FACULTY OF EDUCATION
UNIVERSITI TEKNOLOGI MARA
KAMPUS PUNCAK ALAM
SELANGOR
BACHELOR OF SCIENCE EDUCATION (HONS.) BIOLOGY
BIO462 - BIOCHEMISTRY
LAB REPORT 7 :
QUALITATIVE TEST OF LIPIDS
NAME STUDENT ID CLASS
MEERZA BINTI MD MAHAZAN 2023410814 ED2473A
NIK NUR ALYA AFIFAH BINTI NIK MOHD AMINUDIN 2023217856 ED2473A
NOOR ILYANA SHAFINA BINTI MOHAMAD JAMANI 2023479636 ED2473A
NOR SYAHIRAH NADIRAH BINTI FARA AZUAL 2023486204 ED2473A
SITI NORAZIAH BINTI AZIZ 2023690248 ED2473A
SITI NUR NABIHA BINTI ABDUL JABAR 2023406366 ED2473A
LECTURER:
PROFESOR DR FARIDA ZURAINA BINTI MOHD YUSOF
SUBMISSION DATE:
1 DECEMBER 2024
INTRODUCTION
Lipids are a diverse group of organic compounds that include fats, oils, hormones, and certain
components of cellular membranes. These compounds are characterized by their hydrophobic
nature, meaning they do not interact well with water. Instead, lipids are soluble in nonpolar
organic solvents and exhibit general insolubility in water. Examples of lipids include waxes,
sterols, fat-soluble vitamins (such as vitamins A, D, E, and K), monoglycerides, diglycerides,
triglycerides, phospholipids, and others. Lipids can be classified into four major groups: fatty
acids, glycerides, non-glyceride lipids, and complex lipids. They serve several vital functions in
the body, including energy storage, organ protection, and the production and regulation of
hormones. Additionally, lipids are considered esters due to the presence of carbonyl (-C=O) and
hydroxyl (-OH) functional groups in their molecular structure. This experiment focuses on
identifying and differentiating between various types of lipids through qualitative tests such as
solubility tests, spot tests, and acrolein tests. Observations will be made regarding the chemical
reactions that occur during these lipid tests to accurately interpret the types of lipids present.
OBJECTIVES
1. To identify and differentiate between various types of lipids, such as esters,
phospholipids, and cholesterol, through specific qualitative tests.
2. To observe the chemical reactions that occur during lipid tests and accurately interpret
the results to determine the presence and type of lipids in a sample.
TEST 1: SOLUBILITY TEST
For water and chloroform as solvent:
1. Three test tubes containing 5 mL of clarified butter, vegetable oil, and linseed oil were
prepared and labeled.
2. 5 mL of water was added into each test tube and the test tubes were shaken thoroughly
to mix the sample and the solvent.
3. Observation was made and recorded.
4. Step 1 until 3 were repeated but instead of water, 5 mL of chloroform was added
For ethanol as solvent:
1. Three test tubes containing 5 mL of clarified butter, vegetable oil, and linseed oil were
prepared and labeled.
2. 5 mL of ethanol was added to each test tube.
3. The test tubes were gently heated using a Bunsen burner and shaken thoroughly to mix
the sample and the solvent.
4. The solubility of the samples in ethanol was observed and recorded.
TEST 2: TRANSLUCENT SPOT TEST
PROCEDURE
1. A small drop of vegetable oil was placed on the filter paper.
2. Allowed the vegetable oil to soak into the filter paper for a few minutes. The vegetable oil
will spread and the effect on the paper’s appearance will become more obvious.
3. The filter paper was held close under the candle . The light interaction with the areas
where the vegetable oils have been applied was observed.
4. The observation was recorded.
5. Repeat steps 1-5 with clarified butter and linseed oil.
TEST 3: ACROLEIN TEST
PROCEDURE
1. Three test tubes containing clarified butter, vegetable oil, and linseed oil samples were
prepared.
2. A small amount of potassium hydrogen sulfate crystals was added to each tube.
3. The mixture in each tube was heated over low flame.
4. The smell of the gas, odor and any changes was observed and recorded.
TEST 4 : HUBL’S TEST
1. Two test tubes containing cottonseed oil and linseed oil were prepared.
2. 3 mL of chloroform were added to each test tube and the mixture was mixed.
3. 3 drops of Huble’s reagent were added into each test tube.
