SCHOOL OF BIOSCIENCES
COVER SHEET FOR LABORATORY REPORT
This sheet must to be included with each laboratory report to be submitted. If you are submitting the
report on paper, please staple this sheet to the front of each report. If you are submitting the report
online, please ensure this cover sheet is included at the start of your report.
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 5 - Determination of Effectiveness of Various Emulsifying Agents in Dispersion
System
Module Lecturer: Dr Chan Sook Wah
Student’s Statement:
I have read and understood the TUC Regulations on cheating, plagiarism and collusion and state that this
piece of work is my own and does not contain any unacknowledged work from any other sources.
I authorise the University to test any work submitted by me, using text comparison software, for instances of
plagiarism. I understand this will involve the University or its contractor copying my work and storing it on a
database to be used in future to test work submitted by others.
Note: The attachment of this statement on any electronically submitted assignments will be deemed to have the same authority as a
signed statement.
Signed: Date: 2 July 2020
Received by: Date received from student:
Lab Report 5: Determination of Effectiveness of Various Emulsifying Agents in
Dispersion System
Objectives
• To determine the effectiveness of various substances as emulsifying agents.
Introduction
Emulsions are the mixture of two immiscible liquids, such as oil and water in which one of
these two liquids is dispersed in the other liquid. There are two phases in the system, which are
dispersed phase and continuous phase. The Dispersion phase is a liquid that is dispersed in the
other liquid which is referred to as a continuous phase. The most known emulsions are the
dispersions between lipophilic or oil phase and hydrophilic or watery phase where both can be
either the dispersed phase or the continuous phase. These basic types of emulsions are
commonly known as oil-in-water emulsions and water-in-oil emulsions. These emulsions are
thermodynamically unstable because of the interfacial tension between the immiscible liquids.
Thus, surface-active molecules, known as emulsifiers, are needed to help kinetically stabilize
the emulsion. Moreover, due to the difference in density between the two liquids, undesired
creaming or sedimentation of the dispersed droplets may occur. Therefore, the use of
stabilizers is significant to delay or prevent the physical instability of the emulsions. The
stabilizers are used in emulsions to increase the viscosity of the continuous phase. (Schubert, et
al., 2006)
Basically, the properties of emulsions are the microstructure of emulsion, the emulsifiers used,
and the continuous phase viscosity. Microstructure refers to the function of droplet size and the
distribution of droplet size. The stability of emulsions is highly affected by the droplet size.
Furthermore, it also influences the rheological characteristics, bioavailability or dose-response,
taste, and many more. The function of an emulsifier is to stabilize the emulsion by producing
droplets as fine as possible. (Schubert, et al., 2006)
The hydrophilic-lipophilic balance number (HLB number) of an emulsifier is a measure of its
affinity with oil or water. The properties of hydrophilic are controlled by the association of the
polar gathering of an emulsifier with water, and the lipophilic properties are dictated by the
collaboration of a nonpolar surfactant fragment with oil. HLB number has a significant
influence in the choosing of a certain type of emulsion. (Tereshchuk, et al., 2018)
A significant factor to decide the sort of emulsion being formed is the solvability or
dispersibility of an emulsifier in the fat or watery phase. As per the Bancroft rule, fat-soluble,
(or dispersible in fat) emulsifiers, the HLB estimations of which go from 2 to 6, form water-in-
oil emulsions. Water-soluble (or water dispersible) emulsifiers that have high HLB values –
from 11.0 or more can form oil-in-water emulsions. (Tereshchuk, et al., 2018).
Egg yolk forms stable emulsions at a low pH, this property is uncommon to most emulsifiers.
This makes egg yolk a suitable emulsifier in acidic systems (Magnusson and Nilsson, 2012).
