MCNUR2B: BIOCHEMISTRY LABORATORY

EXPERIMENT NO. 4 LIPIDS ANSWER KEY

I. SOLUBILITY TEST
A. Different Solutions
1. Distilled Water:
Cottonseed oil is generally insoluble in water. Lipids like oils and fats are hydrophobic,
meaning they do not mix with water. In this case, you would observe that the oil forms a
distinct layer on top of the water.
2. Ethyl Alcohol:
Cottonseed oil is partially soluble in ethyl alcohol. Ethyl alcohol can dissolve some lipids, so
you might observe a partial dissolution of the oil in alcohol, resulting in a cloudy or
translucent solution.
3. Ether:
Cottonseed oil is soluble in ether. Ether is a nonpolar solvent that can dissolve lipids, so the
oil would likely completely dissolve in it.
4. Chloroform:
Cottonseed oil is soluble in chloroform. Chloroform, like ether, is a nonpolar solvent capable
of dissolving lipids, leading to a clear solution.
5. Benzene:
Cottonseed oil is soluble in benzene. Benzene is also a nonpolar solvent, and it can dissolve
lipids, resulting in a clear solution.
6. 5% Hydrochloric Acid:
Cottonseed oil is generally insoluble in hydrochloric acid. This is because hydrochloric acid is
an aqueous, polar solution, and lipids like oils do not mix with it. You would see the oil
floating on the acid.
7. 5% Sodium Hydroxide:
Cottonseed oil is partially soluble in sodium hydroxide. Sodium hydroxide can saponify or
react with the oil to form soap and glycerol. You might observe a soap-like substance
forming in the solution, indicating partial solubility.

SOLUTION OBSERVATION
Distilled water Insoluble
Ethyl alcohol Partially soluble
Ether Soluble
Chloroform Soluble
Benzene Soluble
5% HCl Insoluble
5% NaOH Partially soluble

These characteristics are based on the polarity of the solvents and the nature of lipids like
cottonseed oil, which are predominantly nonpolar. Nonpolar solvents like ether, chloroform,
and benzene can dissolve lipids, while polar solvents like water and aqueous solutions have
limited solubility for lipids. Ethyl alcohol falls in between as it is partially polar. The reactions
with hydrochloric acid and sodium hydroxide involve chemical changes due to their alkaline
or acidic nature.

Prepared by: Marlon P. Flores

NATSCI College Instructor
MCNUR2B: BIOCHEMISTRY LABORATORY
EXPERIMENT NO. 4 LIPIDS ANSWER KEY

B. Translucent Spot Test

The solubility of the cottonseed oil-ethyl alcohol mixture and the cottonseed oil-ether
mixture on a piece of bond paper can be compared as follows:
1. Cottonseed Oil-Ethyl Alcohol Mixture:
• Since cottonseed oil is partially soluble in ethyl alcohol, the mixture may have some
solubility characteristics.
• When a few drops of this mixture are dropped onto a piece of bond paper, you might
observe that the alcohol portion of the mixture evaporates relatively quickly, leaving behind
the cottonseed oil.
• The oil, being nonpolar, may interact with the paper fibers, creating a visible stain or
translucent mark on the paper.
2. Cottonseed Oil-Ether Mixture:
• Cottonseed oil is soluble in ether, which is a nonpolar solvent. This means that the
cottonseed oil will readily dissolve in ether.
• When a few drops of the cottonseed oil-ether mixture are dropped onto bond paper, the
ether portion will quickly evaporate, leaving behind the cottonseed oil.
• Similar to the ethyl alcohol mixture, the cottonseed oil, being nonpolar, may interact with
the paper fibers, creating a visible stain or translucent mark on the paper.

SOLUTION OBSERVATION
Cottonseed Oil-Ethyl Alcohol Translucent spot is visible
Cottonseed Oil-Ether Translucent spot is visible

In both cases, you would likely observe a visible stain on the paper due to the cottonseed
oil's interaction with the paper fibers. However, the speed of evaporation may vary, with the
ether portion evaporating more quickly.

