Practical 1: Evaluation of the Effects of Certain Content on the Characteristics of an Emulsion Formulation
Introduction
Emulsion is a two different phases that
is not thermodynamically stable. It contains at least two immiscible liquids in
which one of them (disperse phase) homogenously disperse in another liquid
(continuous phase). It can be classified
as oil-in-water (o/w) emulsion or water-in-oil (w/o) emulsion. The droplet size
affects the distribution of dispersed phase of emulsion. For micro emulsion,
the droplets size range 0.01 to 0.1 mm. For macro emulsion, the droplets size
range approximately 5 mm. Emulsion can be stabilized by adding stabilizing
agents. Emulsifying agents can be divided into 3 types, which are hydrophilic
colloid, finely divided solid particles, and surface active agents or
surfactants.
HLB method (hydrophilic-lipophilic
balance) was used to determine the quantity and type of surfactants that will
needed to prepare a stable emulsion. Each of surfactants, will be given a
number in HLB scale which is from 1 (lipophilic) until 20 (hydrophlilic).
Normally, combination of two emulsifying agents used to produce an emulsion
that is more stable.
Objective
1. To
determine the effects of HLB surfactants on the stability of emulsion
2. To
study the effects of varying surfactant on the physical and stability of
emulsion
Apparatus
8 test tube, 1 50 mL measuring cylinder, 2 set of
Pasteur pipette and dropper, Vortex mixer, weighing boat, mortar and pestle,
light microscope, microscope slide, 5 mL pipette and bulb, 50 mL beaker,
Coulter counter machine, centrifuge tube, Viscometer, water bath ( 45oC),
refrigerator (4 0C).
Material
Palm oil, arachis oil, olive oil, mineral oil, Span
20, Tween 80, Sudan III solution (0.5%), ISOTON III solution.
Procedure
1.
Each test tubes were labelled and one
straight line in 1 cm was sketched at the bottom of test tubes.
2.
4 mL oil (Table 1) and 4 mL distilled water
were mixed in test tubes
Studied oil
|
|
1.
|
Palm oil
|
2.
|
Arachis oil
|
3.
|
Olive oil
|
4.
|
Mineral oil
|
3.
To the mixture of oil and water, Span 20
and Tween 80 (refer Table II) were dropped. Test tube was closed and mixed with
Vortex mixer in 45 seconds. Time taken for separation of 2 phases reaching 1 cm
line was recorded. HLB values were determined for each sample.
4.
A few drops of Sudan III solution was
dropped to few (1g) emulsion formed on weighing boat and distributed evenly.
Colour distribution in the sample were compared and elaborated. A few sample
was distributed evenly on the microscope slide and the sample was observed by
light microscope. The shape and size of globule formed were drew, elaborated
and compared.
5.
By using the wet gum method, formulation
for Mineral Oil Emulsion (50g) was prepared by using these formula.
6.
40 g emulsion formed was inserted into 50
mL beaker and the homogenous process was done in 2 minutes by using homogenous
instrument.
7.
A bit of emulsion formed (2 g) was taken to
the weighing boat and labelled. A drops of Sudan III solution was dropped and
distributed evenly. The texture, consistency, colour distribution and shape of
sample were elaborated and compared below light microscope.
8.
The viscosity of emulsion (15 g in 50 mL
beaker) formed after homogenous process was determined by using viscometer
instrument which is calibrated by using “Spindle” LV-4 type. The sample then
was exposed at temperature 45oC in 30 minutes and 4oC
temperature afterwards in 30 minutes. The viscosity of emulsion was determined
after exposure of temperature cycle and emulsion reached room temperature
(10-15 minutes).
9.
5 g emulsion which already homogenous was
inserted into centrifuge tube and centrifuged (4500 rpm, 10 minutes, 25oC).
the separation height formed was measured and the ratio of separation height
was determined.
