic acid, and linolenic acid.​7 These
unsaturated fatty acids are what makes the corn oil a possible component for soap.

REFERENCES

1. Gupta, R. C.; Kanwar, G. Determination of Iodine Numbers of Edible Oils. 1994.

2. Bockisch, M. In ​Fats and Oils Handbook;​ pp 53–54.
3. Fats. ​http://www.iseo.org/FFO/page2.html​.
4. Odoom, W.; Edusei, V. O. Evaluation of Saponification Value, Iodine Value and
Insoluble Impurities in Coconut Oils from Jomoro District in the Western Region of
Ghana. ​Asian Journal of Agriculture and Food Sciences​ 2015, ​3​ (5).
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Mar 12, 2020).
6. Lipids: Saponification (The Properties and Synthesis of Soap).​
7. Wang, T.; White, P. J. In ​Corn ​; Elsevier, 2019; pp 337–368.
8. (lieber)
9. Kingsley, G.R and Schaffert, R.R. ​Determination of Free and Total Cholesterol by Direct
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March 11, 2020)
10. Sperry, W.M. and Brand, F.C. ​The Calorimetric Determination of Cholesterol. ​1943.
https://www.jbc.org/content/150/2/315.full.pdf​ (Accessed March 11, 2020)
11. Supriya, N. ​Qualitative Analysis of Lipids. ​ 019.
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https://biologyreader.com/qualitative-analysis-of-lipids.html​ (Accessed March 11, 2020)
12. Mandal, A. ​Lipid Biological Functions. ​ 019
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https://www.news-medical.net/life-sciences/Lipid-Biological-Functions.aspx (Accessed
March 11, 2020)
13. Ch26: Lipids http://www.chem.ucalgary.ca/courses/351/Carey5th/Ch26/ch26-0.html
(accessed Mar 11, 2020).
14. Qualitative and Quantitative Tests for Lipids.
http://www.biologydiscussion.com/lipids/tests/qualitative-and-quantitative-tests-for-lipids/
13050 (accessed Mar 12, 2020).
15. Moll, J. Learn the Key Differences Between Saturated and Unsaturated Fats.
https://www.verywellhealth.com/difference-between-saturated-fats-and-unsaturated-fats-
697517 (accessed Mar 12, 2020).