II. TEST FOR UNSATURATION OF FATTY ACIDS

A. Bromine Test
The test for unsaturation of fatty acids involving carbon tetrachloride, oleic acid, and
bromine water is typically referred to as the bromine test. It's used to detect the presence of
unsaturated bonds (double or triple bonds) in organic compounds like oleic acid (a
monounsaturated fatty acid). Here are the characteristics of the results of this test:
• Carbon Tetrachloride: Carbon tetrachloride is used as the solvent for the test. It is
nonpolar and doesn't react with the substances involved. It mainly serves as a medium
for the reaction to take place.
• Oleic Acid (Unsaturated Fatty Acid): Oleic acid contains one or more unsaturated bonds,
which are susceptible to chemical reactions like bromination.
• Bromine Water: Bromine water (a solution of bromine gas in water) is added to the
mixture. Bromine is a reddish-brown liquid at room temperature.
When bromine water is added to the mixture, it reacts with the unsaturated bonds in the
oleic acid. The reddish-brown color of bromine water disappears as bromine is decolorized
during the reaction. The disappearance of the reddish-brown color indicates a positive result
for unsaturation, confirming the presence of double or triple bonds in the oleic acid. The
reaction between the bromine and the unsaturated bonds forms a colorless compound. You
would observe a decolorization of the bromine water solution, turning it from reddish-
brown to colorless.

SOLUTION OBSERVATION INFERENCE

Oleic acid Colorless Unsaturated

Prepared by: Marlon P. Flores

NATSCI College Instructor
MCNUR2B: BIOCHEMISTRY LABORATORY
EXPERIMENT NO. 4 LIPIDS ANSWER KEY

B. Hanus Iodine Test

It is used to determine the degree of unsaturation (the number of double bonds) in fatty
acids and oils. Here are the characteristics of the results for the three test tubes:
1. Test Tube 1 (Palmitic Acid):
• Palmitic acid is a saturated fatty acid, meaning it contains no double bonds.
• When CHCl3 (chloroform) and Hanus iodine solution are mixed with palmitic acid,
there is no reaction, and the solution remains unchanged in color.
2. Test Tube 2 (Oleic Acid):
• Oleic acid is a monounsaturated fatty acid, containing one double bond.
• When CHCl3 and Hanus iodine solution are mixed with oleic acid, the iodine solution
reacts with the double bond(s) present.
• The solution changes color, indicating the formation of iodine-substituted products,
which is a positive result for unsaturation. The color change may range from pale
yellow to brown, depending on the degree of unsaturation.
3. Test Tube 3 (Cottonseed Oil):
• Cottonseed oil is a mixture of various fatty acids, including both saturated and
unsaturated ones.
• When CHCl3 and Hanus iodine solution are mixed with cottonseed oil, the iodine
solution reacts with the unsaturated bonds in the various fatty acids present.
• The solution changes color due to the reaction with the multiple unsaturated
components in the oil. The extent of color change depends on the overall
unsaturation in the oil.

SOLUTION OBSERVATION INFERENCE

Palmitic Acid Remains unchanged in color Saturated
Oleic Acid Pale yellow to brown Unsaturated
Solution changes color
Cottonseed Oil (depends on the overall Unsaturated
unsaturation)

III. ACROLEIN TEST

In both test tubes, you may observe a change in the appearance or physical properties of the
substances being heated. This could include the generation of heat, production of gases, or changes
in color or odor.
The characteristics of the results for the two test tubes, one containing glycerol and a pinch of
KHSO4 (potassium bisulfate) and the other containing cottonseed oil and a pinch of KHSO4, when
heated over a low flame, can be described as follows:
1. Test Tube 1 (Glycerol and KHSO4):
• Glycerol (also known as glycerin) is a trihydric alcohol that is commonly used in various
chemical reactions. KHSO4 is an acidic salt (potassium bisulfate) that can act as a dehydrating
agent.
• When heated over a low flame, glycerol can undergo dehydration in the presence of KHSO4.
Dehydration typically involves the removal of water molecules from a compound. The
reaction between glycerol and KHSO4 will result in the formation of an unsaturated
compound, such as an alkene.
• When glycerol is subjected to the Acrolein test with KHSO4, the characteristic odor released
is a pungent, acrid, and irritating smell.
2. Test Tube 2 (Cottonseed Oil and KHSO4):
• Cottonseed oil is a mixture of various fatty acids and triglycerides, some of which may
contain unsaturated bonds. KHSO4, as in the first test tube, can act as a dehydrating agent
under heat.

