Effects
of Different Composition of Ingredients on the Characteristics of Emulsion
Formulation
Aim
1. To
determine the effects of HLB value of surfactants used on the stability of
emulsion.
2. To
test the physical effect and stability of emulsion formulations having
different amount of emulsifying agents.
Introduction
Emulsion
is a preparation which involved two-phase system. This two-phase system is
thermodynamically non-favourable as it tends to separate into two distinct phases or layers if left over time. The preparation may contain two immiscible
liquids, where one of the liquids is dispersed evenly and homogenously in the
other liquid. There are two types of emulsion which are water-in-oil emulsion
and also oil-in-water emulsion. In water-in-oil emulsion, water or aqueous solution is dispersed in an oil solution. In contrast, oil-in-water emulsion consists of oil as the dispersed phase being dispersed in the continuous phase which is of aqueous or water solution.
Stability of an emulsion can be enhanced by adding emulsifying agent. This agent is made up of hydrophilic and lipophilic part. It generally acts by reducing the surface or interfacial tension. In order to produce a stable emulsion, the amount and the type of surfactants used must first be ensured by an analysis of the characteristics of surfactant. This may be done using HLB method. Usually the combination of two emulsifying agents is used to produce a stable emulsion. The HLB value can be determined using the formula stated below:
Stability of an emulsion can be enhanced by adding emulsifying agent. This agent is made up of hydrophilic and lipophilic part. It generally acts by reducing the surface or interfacial tension. In order to produce a stable emulsion, the amount and the type of surfactants used must first be ensured by an analysis of the characteristics of surfactant. This may be done using HLB method. Usually the combination of two emulsifying agents is used to produce a stable emulsion. The HLB value can be determined using the formula stated below:
Apparatus
i.
8
test tubes
ii.
A
50 mL measuring cylinder
iii.
2
sets of pipette pasture and droppers
iv.
Vortex
mixer
v.
Weighing
boat
vi.
1
set of pestle and mortar
vii.
Light
microscope
viii.
Slide
microscope
ix.
1
set of 5mL pipette and bulb
x.
A
50mL beaker
xi.
Coulter
Counter tools
xii.
Centrifuge
tools
xiii.
Viscometer
tools
xiv.
Water
bath (45 degree Celsius)
xv.
Refrigerator
(4 degree Celsius)
Materials
i.
Palm
oil
ii.
Arachis
oil
iii.
Olive
oil
iv.
Mineral
oil
v.
Distilled
water
vi.
Span
20
vii.
Tween
80
viii.
Sudan
III solution (0.5%)
ix.
ISOTON
III solution
Procedures
1. Each test tube was labelled and a
straight line (+-1cm) was drawn at the bottom of the test tube.
2. 4 mL of mineral oil was mixed
with 4 mL of distilled water in the test tube.
3. In the mixture of mineral oil and
distilled water, Span 20 and Tween 80 were added according to the amount stated
in Table 1. The test tube was sealed and centrifuged for 45 seconds using
Vortex mixer. Time taken for the interface between the separated phases to
reach the 1 cm line was recorded. HLB value for each sample was determined.
Test
tube no.
|
1
|
2
|
3
|
4
|
5
|
6
|
7
|
8
|
Span 20 (Drop)
|
15
|
12
|
12
|
6
|
6
|
3
|
0
|
0
|
Tween 80 (Drop)
|
3
|
6
|
9
|
9
|
15
|
18
|
15
|
0
|
Table
1
4. A few drops of Sudan III solution
were added to small amount (1 g) of emulsion formed in the weighing boat and
mixed evenly. The colour distribution in every sample was described and
compared. Small amount of sample was placed evenly on microscope slide and then
observed under the light microscope. The shape and size of globules formed in
every sample were drawn, described and compared
5. By using wet gum method, the Mineral Oil Emulsion (50g) was prepared using the formulation below.
5. By using wet gum method, the Mineral Oil Emulsion (50g) was prepared using the formulation below.
Mineral oil
|
Referred to next table
|
Acacia
|
6.25g
|
Syrup BP
|
5ml
|
Vanillin
|
2g
|
Alcohol
|
3ml
|
Distilled
water
|
qs 50ml
|
Emulsion
|
Group
|
Mineral oil (ml)
|
I
|
1,2,3
|
20
|
II
|
4,5,6
|
25
|
III
|
7,8,9
|
30
|
IV
|
-
|
35
|
Table III
6. 40g of the emulsion formed was placed in a
beaker and homogenized for 2 minutes using homogenizer machine.
7. 2g of
the sample before and after been homogenized was taken out and placed in the
weighing boats and labelled. Sudan III solution was dropped into the emulsion.
The texture, consistency, appearance of the oil and the color dispersion of the
sample were described and compared under the light microscope.