4. Observations were made and recorded.
RESULT :
SOLUBILITY TEST ACROLEIN
SAMPLE SPOT TEST TEST
WATER ETHANOL CHLOROFORM
Clarified Undissolve Dissolve Dissolve Translucent A pungent
butter completely completely and greasy irritating odour of
after heating without heating spot is present acrolein is
produced
Vegetable Undissolve Dissolve Dissolve Translucent A pungent
oil completely completely and greasy irritating odour of
after heating without heating spot is present acrolein is
produced
Linseed oil Undissolve Dissolve Dissolve Translucent A pungent
completely completely and greasy irritating odour of
after heating without heating spot is present acrolein is
produced
SAMPLE HUBL’S TEST
Cottonseed oil Violet color doesn’t fade away
Linseed oil Violet color fade away
SAMPLE STRUCTURE
Clarified butter
Linseed oil
Vegetable oil
Cottonseed oil
DISCUSSION
TEST 1
In this solubility test, three different samples of lipids which are clarified butter, vegetable
oil, and linseed oil, were tested for their solubility in different solvents. The solvents used are
water, ethanol, and chloroform. Water being a polar solvent, does not dissolve any of the lipids
samples because the lipids structure consists of non-polar fatty acid chains. As a result, the
polar water molecules and the non-polar fatty acid chains in the lipid sample do not interact. On
the other hand, all three lipid samples are dissolved in ethanol, an amphipathic molecule.
Ethanol contains both polar hydroxyl groups and non-polar ethyl groups. The non-polar ethyl
group is able to react with the non-polar fatty acid chain of the lipid sample. The lipid sample's
solubility in ethanol is enhanced by the heating that occurs during the experiment as ethanol is
less effective at dissolving purely non-polar substances compared to solvents like hexane or
chloroform. Last but not least, because chloroform is a nonpolar solvent, all three lipid samples
dissolve entirely without needing any heating. In conclusion, because of polarity differences, the
lipid sample dissolves in nonpolar solvents such as ethanol and chloroform but does not
dissolve in polar solvent, water.
TEST 2
The Translucent spot test is a qualitative test that is used to identify the presence of fats
or oils in a clarified butter, vegetable oil, and linseed oil by observing a translucent spot that
appears when the sample is rubbed onto filter paper. The presence of lipids like fats and oils is
presence if a greasy spot is visible that allows the light to pass through. Based on observation,
clarified butter, vegetable oil and linseed oil show a positive result in which there is a presence
of a greasy spot on the filter paper. Clarified butter produces a lighter spit compared to
vegetable and linseed oil due to the differences in fat composition. Lipids have higher boiling
points. At room temperature, lipids cannot absorb enough heat to evaporate unlike water or
other polar solvents. Lipids are hydrophobic molecules that are composed of non-polar
hydrocarbon chains. The inability to evaporate is because of the strong intermolecular forces,
such as Van der Waals interaction that contribute to their high boiling points.
TEST 3
Acrolein test is used to detect the presence of glycerol or fat by creating a pungent
imitating odor. Acrolein, which has a particular smell, is created when glycerol and potassium
hydrogen react. This test, which involves removing water molecules from glycerol by adding
potassium hydrogen sulfate crystals as a reagent, is the foundation of the dehydration reaction.
Glycerol and potassium hydrogen sulfate crystallize to generate acrolein, which is physically
distinguished by the discharge of a pungent smell. In this experiment, clarified butter, vegetable
oil and linseed oil after the mixture containing a small amount of potassium hydrogen sulfate
crystals, there was an irritating odor smell due to unsaturated acrolein. This, when in touch with
lipids, inherently smells foul. The smell of the acrolein indicates a positive test result.
TEST 4
The Hubl test is used to measure the degree of unsaturation in lipids by assessing their
oxidation rate. In this test, Hubl’s reagent acts as an oxidizing agent. When lipids undergo
oxidation, they reduce the reagent, causing the violet color to fade. The faster the color fades,
the more unsaturated a lipid is. In this experiment, linseed oil, which contains a higher
proportion of polyunsaturated fatty acids, oxidizes more quickly, reducing the reagent faster and
causing the violet color to fade more rapidly. In contrast, cottonseed oil, which contains more
saturated fatty acids, oxidizes more slowly, so the violet color fades less. This indicates that
cottonseed oil is more saturated than linseed oil.
CONCLUSION
The conducted experiments successfully characterized the properties of clarified butter,
vegetable oil, and linseed oil. Since they were soluble in ethanol and chloroform but insoluble in
water, solubility tests demonstrated their non-polar character. Their lipidic nature was validated
using the Translucent Spot Test, and glycerol was detected in every sample by the Acrolein
Test, a crucial component of triglycerides. Hence, Hubl's Test highlighted differences in the
degree of unsaturation, with linseed oil showing a higher degree than cottonseed oil. Overall,
this experiment successfully characterized the physical and chemical properties of the lipid
samples, providing valuable insights into their composition and potential applications.
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