Prepared by: Marlon P. Flores

NATSCI College Instructor
MCNUR2B: BIOCHEMISTRY LABORATORY
EXPERIMENT NO. 4 LIPIDS ANSWER KEY

• When heated, the KHSO4 may react with the unsaturated bonds present in the cottonseed
oil, causing dehydration of these bonds.
• The characteristic odor released by cottonseed oil when subjected to the Acrolein test with
KHSO4 (potassium bisulfate) is similar to the odor associated with acrolein formation.

SOLUTION OBSERVATION INFERENCE

Pungent, acrid, and irritating
Glycerol + KHSO4 Glycerol is present
odor
Pungent, acrid, and irritating
Cottonseed Oil + KHSO4 Glycerol is present
odor

The odor is associated with the formation of acrolein, a volatile organic compound resulting
from the dehydration of glycerol under acidic conditions.

Acrolein has a distinctive, unpleasant odor often described as a combination of burnt or charred
fat, with a somewhat sharp and acrid note. It is known for its strong, irritating, and choking
smell, which can be highly noxious in concentrated form.

The strong and recognizable odor of acrolein in the Acrolein test serves as a qualitative indicator
that glycerol has undergone dehydration, leading to the formation of acrolein as a product of
the reaction. The test is used in various laboratory and industrial applications to identify the
presence of glycerol and to distinguish it from other compounds.

IV. TEST FOR RANCIDITY

The test tubes containing fresh and rancid coconut oil, along with various pH indicators, can help
assess the acidity or alkalinity of these oils. Here are the characteristics of the results for each
combination:
1. Test Tube 1 (Fresh Coconut Oil and Phenolphthalein):
• Fresh coconut oil is typically neutral or slightly acidic in nature.
• Phenolphthalein is a pH indicator that turns pink or red in the presence of a base but
remains colorless in an acidic or neutral environment.
• In this test tube, you would likely observe no color change in the presence of
phenolphthalein, indicating that fresh coconut oil is not significantly alkaline.
2. Test Tube 2 (Fresh Coconut Oil and Methyl Orange):
• Methyl orange is a pH indicator that transitions from red in an acidic environment to
yellow in a neutral to slightly alkaline environment.
• Fresh coconut oil is expected to result in a yellow or possibly an orange color in this test
tube, indicating a slightly alkaline nature.
3. Test Tube 3 (Fresh Coconut Oil and pH Paper):
• pH paper, when dipped into fresh coconut oil, may show a pH reading in the neutral to
slightly acidic range (around 7 or lower on the pH scale).
4. Test Tube 4 (Rancid Coconut Oil and Phenolphthalein):
• Rancid coconut oil may contain free fatty acids, leading to acidity.
• Phenolphthalein, in the presence of an acidic environment, will remain colorless. So, you
would observe no color change in this test tube.
5. Test Tube 5 (Rancid Coconut Oil and Methyl Orange):
• Rancid coconut oil may result in a slightly acidic environment.
• Methyl orange, in an acidic environment, will likely appear red or orangish.
6. Test Tube 6 (Rancid Coconut Oil and pH Paper):
• pH paper dipped into rancid coconut oil may show a lower pH reading compared to
fresh oil, indicating acidity or increased acidity.

Prepared by: Marlon P. Flores

NATSCI College Instructor
MCNUR2B: BIOCHEMISTRY LABORATORY
EXPERIMENT NO. 4 LIPIDS ANSWER KEY

FRESH COCONUT OIL RANCID COCONUT OIL

SAMPLE OBSERVATION SAMPLE OBSERVATION
No color change No color change
Phenolphthalein Phenolphthalein
(Colorless) (Colorless)
Yellow; possibly an Red or deep red-
Methyl Orange Methyl Orange
orange color orangish
Around 7 or slightly Below 5 or even
pH Paper pH Paper
lower than 7 lower

In summary, the pH indicators help reveal the acidity or alkalinity of the coconut oil samples. Fresh
coconut oil typically has a neutral to slightly acidic pH, while rancid coconut oil may become more
acidic due to the presence of free fatty acids resulting from oxidation. The specific colors observed in
the test tubes may vary based on the pH of the oils and the pH indicators used.