Egg yolk contains a large amount of surface active components such as hydrophobic and
hygroscopic domains. Egg yolk can be further fractionated into plasma and granules that
contribute to its emulsifying properties. The plasma fraction contains low-density lipoprotein
and water-soluble plasma proteins. The granules fraction consist mainly of high-density
lipoproteins (Magnusson and Nilsson, 2012). The LDL proteins releases its inner lipid content
into the emulsion droplet, contributing to the emulsifying action The water soluble plasma
protein is known to provide stability against coalescence at pH 6.0 to pH 8.0. The HDL
provides stability which is suggested to be caused when there is enhanced interaction between
the individual proteins that forms a viscoelastic network at the interface (Magnusson and
Nilsson, 2012).
Lecithin is a general term for brownish yellow fatty substances that occur in plant or animal
tissues that are amphiphilic and can be blended with vegetable oil to make a liquid form. .It is
commonly used for oil-in-water emulsions because its hydrophilic lipophilic balance is in the
5-6 range (Hamilton, 2003). It can be used to emulsify emulsions from pH 3 to pH 9 that are in
organic or inorganic nature (Weete, Betageri and Griffith, 1994).
Sucrose esters are synthetically made from the esterification of sucrose and fatty acids. The
sucrose molecule is polar and is the hydrophilic end that forms hydrogen bonds with water
molecules, and the fatty acid chain is the hydrophobic end. It is flexible and creates oil-in-
water or water-in-oil emulsions depending on its HLB number. It has a wide range, from 1 to
16, for its hydrophilic lipophilic balance (Akoh, 1992). The lower the HLB number, there are
relatively fewer free hydroxyl groups and relatively higher fatty acid chains. The higher the
HLB number, there are relatively higher free hydroxyl groups and relatively fewer fatty acid
chains. Low HLB sucrose esters act as water-in-oil emulsifiers whereas higher HLB sucrose
esters act as oil-in-water emulsifiers (Nelen and Cooper, n.d.).
Apparatus and Instruments
• Electric homogenizer
• Light microscope
• Microscope slides
• Test tubes
Reagents and Materials
• Sudan red dye
• Egg yolk
• Lecithin
• Sucrose ester (Ryoto Sugar Ester S-170, HLB 1)
• Sucrose ester (Ryoto Sugar Ester S-170, HLB 15)
Samples
Canola oil
Procedure
1. Color oil with a small amount of the fat-soluble Sudan Red Dye.
2. Mix the emulsifier into the canola oil and water according the following table:
Table 1.0: Preparation of different oil/water (O/W) emulsions system
Emulsifier Oil (mL) Water Liquid for
(mL) Dissolving
Emulsifier
1. Control (no emulsifier) 10 40 -
2. Egg yolk - 0.5 g 10 40 Water
3. Liquid lecithin - 0.5 g 10 40 Oil
4. Sucrose ester (Ryoto Sugar Ester S- 10 40 Oil
170, HLB 1) - 0.5 g
5. Sucrose ester (Ryoto Sugar Ester S- 10 40 Water
170, HLB 15) - 0.5 g
3. Mix the mixture for 50 seconds using the electric homogenizer at 8000 rpm.
4. Pour into the test tubes to observe.
5. Observe the emulsions under the microscope to determine the effectiveness of various
emulsifying agents used in the preparation of O/W emulsion.
Results
Figure 1: Control (no emulsifier) under 400x magnification. Large fat globules formed,
separated from each other as there is no emulsion occur.
Figure 2: Egg yolk added as emulsifier under 400x magnification. Medium and small globules
formed and evenly distributed.
Figure 3: Lecithin added as emulsifier under 400x magnification. Large and medium globules
formed and not evenly distributed.
Figure 4: Sucrose ester (Ryoto Sugar Ester S-170, HLB 1) added as emulsifier under 400x
magnification. Large and medium globules formed and unevenly distributed.
Figure 5: Sucrose ester (Ryoto Sugar Ester S-170, HLB 15) added as emulsifier under 400x
magnification. Medium and small globules formed and evenly distributed.
