SCHOOL OF BIOSCIENCES
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Name:
1. Cathryn Choe
2. Clarissha Vallerie Widjaja (Group Leader)
3. Farah Ula Nabilah
4. Fong Shi Hui
5. Helena Wiranata
Student ID:
1. 0338139
2. 0340155
3. 0340810
4. 0338278
5. 0338467
Module Code: FSC60104
Names of partners: -
Email (Individual/Group Leader): Contact No (Individual/Group Leader) :
1. [email protected] 1. +60166822693
2. [email protected] 2. +6281805882858
3. [email protected] 3. +601120968557
4. [email protected] 4. +60176200168
5. [email protected] 5. +601114248131
Title of experiment:
Practical 4 - Determination of Iodine Value and Peroxide Value of Cooking Oil
Module Lecturer: Dr Chan Sook Wah
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Signed: Date: 19 June 2020
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Lab Report 4: Determination of Iodine Value and Peroxide Value of Cooking Oils
Objectives
• To evaluate the quality of edible oils using iodine value and peroxide value
Introduction
Edible oils contain unsaturated fats and vitamin E which are primarily essential in human
diets and also serve as important sources for energy in the human body. They are also
commonly used for food production in the food industry and home cooking. (MacMahon,
2015)
All fats and oils are susceptible to oxidation during storage. Many factors affect the rate of
oxidation such as the type of fatty acid, moisture, the processing quality, storage temperature,
oxygen concentration, and light. Lipid oxidation affects the quality of the product by
producing rancid flavors and discoloration. It also fastens the degradation of the product,
hence leading to the decreasing of the nutritional quality and safety of the product which
impacts on the human health in harmful way. (Syed, 2016)
Lipid oxidation highly occured in edible oils that contain high amounts of unsaturated fatty
acids, particularly polyunsaturated fatty acids (PUFAs). The oxidation process starts with the
double bonds of fatty acids which yield to the formation of hydroperoxides. Hydroperoxides
do not significantly affect the sensory quality of the oil due to their tasteless and odorless
characteristic. Hence, the problem is in their instability which yields to the formation of
secondary oxidation products like ketones, lactones, hydrocarbons, alcohols, or others. Some
factors that also affect the oxidative stabilities are the presence of non triglyceride materials
such as natural antioxidants, free fatty acids, phospholipids, polymers, mono- and
diglycerides, as well as the position of fatty acids double bonds in the oil. Lipid oxidation is
more susceptible in the sn-2 position of PUFAs than in the sn-1 or sn-3 positions. Many
physical changes are resulted from the oxidative degradation pathway like increased
viscosity, changes in composition, and degradation products formation. (Syed, 2016)
Oils and fat quality can be assessed by conducting the oxidative stability determination like
the Rancimat Metrohm (Europe), the oxidative stability index (OSI) (USA), and some new
methods, OXITEST, which can directly assess the oxidation process without extracting fat
from the foodstuff, or Near Infrared Emission Spectroscopy (NIRES) which is done by
monitoring the intensity of a band at 2,900 nm and is used to know the induction time of
edible oils in accelerated oxidation test, which also links to hydroperoxides formation.
(Ciemniewska-Zytkiewicz, et al., 2014)
On the other hand, oxidative deterioration of oils is commonly measured by determining the
peroxide value. The peroxide value measures hydroperoxide concentration which correlates
with peroxides liberating iodine from potassium iodide, i.e.
ROOH + KI → ROH + KOH +I2
The measure of ROOH is then controlled by estimating the measure of iodine formed, which
is done by titration with sodium thiosulfate and utilizing a starch indicator,
I2 + starch + 2Na2S2O3 (blue) → 2NaI + starch + Na2S4O6
The measure of peroxides is determined back by the measure of sodium thiosulfate (Na2S4O6)
expended. It is shown as peroxide value (PV) in units of milli-equivalents (meq) peroxide per
1 kg of fat removed from the food. A general standard is that PV ought not be over 10–20
meq/kg fat to dodge the rancidity flavor. (Kong & Singh, 2011)
The degree of unsaturation in oil can be measured by determining iodine value. It has been
discovered that the C18:2 fatty acid is relatively lost, and the iodine value of oil is decreased
after heating treatment which is caused by the increase of intensive thermo-oxidative
transformations that occur as compared to heated oil that contains food. To determine the
iodine value, the sample is treated with an excess of solutions of iodine monochloride (ICl)
in glacial acetic acid. Unreacted iodine monochloride will react with potassium iodide and
convert it to iodine. The concentration of iodine is determined by titration with sodium
thiosulfate. The destruction of double bonds by oxidation, scission, and polymerization can
be linked to the decrease of iodine value. (He & Liu, 2019)
Procedure
Section 1: Determination of Iodine Value of Cooking Oils
Samples
Corn oil and palm oil
Sample Preparation
If necessary, melt the sample at 60 to 70 °C and thoroughly homogenised before taking a test
portion.