PRE-RESULT EXPLANATION BEFORE INSOLUBLE SOAP TEST:

A. PRINCIPLES OF SAPONIFICATION AND SALTING OUT PROCESS
The "salting out" process, also known as "salting out of soap," is used to precipitate soap from a
soap solution. In the procedure using a soap solution made from fresh coconut oil and 10% alcoholic
KOH, along with the addition of NaCl and distilled water, the characteristic result is the separation
and precipitation of soap. Here's how this process works:
• Soap Solution: The soap solution is initially prepared by saponifying the fresh coconut oil
with 10% alcoholic KOH. This results in the formation of soap molecules in the solution.
• Salting Out: When NaCl (table salt) is added to the soap solution and mixed, it reduces the
solubility of soap in the water-alcohol mixture. This is because the addition of a highly ionic
salt like NaCl disrupts the interactions between soap molecules and the solvent.
• Distilled Water: Distilled water is then added to the mixture. The presence of excess water
further reduces the solubility of soap, causing it to precipitate out of the solution. The soap
forms solid particles.
• Crushed Ice: Placing crushed ice above the watch glass helps to cool the mixture. Cooling
encourages the soap particles to aggregate and form a solid mass.
• Boiling Over a Water Bath: Boiling the mixture over a water bath serves to complete the
separation process. As the mixture heats up, the remaining soap in solution is removed
through the vapor phase, leaving behind the precipitated soap on the watch glass.
As the mixture is cooled and heated over a water bath, you would observe the soap molecules
coming out of solution and forming solid soap flakes or particles on the watch glass.
This solid soap can be scraped off and separated from the remaining liquid.
The key characteristic of this process is the separation and precipitation of soap from the soap
solution due to the addition of salt (NaCl) and the subsequent cooling and heating steps. This
method is often used in soap-making and purification processes.

B. FORMATION OF FATTY ACIDS

When 10% HCl (hydrochloric acid) solution is added to the soap solution, it results in the
neutralization of the soap. The acid reacts with the soap (which is a base) to form a salt and water.
The characteristic result of adding 10% HCl to the soap solution is:
• Neutralization: The HCl solution neutralizes the soap, resulting in the formation of a sodium
salt (sodium chloride, NaCl) and water (H2O). The soap molecules are broken down into their
component parts.
• Formation of Sodium Chloride: The primary product of this reaction is sodium chloride
(table salt), which is a water-soluble ionic compound.

Prepared by: Marlon P. Flores

NATSCI College Instructor
MCNUR2B: BIOCHEMISTRY LABORATORY
EXPERIMENT NO. 4 LIPIDS ANSWER KEY

• Separation: The sodium chloride formed in the reaction remains in solution, while any
remaining impurities or insoluble components of the soap solution may precipitate or
separate out.
The characteristic result is a change in the chemical composition of the soap solution, with the soap
being converted into a salt and water. The resulting solution may contain dissolved salt (NaCl) and
other substances, depending on the initial composition of the soap solution.

V. INSOLUBLE SOAP
A. Calcium Chloride and Magnesium Chloride Test
The characteristics of the results in the two test tubes with distilled water, calcium chloride
(CaCl2) solution, magnesium chloride (MgCl2) solution, and soap solution can be described as
follows:
1. Test Tube 1 (Distilled Water, 5% CaCl2 Solution, and Soap Solution):
• Calcium chloride (CaCl2) is a calcium salt that can react with soap, forming calcium
soap, which is often insoluble in water.
• When CaCl2 is added to the soap solution in distilled water, you may observe the
formation of a white or cloudy precipitate. This precipitate is calcium soap. Calcium
soap is not soluble in water and tends to separate from the aqueous solution.
2. Test Tube 2 (Distilled Water, 5% MgCl2 Solution, and Soap Solution):
• Magnesium chloride (MgCl2) is another salt that can react with soap, forming
magnesium soap, which is also typically insoluble in water.
• When MgCl2 is added to the soap solution in distilled water, you may observe the
formation of a white or cloudy precipitate. This precipitate is magnesium soap.
• Like calcium soap, magnesium soap is not soluble in water and tends to separate
from the aqueous solution.
In summary, the characteristic results in both test tubes involve the formation of a white or cloudy
precipitate due to the reaction of the calcium or magnesium salts with the soap solution.
The formation of calcium soap in Test Tube 1 and magnesium soap in Test Tube 2 is an indication of
the interaction between the metal ions and the soap molecules, leading to the precipitation of the
soap as an insoluble compound.
This is a common qualitative test for the presence of soap or the identification of hardness ions
(calcium and magnesium) in water.