Emulsion
|
Characteristics
|
I
|
Before
homogenization
· Naked
eyes: less milky, less viscous, more oily
· Microscopic: more
globules, red stain is unevenly dispersed
After
homogenization
· Naked
eyes: milky, more viscous, less oily
· Microscopic:
less and smaller globules, red stain evenly dispersed
|
II
|
Before
homogenization
· Naked
eyes: milky, oily, less viscous
· Microscopic:
many globules, red stain is unevenly dispersed
After
homogenization
· Naked
eyes: more viscous, less oily
· Microscopic:
less and smaller globules, red stain evenly dispersed
|
III
|
Before
homogenization
· Naked
eyes: cloudy, less viscous, more oily
· Microscopic:
more globules, red stain is unevenly dispersed
After
homogenization
· Naked
eyes: milky, more viscous, less oily
· Microscopic:
smaller globules, red stain evenly dispersed
|
IV
|
Before
homogenization
· Naked
eyes: cloudy, less viscous, more oily
· Microscopic:
not uniform in size, red stain is unevenly dispersed
After
homogenization
· Naked eyes: milky, more viscous, less
oily
· Microscopic:
smaller globules , uniform in size, red stain evenly dispersed
|
Discussion/Questions
1. What are the
HLB values that can produce a stable emulsion? Discuss.
Palm oil (Group 1 and
Group 5)
Tube number
|
1
|
2
|
3
|
4
|
5
|
6
|
7
|
8
|
Span 20 (drops)
|
15
|
12
|
12
|
6
|
6
|
3
|
0
|
0
|
Tween 80 (drops)
|
3
|
6
|
9
|
9
|
15
|
18
|
15
|
0
|
HLB value
|
9.67
|
10.73
|
11.34
|
12.44
|
13.17
|
14.09
|
15.00
|
0
|
Average phase separation time
(min)
|
74.5
|
55.0
|
67.0
|
29.0
|
39.0
|
37.0
|
16.0
|
8.0
|
Stability
|
No
|
No
|
No
|
No
|
No
|
No
|
No
|
No
|
Arachis Oil (Group 2
and Group 6)
Tube number
|
1
|
2
|
3
|
4
|
5
|
6
|
7
|
8
|
Span 20 (drops)
|
15
|
12
|
12
|
6
|
6
|
3
|
0
|
0
|
Tween 80 (drops)
|
3
|
6
|
9
|
9
|
15
|
18
|
15
|
0
|
HLB value
|
9.67
|
10.73
|
11.34
|
12.44
|
13.17
|
14.09
|
15
|
0
|
Average separation time (min)
|
No
separation
|
No
separation
|
No
separation
|
84
|
74
|
98
|
53
|
5
|
Stability
|
Yes
|
Yes
|
Yes
|
No
|
No
|
No
|
No
|
No
|
Olive oil (Group 3 and
Group 7)
Tube number
|
1
|
2
|
3
|
4
|
5
|
6
|
7
|
8
|
Span 20 (drops)
|
15
|
12
|
12
|
6
|
6
|
3
|
0
|
0
|
Tween 80 (drops)
|
3
|
6
|
9
|
9
|
15
|
18
|
15
|
0
|
HLB value
|
9.67
|
10.73
|
11.34
|
12.44
|
13.17
|
14.09
|
15.00
|
0
|
Average separation time (min)
|
107
|
117
|
76
|
38.39
|
112
|
98
|
20.35
|
11.31
|
Stability
|
No
|
No
|
No
|
No
|
No
|
No
|
No
|
No
|
Mineral oil (Group 4
and Group 8)
Tube number
|
1
|
2
|
3
|
4
|
5
|
6
|
7
|
8
|
Span 20 (drops)
|
15
|
12
|
12
|
6
|
6
|
3
|
0
|
0
|
Tween 80 (drops)
|
3
|
6
|
9
|
9
|
15
|
18
|
15
|
0
|
HLB value
|
9.67
|
10.73
|
11.34
|
12.44
|
13.17
|
14.09
|
15.00
|
0
|
Average separation time (min)
|
No
separation
|
No
separation
|
No
separation
|
91
|
57
|
15
|
9
|
0
|
Stability
|
Yes
|
Yes
|
Yes
|
No
|
No
|
No
|
No
|
No
|
Based on the results,
the HLB values which produce stable emulsions are 9.67, 10.73 and 11.34.
Arachis oil and mineral oil emulsions having these values do not separate by
1cm. Meanwhile, palm oil and olive oil emulsions having these values take
longer time to separate by 1cm compared to other HLB values.
Generally, emulsifiers
with HLB below 10 exhibits lipophilicity while above 10 exhibits
hydrophilicity.
2. Compare the
physical characteristic of the emulsion Mineral Oil formed and explain. What is
Sudan Test III? Compare the color dispersion in the emulsion formed and
explain.