Discussion
Understanding the HLB (Hydrophilic Lipophilic Balance) and the type of emulsion that differ
in dispersed phase and continuous phase are essential in order to choose the proper emulsifier.
Since the stability of emulsion is affected by the mean droplet diameter and droplet distribution
(Schubert, Engel and Kempa, 2006), the smaller and the more droplets of dispersed phase are
desired characteristics of stable emulsion. Thus, the larger interfacial layer between both
phases is where the emulsifier can work effectively to keep droplets from aggregating and
coalescing.
Control sample was made by mixing 10 mL canola oil and 40 mL water without an emulsifier
as shown in Figure 1. Under 400x magnification, large fat globules can be observed with the
separation between these two immiscible liquids clearly occurring. After the process of
homogenization, immediately flocculation and coalescence or the process of Ostwald ripening
was immediately taking place. Separation by the layer formation will happen in a short time.
Therefore, the oil in water (O/W) emulsion of control mixture is thermodynamically unstable.
As shown by the result of this experiment, both Figure 2 and Figure 3 have different results.
The addition of egg yolk as emulsifier as presented in Figure 2 resulted in medium and small
globules formed and evenly distributed meanwhile the Figure 3 which was the incorporation
of lecithin has large and medium globules and it did not distribute evenly. The difference of
the result was questioned since the type of emulsifier is the same which is lecithin.
Main reason is affected by the type of emulsion made in this experiment. Oil in water (o/w)
emulsion is the preferred emulsion type in this observation. Hence, when lecithin in egg yolk is
dissolved to water, the stabilization of emulsion can be formed since water plays a role as a
continuous phase. The aqueous chemical reaction influences the interface as a place where the
emulsifier works and eventually has an effect on emulsion stability. Thus, the small and
medium globules as well as even distribution can be observed in Figure 2 as comparable to
Figure 3.
In Figure 3, where the liquid lecithin is used, the emulsion is considered to be unstable
compared to the emulsion using egg yolk. It is because oil as a dispersed phase that gives
limited effect to the stability of emulsion will make the emulsion unstable when the emulsifier
(liquid lecithin) is dissolved to oil. Therefore, large and medium globules formed with uneven
distribution can be noted. Moreover, the usage of lecithin in this experiment is not proper since
lecithin will only work productively in water in oil emulsion where the lecithin should be
dissolved in oil as the continuous phase. A prediction can be made to result in more stable
emulsion that can be formed with small globules and evenly distributed droplets. In summary,
lecithin will work productively when dissolved in the correct phase and used in the correct
type of emulsion that should be in water in oil (w/o) emulsion.
Sucrose esters as one of emulsifying agents having a particular chemical structure that helps in
the emulsification process which needs hydrophilic sites and lipophilic sites. Sucrose esters are
synthesized by the esterification of fatty acids with sucrose, a polyhydric alcohol with eight
hydroxyl groups that has three primary and five secondary hydroxyls; the primary ones are
more reactive and more easily replaced with fatty acids (Anarjan and Tan, 2013). The
illustration of sucrose esters is shown in Figure 6. Due to this structure, sucrose esters are
suitable for large ranges of oil in water (O/W) and water in oil (W/O) emulsion depending on
the modification of HLB value. The higher the HLB value meaning the higher the
hydrophilicity that is more suitable for oil in water (O/W) emulsion. While lower HLB value
means higher lipophilicity will be appropriate for water in oil (W/O) emulsion (van Ingen,
2011).
Figure 6. Sucrose esters chemical structure (Healthcare, n.d.)
Similar mixture of canola oil and water was made with the addition of sucrose ester (Ryoto
Sugar Ester S-170). The emulsifier with HLB value equals 1 was dissolved in the canola oil,
while HLB value equals 15 of the same emulsifier was dissolved in water. Under 400x
magnification, Figure 4 showed large and medium globules diameter with uneven distribution
in the usage of sucrose esters HLB 1. Large and medium oil globules formed because the
emulsifier was dissolved into the oil which is the dispersed phase of Oil in Water (O/W)
emulsion. Dispersed phase has limited effects on the properties of the emulsion. Therefore,
emulsifiers can not work effectively. Besides, the low HLB value indicated that emulsifiers
will work better in water in oil (W/O) emulsion compared to oil in water (O/W) emulsion.