8. 15g
of the emulsion that have been homogenized was taken and the viscosity was
determined using the viscometer that has been calibrated using the “Spindle”
LV-4 type. The sample was then placed at 45oC (water bath) for about
30 minutes and at 4oC (freezer) for 30 minutes afterward. The sample
that has been exposed to temperature cycle was then allowed to reach room
temperature (10-15minutes). The viscosity was determined afterward.
9. 5g of
the emulsion that has been homogenized was placed into centrifuge tube and was
centrifuged in 4500rpm, for 10 minutes at 25oC. The separation
height produced was measured and the ratio of separation height was determined.
Results
Test
tube no.
|
1
|
2
|
3
|
4
|
5
|
6
|
7
|
8
|
Span
20 (Drop)
|
15
|
12
|
12
|
6
|
6
|
3
|
0
|
0
|
Tween
80 (Drop)
|
3
|
6
|
9
|
9
|
15
|
18
|
15
|
0
|
HLB
Value
|
9.7
|
10.7
|
11.3
|
12.4
|
13.2
|
14.1
|
15.0
|
0.0
|
Phase
separation time (min)
|
196.0
|
173.0
|
129.0
|
91.0
|
57.0
|
15.0
|
9.0
|
2.0
|
Stability
|
Stable
|
Stable
|
Stable
|
Slightly stable
|
Slightly stable
|
Not stable
|
Not stable
|
Not stable
|
Amount of mineral oil (ml)
|
Average Viscosity (cP)
(mean±SD)
|
Difference of viscosity (%)
| |
Before heat exposure
|
After heat exposure
| ||
20
|
18.93 ± 2.0422
|
19.30± 3.9800
|
1.95
|
25
|
205.10±98.72
|
460.34±131.08
|
124.45
|
30
|
520±70
|
2650±350
|
409.62
|
35
|
5350±334.17
|
6373.33±412.65
|
19.13
|
Discussion:
1. 1. What are the HLB values that will produce a
stable emulsion? Discuss.
Based on this experiment, we are able to detect a stable
emulsion based on the separation time. The longer the separation time, the more
stable will be the emulsion. Note that our group is doing the mineral oil, and
based on our results, the most optimum HLB value that produce a stable emulsion
are 9.7, 10.7 and 11.3 which results in the longest time separation that is
more than 120 minutes. When two or more emulsifiers are blended, the
resulting HLB of the blend is easily calculated by using the formula:
HLB value =
(quantity of surfactant 1)(HLB of surfactant
1)+(quantity of surfactant 2)(HLB of surfactant 2)
quantity of surfactant
1 + quantity of surfactant 2
Moreover we also have to take into an account about the emulsifying agents that added to form
an emulsion are Tween 80 and Span
20. The
emulsions are thermodynamically favourable due to the emulsifying agents form
around the globules to reduce energy of the system.Span 20 and
tween 80 has a HLB value of 8.6 and 15 respectively. Based in theory, the lower
the HLB value, the more oil soluble it is. Hence, we can say that span 20 is
more lipophilic while tween 80 is more hydrophilic. HLB of an emulsifier can
affect type of emulsion that been produced whether it is oil-in-water emulsion
or water-in-oil emulsion. Combination of different amount and different types
of emulsifying agents is able to produce a more stable emulsion.
Note that the role of the emulsifying agent is to stabilize the emulsion
through the formation of micelles. Micelles will function to keep the hydrophobic
drug particles or lipid globules in the core with tail pointing inward center
while the head will remain in aqueous phase. It will reduce the surface tension
of the interface between the oily and aqueous phase. As a result, it can
prolong the time of phase separation and phase separation occurs slowly. To be compared with constant condition
that are provided in tube 8 which there is no emulsifying agent being added,
the time separation are occur rapidly. This
high viscosity film is sufficiently flexible to permit distortion of the
droplets, resisted rupture, and gave a lower interfacial tension.
2. Compare the physical appearance of the mineral
oil emulsions produced and give comments.
What is Sudan III test? Compare the color dispersion in emulsions
produced and give comments.
Based on the experiment, our group
had conducted the experiment for 25ml of mineral oil. Note that before the
homogenization, the emulsion is unstable because the oily phase is immiscible with
aqueous phase. Under the microscope, the globules appear in different
sizes and larger. They are all unevenly dispersed. But after the homogenization
process, we can denote that the emulsion become more stable as it become less
oily and more viscosity. When it is observed under microscope, the globules are
in smaller size and more evenly distributed throughout the emulsion.