For the experiment,
different amount of mineral oil was used which the Emulsion I contain 20 ml,
Emulsion II 25 ml, Emulsion III 20 ml and Emulsion IV 35 ml. before the
homogenation, the globule size formed is big and less uniform in dispersion of
colour. The emulsion looks a bit oily and has bad consistency. The smooth and
non-homogenous was produce for the texture of the emulsion. This shows that
water in oil emulsion has been produced .After homogenation, the phase
inversion occur where the oil in water emulsion is produced which it shows a
small globule size and colour dispersion is uniform. It has less oily compared
to before homogenation and more smooth and homogenous texture has been formed.
Sudan III test is used to
determine whether the type of emulsion is w/o or o/w emulsion. This can be done
by seeing the distribution of red colour of Sudan III solution in the emulsion
tested. When the dispersion of red colour is distributed evenly, it can be
conclude that the emulsion is oil in water phase. If the colour of red is not
distributed evenly, the water in oil emulsion is produce.
3. Plot a graph and give comments:
(1)
Graph of sample viscosity before and after
temperature cycle versus different amount of Mineral Oil.
Mineral
oil (ml)
|
Viscosity
average (cP) x ± SD
|
Difference
in viscosity (%)
|
|
Before
|
After
|
||
20
|
830±14.14
|
700±156.84
|
16.99%
|
25
|
1360±519.81
|
1180±61.64
|
14.17%
|
30
|
4998
 |
833±23.57
|
142.86%
|
35
|
3290±14.14
|
13010±1083.42
|
119.26%
|
Based on the graph of sample viscosity before and after temperature cycle versus different amount of olive oil plotted above, there are different of viscosity of the emulsions which formed by different volume of olive oil in the emulsion formulation.
The linear curve of before temperature cycle shows
that viscosity of emulsion which added with 30ml mineral oil is higher than the
emulsion which added with 20ml and also 25ml of mineral oil. The linear curve
of after temperature cycle also shows that the emulsion with higher amount of
mineral oil (30ml) is more viscous if compared to the emulsion of lower amount
of mineral oil (20ml and 25ml). It is also obvious that the viscosity average
all emulsions after the temperature cycle is higher than before the temperature
cycle. However, the viscosity average of the emulsion with 35ml mineral oil
increased significantly rather than emulsion with 20ml mineral oil which only
increased slightly. The emulsion with 35ml mineral oil also shows great
increase in viscosity if compared to emulsion with 20ml.
Theoretically, an oil in water emulsion which
stabilized by non-ionic emulsifying agents will undergo phase inversion and
invert to form a water in oil emulsion. This inversion occurs under a
well-defined condition, such as a change in emulsifier solubility which caused
by temperature effects/special interactions with other additives. During
temperature cycle, an increase in temperature decreases HLB value of non-ionic
surfactant as it becomes more hydrophobic. In this case, non-ionic surfactant
refers to olive oil which used in emulsion formulation.
As temperature increased, emulsions invert they
reach phase inversion temperature, which refer to temperature at which
emulsifying agent has equal hydrophilic and hydrophobic tendencies. An
increased temperature will cause a fall in apparent viscosity of continuous
phase and increased kinetic motion of disperse droplets and emulsifying agent
at oil in water interface. At low temperature (40c), the viscosity
of continuous phase will be increased and also the kinetic energy of the system
will be reduced. This subsequently reduces the rate of migration of the
globules in the disperse phase. Thus, the viscosity of the emulsion should be
increases after the temperature cycle. Hence, the higher viscosity average of
emulsion obtained after the temperature cycle can be explained by the formation
of water in oil emulsion after the inversion process. This is because the water
in oil emulsion often has higher viscosity rather than oil in water emulsion.
(2) Graph of
viscosity difference (%) versus different amount of oil.
The graph of percentage of viscosity difference of
different emulsions versus amount of oil plotted above show that the percentage
of viscosity difference before and after the temperature cycle increased with
the increased amount of mineral oil used in the formulation but then the
percentage decreased again. This does not obeys the theory that the viscosity
differences before and after temperature cycle is increased as the amount of
olive oil used is increased. 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 as the viscosity approaches that oil continuous phase. Therefore the
graph obtained supposed to have an increasing curve. The result above may due
to several errors that had occurred in the procedure.
4. Plot a graph of ratio of phase separation resulting
from centrifugation process against different mineral oil content. Describe.