SOLUTION OBSERVATION INFERENCES

5% CaCl2 White or cloudy precipitate Formation of calcium soap
(Insoluble)
5% MgCl2 White or cloudy precipitate Formation of magnesium
soap (Insoluble)

B. Liquid Detergent Test

When conducting the same test using liquid detergent instead of a soap solution, the
characteristics of the results may differ. Here's what you can expect:
1. Test Tube 1 (Distilled Water, 5% CaCl2 Solution, and Liquid Detergent):
• Liquid detergents typically contain surfactants, which are designed to be more
soluble in water and form stable micelles.
• When CaCl2is added to the liquid detergent in distilled water, you may not observe
the same type of precipitate formation as with soap. Calcium ions may not readily
react with the surfactants in the liquid detergent to form insoluble compounds.
• The solution may remain clear, and you might not see a visible precipitate.

Prepared by: Marlon P. Flores

NATSCI College Instructor
MCNUR2B: BIOCHEMISTRY LABORATORY
EXPERIMENT NO. 4 LIPIDS ANSWER KEY

2. Test Tube 2 (Distilled Water, 5% MgCl2 Solution, and Liquid Detergent):

• Similar to Test Tube 1, when MgCl2 is added to the liquid detergent in distilled water,
you may not observe the formation of a visible precipitate.
• The magnesium ions may not readily react with the surfactants in the liquid
detergent in a way that results in the formation of an insoluble compound.

SOLUTION OBSERVATION INFERENCES

5% CaCl2 No precipitate Surfactant is present
5% MgCl2 No precipitate Surfactant is present

In summary, the characteristic results when using liquid detergent in the test tubes are that you are
less likely to see the formation of visible precipitates as you would when using traditional soap.
Liquid detergents are formulated to remain soluble and form stable micelles in water, which might
prevent the same type of reaction with calcium and magnesium salts that is observed with soap.
Instead, you are more likely to have a clear or slightly cloudy solution in both test tubes.

Prepared by: Marlon P. Flores

NATSCI College Instructor
MCNUR2B: BIOCHEMISTRY LABORATORY
EXPERIMENT NO. 4 LIPIDS ANSWER KEY

POST-EXPERIMENT GUIDE QUESTION DISCUSSION

1) Why are fatty acids insoluble in water?
Fatty acids are insoluble in water because of their chemical nature and the differences in
polarity between water molecules and fatty acid molecules. This insolubility is primarily due
to the following reasons:
• Polar vs. Nonpolar: Fatty acids consist of a long hydrocarbon chain with a carboxyl
group (-COOH) at one end. The hydrocarbon tail is nonpolar, meaning it does not
have a significant electric charge distribution. In contrast, water (H 2O) is a highly
polar molecule with a partial positive charge on the hydrogen atoms and a partial
negative charge on the oxygen atom. Polar substances tend to dissolve in other
polar substances, while nonpolar substances dissolve in other nonpolar substances.
Fatty acids, being nonpolar, do not readily mix with polar water.
• Hydrophobic Nature: The nonpolar hydrocarbon tail of fatty acids is hydrophobic,
meaning it repels water. Water molecules are strongly attracted to each other due
to hydrogen bonding and do not interact favorably with nonpolar substances like
fatty acids.
• Lack of Hydrophilic Groups: While the carboxyl group at the end of a fatty acid
molecule is polar and hydrophilic (water-attracting), it is not sufficient to overcome
the hydrophobic nature of the hydrocarbon tail. The carboxyl group can form weak
interactions with water, but these interactions are not strong enough to make the
entire fatty acid molecule soluble.
Due to these factors, fatty acids, as well as fats and oils, do not mix or dissolve in water
and tend to separate, forming distinct layers or droplets. This property is essential for
many biological functions, such as the formation of cell membranes and energy storage,
as it allows for the compartmentalization of different cellular components.

2) Explain why cis-form is the predominant configuration of unsaturated fatty acids.