(25 mL of Mineral oil)
Description
|
|
Before
homogenization
|
Physical appearance: smooth and cloudy, not
consistent, very greasy
Globules:More globules, not uniform with
both big and small size
Color dispersion: unevenly dispersed, less
red spot
Viscosity: less viscous
|
After
homogenization
|
Physical appearance: smooth and milky,
consistent, less greasy
Globules:Less globules, uniform size
Color dispersion: evenly dispersed, more red
spot
Viscosity: more viscous
|
Sudan III is an oil soluble dye
which is red in color and usually being added to o/w emulsion, in background
will be colorless with colored spots if dispersed phase. Note that this test
may fail if ionic emulsifiers are used in the preparation. Sudan III testing
shows the shape and physical characteristic of oily phase in an emulsion by
staining the oil phase and observed under light microscope. We can also
determine the type of the emulsion whether water in oil (w/o) or oil in water
emulsion (o/w) by using this test. Red environment with globules indicate a w/o
emulsion whereas red globules in a clear field indicate an o/w emulsion.
3:
(a)
Graphs of emulsion viscosity before and after heat exposure against various amounts
of mineral oil added.
Emulsion is a mixture of two
liquids which are normally immiscible. It is classified into 2 types which are oil in water emulsion and water in oil
emulsion. In
this experiment, the emulsion formed is oil in water emulsion. It can be
determined through the usage of emulsifier which is acacia. Acacia is used to
produce oil in water emulsion.
The formulation of emulsion may
affect the physical properties and stability of an emulsion where one of them
is viscosity. According to the results, the emulsion viscosity increases with
the amount of mineral oil contained.The
results showed all the emulsion
are acceptable. The
emulsion viscosity is expected to augment with the increase of internal phase
content which
is mineral oil.
There are many factors that can
affect viscosity but only temperature is discussed here. Generally, emulsion viscosity will decrease
with the increase of temperature.
According to Kokal, this condition is due to the decrease in oil
viscosity. However, there is an exception where certain surfactants may cause
an opposite effect. For example, cationic surfactant will increase emulsion
viscosity with temperature due to the formation of wormlike micelles. During
high temperature, the micelles length will unusually increase and thus make the
emulsion more viscous. As for this experiment, acacia is a natural emulsifying
agent that usually form hydrated lyophilic colloids called as hydrocolloids. The
results showed did not follow the basic rule of temperature effect. Instead,
the viscosity increases with temperature. This may be explained by the nature
of acacia gum. Acacia gum will swell when temperature increases, the swelling
causes the emulsion to become more viscous.
(b) Graph above shows
that difference in viscosity versus different amount of mineral oil added. From
the graph it is shown that overall, the viscosity changes
before and after heat exposure increases and decreases again. In theory, increase in amount of mineral oil,
increase viscosity difference should be obtained. The changes of
viscosity refer to the emulsion stability. The larger the difference, the more
instable the emulsion will be. As the concentration of dispersed phase increases, so
does the apparent viscosity of the product. This means that as the amount of
oil globules increase in continuous phase, the viscosity of the emulsion will
increase. Since mineral oil is the dispersed phase, if dispersed phase increased,
viscosity of the emulsion should be increased. However, in the
laboratory experiment, the result deviates from the trend due to homogenization
error. On the other hand, the results can also be explained by the various
amount of mineral oil added. As mentioned earlier, the higher the internal
phase which is mineral oil, the more viscous it is. The effort
to change the viscosity should be decreasing from the first till the last
emulsion as the viscosity will keep increasing, thus the small viscosity
change. But as what
this graph obtained, the result is bias from what that is expected.
Experimental error might have occurred such as during the preparation of
emulsion, the amount of materials used was not in the exact proportion, for
example, excipients and active ingredients. Another error is of the viscometer,
continue using the machine without washing every time finish measuring
viscosity will lead to the inaccuracy of the data.
4. Plot graph of
phase separation ratio against different volume of Mineral Oil. Give your
comment.
SD =
SEPARATION HEIGHT
Height ratio = Separation phase
Emulsion phase
Mineral Oil (ml)
|
Group
|
Separation phase (mm)
|
Initial emulsion (mm)
|
Ratio of Separation
Phase
|
Average Ratio
(Average ± SD)
|
Emulsion I (20mL)
|
1
|
1.8
|
4.4
|
0.41
|
0.49
± 0.08
|
2
|
2.6
|
4.6
|
0.57
|
||
Emulsion II (25mL)
|
3
|
3.4
|
5.0
|
0.68
|
0.61
± 0.07
|
4
|
2.7
|
5
|
0.54
|
||
Emulsion III (30mL)
|
5
|
1.5
|
7
|
0.21
|
0.375
± 0.165
|
6
|
27
|
50
|
0.54
|
||
Emulsion IV (35mL)
|
7
|
12.6
|
43
|
0.29
|
0.295
± 0.005
|
8
|
14
|
46
|
0.30
|
Phase separation ratio indicate the stability of an
emulsion. If the emulsion is unstable emulsion and form two distinct phase, the
ratio of phase separation will be high. In this experiment, we used centrifuges
to accelerate phase separation processes in emulsion by enhancing the specific
gravity differences. The concept of the phase separation by centrifugal is
based on density difference of the oil and water phase in an emulsion. After
the process, phase separation will occur which is water phase and oil phase
will separate into two layers. Since the oil has lower density, it will become
upper layer while water is the bottom layer. There is no much difference
between densities of oil being used. Hence, type of oil did not give much
influence to the result. According to theory, as the amount of oil increase,
the separated phase ratio will increase. This is because the added amount of
oil in emulsion is beyond the oil amount at which a stable emulsion can be
formed. Phase separation will occur at a faster rate.