Mineral Oil (ml)
|
Group
|
Separation phase(mm)
|
Initial emulsion (mm)
|
Ratio of Separation Phase
|
Average Ratio of
separation
(Average ± SD)
|
Emulsion I (20mL)
|
1
|
1.8
|
4.4
|
0.41
|
0.49 ± 0.08
|
5
|
2.6
|
4.6
|
0.57
|
||
Emulsion II
(25mL)
|
2
|
3.4
|
5.0
|
0.68
|
0.61 ± 0.07
|
6
|
2.7
|
5
|
0.54
|
||
Emulsion III
(30mL)
|
3
|
1.5
|
7
|
0.21
|
0.375 ± 0.165
|
7
|
2.7
|
50
|
0.54
|
||
Emulsion IV
(35mL)
|
4
|
12.6
|
43
|
0.295 ± 0.005
|
|
8
|
14
|
46
|
0.30
|
Stability of emulsion is indicated by the phase separation. Unstable emulsion will have high ratio of phase separation which will form non-homogenized phase. The phase separation is accelerated by centrifugation. It based on the different density of oil and water phase in emulsion. The low density material will be upward while the high density material at bottom layer.
Based on graph plotted, the separation phase ratio increase from 20 to
25 ml but decrease to 30 and 35 ml. This is uneven with the theory. According
to the theory, as the amount of oil increase,
the separated phase ratio will increase. This is because the added amount of
oily phase in emulsion has exceeded the oil amount at which stable emulsion is
formed. Therefore, separation will occur in a faster rate.
This maybe due to some errors happen when conducting the experiment. The
error can be the inaccuracy of measuring the amount of oil in making the
emulsion. Parallax error can occur while measuring the height of separation
phase. Besides that, the emulsion may be not homogenized well during
homogenisation. The quality of the emulsion also may affect the result because
good emulsion will have good stability. The emulsion must be in good form and
use the right method as instructed.
5.) What are the functions of each ingredients used in
the preparation of the emulsion? How does the usage of these ingredients affect
the physical characteristics and stability of the emulsion formulation?
INGREDIENT
|
FUNCTION
|
Mineral Oil
|
As a lubricating laxative
|
Acacia
|
Act as an emulsifying agent to reduce coalescense
|
Vanillin
|
As a flavorant
|
Alcohol
|
As a preservative to prevent the growth of
microorganism
|
Syrup
|
|
Distilled water
|
To make up to the required volume
|
The ingredients used in preparing emulsion will affect
the physical and chemical characteristics of the emulsion. Different types of
mineral oil used in the preparation of emulsion will give rise to a different
physical characteristics and chemical stability in an emulsion formulation.
Besides that, the different compositions of mineral oil and distilled water
used in the preparation of emulsion will affect the types of emulsion produced,
either oil in water (o/w) emulsion or water in oil (w/o) emulsion. The oil
content is set in range of 40% to 60% in order to produce a stable o/w
emulsion. The oil phase of an emulsion can provide emollient effects. If the
amount of distilled water used is in excess compared to the oil used, then an
oil in water emulsion will be produced. While water in oil emulsion is produced
if the amount of oil used is in excess. Hence, any change in the content of
water or oil will results in phase inversion.
Furthermore, the
content of syrup can affect the flow and physical characteristics of the
emulsion formed. The content of the syrup used must be controlled to prevent
any problems related to the emulsion's rheology properties. Physical
characteristics can also be affected in this case. Some of the oils used with different
colours will produce emulsion with different colours. In addition, palm oil has
antioxidant property and this can improve the stability of the emulsion formed.
The differences discussed above will produce an emulsion with different
physical characteristics and chemical stability.
The amount of alcohol used
as a preservative depends on the amount of water phase in the emulsion as water
is the best medium for microorganisms to grow. Distilled water on the other
hand, act as the aqueous phase (external phase or continue phase) in the oil in
water (o/w) emulsion.
Conclusion:
Span 20 in larger volume compare to Tween 80 gives a stable emulsion, this has proven the emulsion formed from four types of oil is oil in water emulsion. Palm oil, Arachis oil, and olive oil increase the viscosity of emulsion Arachis oil and olive oil give an unstable emulsion compare to mineral oil and palm oil as the change in viscosity is high.
Span 20 in larger volume compare to Tween 80 gives a stable emulsion, this has proven the emulsion formed from four types of oil is oil in water emulsion. Palm oil, Arachis oil, and olive oil increase the viscosity of emulsion Arachis oil and olive oil give an unstable emulsion compare to mineral oil and palm oil as the change in viscosity is high.