Procedure
1. Weigh 0.5 g sample accurately to the nearest 0.0001 g into a 500 mL conical flask.
Prepare the samples in duplicates.
2. Add 20 mL of cyclohexane to dissolve the fat. If necessary, warm the flask slightly to
facilitate dissolving of the fat.
3. Add exactly 25 mL of the Wijs solution, insert the stopper, shake gently and place the
bottle in the dark for 1 hour.
4. After standing, add 20 mL of the 15% KI solution and 100 mL of water.
5. Add 2-3 drops of starch indicator solution.
6. Titrate with 0.1N Na2S2O3 solution until the blue color just disappeared.
7. Carry out a blank test in duplicates simultaneously under the same conditions.
Calculations:
where,
N = the normality of sodium thiosulphate solution
Vs = the volume, in mL, of the sodium thiosulphate solution used for the
determination;
Vb = the volume, in mL, of the sodium thiosulphate solution used for the blank test;
W = the weight, in grams, of the test portion
Express the result to one decimal place.
Section 2: Determination of Peroxide Value of Cooking Oils
Samples
• Fresh corn oil (A)
• Fresh palm oil (B)
• A and B, exposed to strong sunlight for 2 weeks
• A and B, treated with saturated ethanolic copper sulphate (4%, v/v)
• A and B, treated with 200 ppm of tocopherol solution
Sample Preparation
All samples should be analysed as soon as possible, or should be kept in a cool, dark place
before analysis.
Procedure
1. Weigh to the nearest 0.1 mg, 5.00 0.05 g of the sample into the 250 mL flask. Prepare
the samples in duplicates.
2. Add 30 mL of the acetic acid-chloroform solution. Swirl the flask until the sample is
dissolved in the solution.
3. Add 0.5 mL of saturated KI with a graduated pipette. Swirl the solution for 1 minute
and then add 30 mL of distilled water. Add 2-3 drops of starch solution.
4. Titrate with 0.1N Na2S2O3 solution, adding it gradually and with constant and vigorous
shaking. Continue the titration, shaking the flask vigorously near the end point to
liberate all the iodine from the chloroform layer.
5. Add the sodium thiosulphate solution dropwise until the blue colour just disappears.
6. Carry out a blank test (in duplicates) in parallel with the determination. The blank
titration must not exceed 0.1 mL of the 0.1N sodium thiosulphate solution.
Calculations:
(Vs - Vb ) x N x 1000
Peroxide Value (meq. peroxide/kg fat) =
W
where,
N = the normality of sodium thiosulphate solution
Vs = the volume, in mL, of the sodium thiosulphate solution used
for the determination;
Vb = the volume, in mL, of the sodium thiosulphate solution used
for the blank test;
W = the weight, in grams, of the test portion
Express the result to one decimal place.
Results
Section 1
Table 1. Weight of sample and volume of 0.1N Na2S2O3solution used for titration
Sample Weight Average Volume of 0.1N Average volume of Iodine
(g) weight Na2S2O3solution 0.1N value
(g) used (mL) Na2S2O3solution
used (mL)
0.0000 47.4
0.0000 47.25 0.00
Blank
0.0000 47.1
0.5079 6.00
0.5116 5.60 103.31
Corn
0.5154 5.2
Oil
0.5428 17.70
0.5222 18.70 69.41
Palm
0.5016 19.70
Oil
12.69 × 0.1 × (47.25−5.60)
Iodine value of corn oil =
0.5116
= 103.31
12.69 × 0.1 × (47.25−18.70)
Iodine value of palm oil =
0.522
= 69.41
Section 2
Table 2. Weight and volume of 0.1N Na2S2O3solution used for titration
Sample Weight Average Volume of 0.1N Average volume Peroxide
(g) weight (g) Na2S2O3solution of 0.1N value (meq.