The cis-configuration is the predominant configuration of unsaturated fatty acids for several
reasons, primarily based on the steric hindrance and thermodynamic stability of this
arrangement:
• Steric Hindrance: In a cis-configuration, the two hydrogen atoms attached to the
carbon atoms on the same side of the double bond create steric hindrance. This
hindrance makes it energetically less favorable for the molecule to adopt the trans-
configuration, where the hydrogen atoms would be on opposite sides of the double
bond. In contrast, the trans-configuration would lead to greater steric hindrance,
which is why the cis-configuration is favored.
• Thermodynamic Stability: The cis-configuration is typically more thermodynamically
stable than the trans-configuration. The electrons in the pi bond of the double bond
experience less repulsion in the cis-configuration, leading to lower energy and
greater stability.
• Biological Synthesis: Enzymes involved in the biosynthesis of fatty acids, such as
desaturases, tend to produce cis-configured double bonds. This is because the cis-
configuration is more stable and requires less energy to form.
• Biological Function: Many biological processes and functions depend on the
presence of cis-unsaturated fatty acids in cell membranes and metabolic pathways.
These molecules provide fluidity and flexibility to cell membranes, which is essential
for the proper functioning of cells and organisms. The kinks or bends introduced by
the cis-double bonds enable membranes to remain fluid at lower temperatures.

Prepared by: Marlon P. Flores

NATSCI College Instructor
MCNUR2B: BIOCHEMISTRY LABORATORY
EXPERIMENT NO. 4 LIPIDS ANSWER KEY

3) Why is acrolein test a general test for fats?

The acrolein test is not a general test for fats but rather a specific test for a component of
fats and oils known as glycerol, which is a polyhydric alcohol and a major constituent of
triglycerides. Here's why the acrolein test is used to detect glycerol:
• Formation of Acrolein: In the acrolein test, glycerol is treated with a strong
dehydrating agent, often concentrated sulfuric acid (H2SO4). Under these conditions,
glycerol undergoes dehydration, which leads to the formation of acrolein, a
compound with a pungent, irritating odor. The reaction can be represented as
follows:

Glycerol → Acrolein + Water

• Acrolein's Characteristics: Acrolein's distinct and strong odor, along with its ability
to cause eye and respiratory irritation, makes it easily detectable and recognizable.
• Specificity: The acrolein test specifically detects glycerol because it involves the
unique dehydration of glycerol molecules. While glycerol is a component of fats and
oils (as the backbone of triglycerides), fats themselves are not directly converted
into acrolein by this test. Instead, it targets glycerol, which is common to various fat
and oil sources.

4) What type of rancidity occurs in vegetable shortenings and how can it be prevented?
The type of rancidity that occurs in vegetable shortenings is typically oxidative rancidity.
Oxidative rancidity is a chemical reaction that involves the oxidation of fats and oils. It is
characterized by the development of off-flavors and odors in the product, often described as
"stale" or "rancid." This process is accelerated by exposure to oxygen, heat, and light.
Preventing oxidative rancidity in vegetable shortenings involves several strategies:
• Antioxidants: Adding antioxidants to vegetable shortenings is a common method for
preventing oxidative rancidity. Antioxidants, such as tocopherols (vitamin E) and
BHA (butylated hydroxyanisole), help inhibit the oxidation of fats and oils by
scavenging free radicals and preventing the chain reaction of lipid oxidation.
• Packaging: Proper packaging is crucial to protect vegetable shortenings from
exposure to oxygen and light. Packaging should be airtight and light-resistant to
minimize the contact between the product and these rancidity-promoting factors.
• Storage Conditions: Vegetable shortenings should be stored in cool, dark, and dry
conditions to minimize temperature and light exposure. Reducing the storage
temperature can slow down the oxidation process.
• Inert Gas Packaging: Some commercial applications use inert gas packaging, where
the headspace in containers is filled with inert gases like nitrogen or carbon dioxide.
This displaces oxygen and reduces its contact with the product.
• Quality Ingredients: Using high-quality ingredients in the production of vegetable
shortenings, such as low-peroxide oils, can help extend the shelf life and reduce the
potential for oxidative rancidity.
• Hydrogenation: Partial hydrogenation of vegetable oils can create more stable fats
with higher resistance to oxidation, but this process has health implications and is
being used less frequently due to trans-fat concerns.
• Rapid Cooling: Rapidly cooling the vegetable shortenings after production and
packaging can help lock in freshness and reduce the oxidation rate.