On the other hand, sucrose ester HLB 15 that dissolves in water has small and medium oil
globules dispersed in the emulsion system and even distribution can be noticed in Figure 5.
The medium to dissolve the emulsifier used is water that also serves as a continuous phase that
will directly affect the interface. Hence, the more stable emulsion can be produced
synergistically with the effectiveness of the emulsifier to reduce the surface tension at the
interfacial layer. High HLB value of sucrose ester, 15, was appropriate in the oil in water
(O//W) emulsion because of the hydrophilicity trait.
Conclusion
In conclusion, the emulsifier that most suitable for oil in water (O/W) emulsion are egg yolk
and sucrose ester (Ryoto Sugar Ester S170, HLB 15) whereas the most suitable for water in oil
(W/O) are lecithin and sucrose ester (Ryoto Sugar Ester S170, HLB 1). Oil in water (o/w)
emulsion is the preferred emulsion type in this observation. Thus, egg yolk and sucrose ester
(Ryoto Sugar Ester S170, HLB 15) are more stable than lecithin and sucrose ester (Ryoto
Sugar Ester S170, HLB 1) because the fat globules formed in medium and small size and
distributed evenly from the observation of microscope.
References
1. Akoh, C., 1992. Emulsification properties of polyesters and sucrose ester blends I:
Carbohydrate fatty acid polyesters. Journal of the American Oil Chemists Society, 69(1),
pp.9-13.
2. Anarjan, N. and Tan, C., 2013. Effects of Selected Polysorbate and Sucrose Ester
Emulsifiers on the Physicochemical Properties of Astaxanthin Nanodispersions.
Molecules, 18(1), pp.768-777.
3. Hamilton, R., 2003. Physical properties of fats, oils and emulsifiers. Edited by N
Widlak. American Oil Chemists' Society Press, Champaign, IL, 2000. 260 pp. ISBN 0-
935315-95-0. Journal of the Science of Food and Agriculture, 84(1), pp.112-112.
4. Healthcare, A., n.d. SUCROSE FATTY ACID ESTER "RYOTO™ SUGAR ESTER" |
Products | Mitsubishi Chemical Corporation. [online] Mitsubishi Chemical
Corporation. Available at: <https://www.m-
chemical.co.jp/en/products/departments/group/mfc/product/1201443_7739.html>
[Accessed 25 June 2020].
5. Magnusson, E. and Nilsson, L., 2012. Eggs: Nutrition, Consumption And Health.
Sweden: Nova Science Publishers, Inc, pp.2-3.
6. Nelen, B. and Cooper, J., n.d. Sucrose Esters. Emulsifiers in Food Technology, pp.131-
161.
7. Schubert, H., Engel, R. and Kempa, L., 2006. Principles of Structured Food Emulsions:
Novel formulations and trends. 13th World Congress of Food Science & Technology,.
8. Tereshchuk, L. V., Starovoytova, K. V. & Ivashina, O. A., 2018. Practical Aspects of
The Use of Emulsifiers in Manufacturing Emulsion Fat-and-Oil Products. Food and
Raw Materials, 6(1), pp. 30-39.
9. van Ingen, B., 2011. Sucrose Esters, Not A ‘Sweet’ Solution.. [ebook] Available at:
<http://www.flavours.asia -> white_paper_-_sucrose_esters_not_a_sweet_solution.pdf>
[Accessed 25 June 2020].
10. Weete, J., Betageri, S. and Griffith, G., 1994. Improvement of lecithin as an emulsifier
for water-in-oil emulsions by thermalization. Journal of the American Oil Chemists’
Society, 71(7), pp.731-737.