used (mL) Na2S2O3solution peroxide/kg
used (mL) fat)
0.0000 0.00
Blank 0.0000 0.00 0.00
0.0000 0.00
5.0000 1.20
Fresh palm oil 5.0459 1.40 27.74
5.0918 1.60
Palm oil 5.0249 4.10
exposed to 5.0132 3.80 75.80
sunlight for
5.0018 3.50
two weeks
Palm oil 5.0068 2.60
treated with 5.0064 6.50 129.83
4% saturated
ethanolic 5.0059 3.90
copper
sulphate
Palm oil 5.0000 1.00
treated with 5.0005 1.10 22.00
200 ppm of
tocopherol 5.0009 1.20
solution
(1.40−0) X 0.1 X 1000
Peroxide value of fresh palm oil =
5.0459
= 27.74
Peroxide value of palm oil exposed to sunlight for two weeks = (3.80−0) X 0.1 X 10005
5.0132
=75.80
Peroxide value of palm oil treated with 4% saturated
(6.50−0) X 0.1 X 1000
ethanolic copper sulphate =
5.0064
= 129.83
Peroxide value of palm oil treated with 200 ppm of tocopherol solution
(1.10−0) X 0.1 X 1000
=
5.0005
= 22.00
Discussion
Section 1: Determination of Iodine Value of Cooking Oils
Understanding the purpose of Iodine value test is essential in determining different results from
corn oil and palm oil. The principle of the test is based on the degree of the unsaturation of fatty
acid components in oil. Unsaturation of fatty acid means the amount of double bonds contained
that will react with iodine compound from Potassium iodide (KI) solution (Iodine Value test with
CDR PalmOilTester palm oil analysis system, n.d.; Bioline International Official Site (site up-
dated regularly), n.d.; Estimation of Iodine Value of Fats and Oils (Theory) : Biochemistry Virtual
Lab I : Biotechnology and Biomedical Engineering : Amrita Vishwa Vidyapeetham Virtual Lab,
2020). Thus, it can be stated that the higher the iodine value, the higher the degree of unsaturated
fatty acids present in oil. The higher degree of unsaturation of oil results in more unstable
properties due to being more susceptible to oxidation. Oxidation of fat and oil affects several
damages, such as rancidity, flavor and nutritional quality loss.
Observation was made to compare control, corn oil, and palm oil in difference on Iodine Value
as presented in Table 1. Duplication was made in order to eliminate error bias in weight of oil
used and volume of 0.1N of Sodium thiosulfate. Compared to blank as control sample, outcome
of this experiment for Iodine Value of corn oil and palm oil subsequently are 103.31 and 69.41.
Therefore, degrees of unsaturation in corn oil are significantly much higher than palm oil. It can
be concluded that palm oil is more stable against oxidation than corn oil.
This statement can be supported by the observation made by Montoya et al. (2014), “Palm oil
contains approximately 50% saturated fatty acids, with 44% palmitic acid (C16:0), 5% stearic
acid (C18:0), and trace amounts of myristic acid (C14:0). The unsaturated fatty acids are
approximately 40% oleic acid (C18:1) and 10% polyunsaturated linoleic acid (C18:2) and
linolenic acid (C18:3).” While, total of saturated fatty acid in corn oil only about 13% (consist of
10% palmitic acid and 3% stearic acid) and 86% of unsaturated fatty acid (consist of
approximately 30% oleic (18:1) acid and 56% linoleic acid (18:2)) (Sahara and profile, 2011;
Baughman and Jamieson, 1921). With reference in these statements, Figure 1 shows the detail
component of fatty acids composition for both oils.
Figure 1. Comparison of Saturated and Unsaturated Fatty Acids for Corn Oil and Palm Oil
Thus, 86% unsaturated fatty acid in corn oil caused the oil to become less stable towards oxidation
compared to palm oil, which has about 50% unsaturated fatty acids. The ratio of palm oil and
corn oil oleic and linoleic acid also give particular effect to stability of oil. It is obvious that a
higher amount of linolenic acid, that has two double bonds in one chain, in corn oil is 56% will
cause rapid oxidation because the more double bonds being exposed compare to ~10% linoleic
acid in palm oil.
Section 2: Determination of Peroxide Value of Cooking Oils
The most common method for measuring oxidative decreases in oil is Peroxide Value (PV),
which is determined through the titration of iodine emitted by the lipid extract reaction to the
solution of potassium iodide. PV was correlated with the rancidity of the oils, the higher PV the
higher oxidation (Irwin and Hedges, 2004). The rates of oxidation are affected by factors such as
temperature, light, moists, metals and oxygen (R, SA and Olayanju, 2010). With the higher
oxidation level of oil, decline in oil quality will be occured.