Prepared by: Marlon P. Flores

NATSCI College Instructor
MCNUR2B: BIOCHEMISTRY LABORATORY
EXPERIMENT NO. 4 LIPIDS ANSWER KEY

5) Explain the cleansing action of soaps.

The cleansing action of soaps is based on their unique chemical properties as surfactants,
which stands for surface-active agents. Here's how soap achieves its cleansing action:
• Emulsification of Grease and Oil: Soaps have a polar (hydrophilic) head and a
nonpolar (hydrophobic) tail. This dual nature allows them to interact with both
water and oily substances. When soap is applied to a greasy or oily surface, the
hydrophobic tail of the soap molecule is attracted to the grease and oil, while the
hydrophilic head is attracted to water.
• Formation of Micelles: As you scrub or agitate the soapy solution, soap molecules
surround and encapsulate the oil or grease, forming structures called micelles. In a
micelle, the hydrophobic tails cluster together to protect themselves from water,
while the hydrophilic heads face outward, interacting with the water.
• Removal of Particles: The micelles act as tiny, spherical structures with the
hydrophobic tails trapping the oily or greasy substances inside. These micelles are
easily suspended in water because of their hydrophilic exteriors.
• Rinsing Away: When you rinse with water, the micelles disperse in the water,
carrying the trapped grease or oil with them. This process effectively removes the oil
or grease from the surface being cleaned.
• Dispersal of Particles: Additionally, the soap molecules help disperse dirt, dust, and
other particulate matter by surrounding these particles and preventing them from
re-depositing on the cleaned surface.

This mechanism allows soap to act as an effective cleanser, making it possible to remove oil,
grease, dirt, and various contaminants from surfaces, including skin, clothing, dishes, and
more. It is essential to rinse thoroughly to remove the micelles and the trapped particles
from the cleaned surface.

The cleansing action of soaps is a fundamental concept in daily hygiene and cleaning
processes. However, it's important to note that soap may not be as effective for removing
heavy or non-polar substances, such as some industrial oils and waxes, which may require
more specialized cleaning agents.

Prepared by: Marlon P. Flores

NATSCI College Instructor
MCNUR2B: BIOCHEMISTRY LABORATORY
EXPERIMENT NO. 4 LIPIDS ANSWER KEY

GOALS OF EACH TESTS:

I. SOLUBILITY TEST
Purpose: To assess the solubility characteristics of lipids and gain insights into their chemical
nature; also, to determine lipid solubility in various solutions to check whether it is soluble
or insoluble in polar or non-polar solutions.

II. TEST FOR UNSATURATION OF FATTY ACIDS

Purpose: To determine the degree of unsaturation in a given sample of fatty acids, whether
it is saturated or unsaturated.

III. ACROLEIN TEST

Purpose: To detect the presence of glycerol, a component of lipids, and indicate the possible
presence of lipids in a sample. It is not a comprehensive test for lipid identification but
serves as a preliminary step in lipid analysis by providing an initial clue about the presence of
glycerol-containing compounds like lipids.

IV. TEST FOR RANCIDITY

Purpose: To assess the quality and freshness of fats and oils; to detect if the lipid manifests a
sign of deterioration and spoilage, which can negatively impact the taste, odor, and safety
especially in food products.

V. INSOLUBLE SOAP TEST

Purpose: To assess the presence of certain insoluble soaps formed from the reaction
between fatty acids and alkali metals; to determine the effect of cations on soap solubility.

VI. SAPONIFICATION TEST

Purpose: To produce soap; saponification is a crucial chemical process in the production of
soap and detergents, and it is also widely used for cleaning, hygiene, and emulsification
purposes in various applications. It plays a significant role in daily life and various industries
where cleaning and emulsification are required.

VII. SALTING OUT TEST

Purpose: To improve the separation, purification, and concentration of soap or specific
components in a solution. This technique is employed in soap production, soap purification,
and related processes to enhance the quality and efficiency of soap manufacturing.

Prepared by: Marlon P. Flores

NATSCI College Instructor