However, according to the graph above,
the phase separation ratio decrease when the amount of oil increases after 20
ml. The result didn’t follow the theory. This is because some errors occur
during the experiment. For example, present of contaminant in emulsion has
influence the accuracy of the result. Besides that, incorrect amount of oil
also affect the accuracy. Parallax error occurs when measuring the height of
separation phase. There is also possibility that some groups measure separation
phase by using height of water phase instead of oil phase.
5. Palm oil, arachis oil, olive oil and
mineral oil function as oil phase (internal phase) in the oil in water emulsion
(o/w emulsion). Acacia, Span 20 and Tween 80 are emulsifying agents used to emulsify two
immiscible liquid which are liquid and oil into a miscible form called
emulsion. The hydrophobic tails will be in contact with the oily phase while
the hydrophilic head group will be in contact with the aqueous phase. This
lower the surface tension of water molecule and provide an evenly mixing
between oil and water molecule. This makes the emulsion more stable. However,
it promotes the growth of microorganism, hence antimicrobial agents should be
added to prevent the growth of the microorganisms.
Alcohol is used as a preservative in this oil-in-water emulsion, as emulsion contains a high proportion of water present so it is very susceptible to microbial contamination. Besides that, syrup is to mask the unpleasant taste of the mineral oil and increase patient compliance as it contains a high concentration of sugar. It can also be used to increase the viscosity of the emulsion. However, the amount of syrup used should be controlled to prevent too viscous and form a layer on the side of the container.
Vanillin acts as flavouring agent which can increase the taste of emulsion to increase the compliance of patient as an emulsion always contains a taste not preferred by most of the patients.
Distilled water can function as aqueous phase (continuous phase) in oil-in-water emulsion.
Different type of oil will have different viscosity. The more viscous the oil , the more stable the emulsion. Different composition of oil and water will determine the type of emulsion either oil in water or water in oil emulsion. If there is too much oily phase in an o/w emulsion, the emulsion will become very unstable, and phase inversion will occur where it is converted into w/o emulsion. Hence, suitable emulsifying agents with suitable HLB value should be selected in order to produce a stable emulsion. Different proportion of emulsifying agents will give different stability and emulsifying effect. Sometimes, a combination of the surfactant can be used to improve the stability of the emulsion. Unsuitable surfactants will produces emulsions with different physical properties such as globule size, texture, consistency, oily phase dispersion, etc. These may affect the therapeutic effects of the emulsion. The use of different type of mineral oil will affect the physical characteristics and chemical stability of emulsion. For example, palm oil has anti-oxidant properties which increase the chemical stability of the emulsion. This type of emulsion will be less prone to oxidation than using other types of oil.
Conclusion :
In conclusion, HLB method is required to determine the stability of
emulsion. The most suitable HLB value of Mineral oil is range from 9.7 to 11.3.
For the emulsion
preparation, mineral oil is used as the oil phase as it is inert. Suitable
emulsifying agents with suitable HLB value should be chosen to produce a stable
emulsion. The higher the amount of mineral oil added, the higher the viscosity
of the emulsion formed.
REFERENCES
1. Aulton, M.E. Collet, D.M. Pharmaceutical Practice. Edinburgh: Churchill Livingstone.
2. Salager, J.L, Emulsion Properties and Related Know-how to Attain Them,
Pharmaceutical Emulsions and Suspensions, 2000, Marcel Dekker Inc.
3. Kalur, G. C, Frounfelker, B. D, Cipriano, B. H, Norman, A.L, Raghavan, S. R,
ViscosityIncrease with Temperature in Cationic Surfactant Solutions Due to the
Growth of Wormlike Micelles, 2005, American Chemical Society.
4. http://media.wiley.com/product_data/excerpt/3X/04701709/047017093X.pdf
5. http://www.rjpbcs.com/pdf/Old%20files/51.pdf
6. http://www.monzir-pal.net/Industrial/Characteristics%20of%20Surfactants%20and%20Emulsions.htm
7. Pharmaceutics, The Science of Dosage Form Design, Michael Aulton, 3rd Edition