As noted in Table 2, a blank sample was conducted as control. The comparison of fresh palm oil
with palm oil exposed to sunlight for two weeks shows different results, palm oil exposed to
sunlight has a higher PV value of 75.80 meq.peroxide/kg fat while fresh palm oil is 27.44 meq.
peroxide/kg fat so we can say that fresh palm oil that is kept in the right place for storage has
better quality.
Photooxidation is an oxidation process of oil that is accelerated by light, especially with
sensitizers, which absorb light energy very quickly in the singlet state. Singlet sensitizers may
return via light emission, inner conversion or device crossing into their ground state (Choe and
Min, 2006). In the initiation stage, hydrogen is abstracted from a free radical olefin (i.e.
unsaturated hydrocarbon) the hydrogen is separated from the carbon atom next to the double
bond, which is caused by light presence. The peroxides produced at the stage of propagation are
the alternation of primary products of oxidation, which does not have any importance to the
deterioration of flavor. This unstable primary oxidation outcome formed secondary oxidation
products that caused flavor deterioration which was called peroxide.
Figure 2. Photooxidation steps with hv as light sensitiser
Therefore, it can be concluded that the higher intensity of oxidative agents such as light will
increase the oxidation level. Higher peroxide value in palm oil exposed in sunlight for 2 weeks
was caused by the changes of oxidation’s primary products into secondary products, peroxide,
that impart rancid color and flavor.
It is observed that palm oil treated with saturated ethanolic copper sulphate has a relative higher
peroxide value. Indicating a rapid rate of oxidation has occurred. The results suggest that
ethanolic copper sulphate is an oxidizing agent, which is proven by the study conducted by
Algirdas and Griskonis (2019). Copper is an element that causes its reactants to lose electrons
which makes it an oxidizing agent, the reaction can be seen in Figure 3.
Figure 3. The progression of copper as an oxidizing agent.
Oxidation of edible oils can be influenced by an energy input such as metal, in this experiment,
it is copper. This experiment could have been done to mimic the property of oil as if it had not
undergone refinement yet as natural palm oil contains a relatively high quantity of transition metal
such as copper. The copper metal reacts directly with the palm oil to produce lipid alkyl radicals
which in turn produce reactive oxygen and hydroxyl radical which will act as a catalyst for oil
oxidation (Choe & B. Min, 2006).
In Table 2, palm oil treated with tocopherol solution recorded the lowest PV
22.00 meq.peroxide/kg fat, which is even better than the PV 27.74 meq.peroxide/kg fat of fresh
palm oil. This suggests that the palm oil has been treated with an anti-oxidant given that it has a
lower rate of PV than the palm oil that underwent auto oxidation. Tocopherol belongs to a group
of lipophilic compounds called vitamin E. They are considered powerful antioxidants because
they protect fatty acids in the lipid from lipid peroxidation. They do this by interacting with
polyunsaturated acyl groups and lipid peroxyl radicals and reactive oxygen radicals which
prevents them from reacting with fatty acid substrates (Fritsche, Wang & Jung, 2017).
Tocopherols have the ability to donate their phenolic hydrogen which is vital in the antioxidant
activity (Wagner, Kamal-Eldin & Elmadfa, 2004). The antioxidant delays the onset of oxidation
by restricting the formation of free radicals during the initiation phase. Naturally, vegetable oils
contain a relatively higher amount of tocopherols compared to animal fat. However, palm oil
contains one of the least amount of tocopherols,it has a rather higher concentration of tocotrienols
(Choe & B. Min, 2006). This suggests that palm oil could be treated with tocopherol to increase
shelf life due to its antioxidant properties.
Conclusion
In conclusion, the higher the iodine value, the higher the degree of unsaturated fatty acids present
in oil. The iodine value of corn oil is higher than palm oil because the degrees of unsaturated fatty
acids in corn oil are significantly much higher than palm oil. Therefore, palm oil is more stable
against oxidation than corn oil.
Consecutively, the higher peroxide value the higher oxidation level resulted in low oil quality.
Besides, the higher intensity of oxidative agents such as light and metal ions will increase the
oxidation. The rates of oxidation are affected by factors such as temperature, light, moists, metals
and oxygen. The palm oil exposed in sunlight for two weeks has a higher peroxide value which
was caused by the changes of oxidation’s primary products into secondary products, peroxide,
that impart rancid color and flavor. Moreover, ethanolic copper sulphate is an oxidizing agent,
therefore, the palm oil treated with saturated ethanolic copper sulphate has a relative highest
peroxide value. Lastly, tocopherol is an antioxidant, it lowers the rate of peroxide value of the
palm oil. Therefore, the peroxide value is lower than the palm oil that undergoes autoxidation.
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