TITLE
Effects of different
ingredients on the characteristics of emulsion formulation
DATE
13rd MAY 2015
OBJECTIVE
1) To determine the effects of surfactant’s HLB
value on the emulsion stability
2) To determine the
different physical effects and stability of the emulsion formulation by using
different emulsifying agent.
INTRODUCTION
Emulsion is a two-phase-system which is thermodynamically unstable. It
contains at least two immiscible liquids in which one of them
(internal/dispersed phase) is dispersed homogenously in the other liquid
(external/continuous phase). Emulsion can be classified into two: oil-in-water
emulsion (o/w) and water-in-oil emulsion (w/o). The emulsion can be stabilized
by adding the emulsifying agent. The emulsifying agent can be divided into 4
types: hydrophilic colloid, finely divided solid particle, and
surface-active-agent or surfactant.
The
HLB (hydrophilic-lipophilic balance) method is used to determine the quantity
and the type of surfactant required to prepare a stable emulsion. Each
surfactant is given a number in HLB scale, ranged from 1 (lipophilic) to 20
(hydrophilic). Normally, combination of two emulsifying agents is used to yield
a more stable emulsion preparation. The HLB value for the emulsifying agent
combination can be determined by using the formula:
HLB value
= (quantity of surfactant
1)(HLB of surfactant 1) + (quantity of surfactant 2)(HLB surfactant 2)
quantity of surfactant 1+ quantity of surfactant 2
MATERIAL AND APPARATUS
Apparatus: Ingredients:
8 test tubes palm oil
50 ml measuring cylinder arachis oil
2 sets of pipette pasture
and droppers olive oil
Vortex mixing device mineral oil
Weighing boat distilled water
Mortar and pestle Span 20
Light microscope Tween 80
Microscope
slides
Sudan III solution (0.5%)
1 set of pipette (5 ml) and
pipette-bulb ISOTON solution
III
50
PROCEDURES
1. Each of the test tubes was labeled and marked a
straight line 1cm from the base.
2. 4ml of the oil (table I) and 4ml of distilled water
were mixed in each test tube.
Table I
|
Group
|
Oil
|
|
1,5,9
|
Palm oil
|
|
2,6,10
|
Arachis oil
|
|
3,7,11
|
Olive oil
|
|
4,8,12
|
Mineral oil
|
3. Span 20 and tween 80 are added to each of the oil
and water mixture (refer table II). The test tubes were closed and mixed by
using Vortex mixing device for 45
second. The time needed for the separated phase to reach the 1cm line was
recorded. The HLB value for each sample was determined.
 |
| Tween 80 and Span 20 |
 |
| 8 tubes with mixing of oil, distilled water, Tween 80 and Span 20 |
 |
| Vortex Mixing Device |
For mineral oil:
|
Tube No
|
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
|
4.
A few drops of Sudan III solution were added to a little (1g) of emulsion
in the weighing boat and was flattened. The colour dispersity of the sample was recorded and compared with
other sample. A little sample was flattened on the microscope slide and
observed under the light microscope. The appearance and globule size formed for
each sample were drawed, explained and compared with others.
 |
| Sudan III solution |
5.
By using the wet gum method, a mineral oil emulsion was prepared using the
following formula:
Mineral oil
|
(refer table III)
|
|
Acacia
|
6.25g
|
|
Syrup
|
5ml
|
|
Vanilin
|
2g
|
|
Alcohol
|
3ml
|
|
Distilled water,qs
|
50ml
|
Table
III
|
Emulsion
|
Group
|
Mineral Oil (ml)
|
|
I
|
1, 5
|
20
|
|
II
|
2,6
|
25
|
|
III
|
3,7
|
30
|
6. 40g of
emulsion formed is put into a 50ml beaker and homogenization is done for 2
minutes using a homogenizer.
7. Some (2g) of emulsion formed is taken (before and after homogenization)
and put into a weighing boat and labelled. A few drops of Sudan III solution is
added and mixed. The texture, consistency, the degree of oily appearance and
the spreading of colour in the sample is stated and compared under the light microscope.
8.
The viscosity of the emulsion formed after homogenization (15g in 50ml
beaker) is determined using the viscometer that is calibrated with “Spindle”
type LV-4. The sample is then exposed to 45°C (water bath) for 30 minutes and
then to 4°C (refrigerator) for another 30 minutes. After the exposure to the
temperature cycle is finished and the emulsion had reached room temperature
(10-15 minutes), the viscosity of the emulsion is determined.
9.
5g of emulsion homogenised is put into a centrifugation tube and
centrifuged (4500 rpm, 10 minutes, 25°C). The height of the separation formed
is measured and the ratio of the height separation is determined.
DISCUSSION
1.What are the HLB values to form a stable emulsion?
Discuss.
|
Tube
no.
|
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
|
0.00
|
|
Average phase
separation time (min)
(Palm
oil)
|
16.28
|
40.55
|
67
|
25.7
|
38.58
|
39.69
|
22.1
|
4.68
|
|
Stability
(Palm
oil)
|
+++
|
++++++
|
++++++++
|
++++
|
+++++++
|
+++++
|
++
|
+
|
|
Average phase separation
time (min)
(Arachis
oil)
|
Did not
separate
|
Did
not seperate
|
Did
not seperate
|
49.5
|
53
|
56.5
|
32.0
|
22.5
|
|
Stability
(Arachis
oil)
|
++++++++
|
+++
|
++++++
|
+++++++
|
++++
|
++
|
+++++
|
+
|
|
Average phase
separation time (min)
(Olive
oil)
|
25.58
|
28.24
|
32.31
|
21.12
|
22.19
|
25.03
|
12
|
9.4
|
|
Stability
(Olive
oil)
|
+++++
|
++++++++
|
++
|
++++++
|
+++++++
|
++++
|
+++
|
+
|
|
Average phase
separation time (min)
(Mineral
oil)
|
69
|
65
|
60
|
59
|
27.5
|
14.5
|
7.85
|
0.84
|
|
Stability (Mineral oil)
|
++++++
|
++++++
|
++++++
|
+++
|
+++++
|
++++
|
++
|
+
|
HLB
value for Span 20 = 8.6
HLB
value for Tween 80 = 15.0
In
this experiment, the emulsifying agents that added to form an emulsion are
Tween 80 and Span 20. Span 20 and
tween 80 has a HLB value of 8.6 and 15 respectively. 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.
In this experiment, palm
oil, arachis oil, olive oil and mineral oil are used to produce an emulsion. Different
oil requires different optimum HLB value of emulsifying agents to produce a
stable emulsion. The longer the phase separation time, the more stable an
emulsion is. From this experiment, optimum HLB value for palm oil, arachis oil
and olive oil emulsion is 11.34, 9.67 and 10.73 respectively. Meanwhile,
mineral oil emulsion has 3 optimum HLB values which are 9.67, 10.73 and 11.34. Hence,
it can be said that the emulsion is slightly lipophilic. So, this emulsion is
water in oil emulsion. In these four type oil emulsion, tube 8 has shortest
separation time. This mean the emulsion produced is the least stable. HLB value
for tube 8 is 0.00. There is no emulsifying agent being used.
Emulsifying agent
is added into an emulsion to stabilise the emulsion through the formation of
micelles. Micelles will try 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. From tube 8, we can
clearly see that phase separation occur rapidly due to lack of emulsifying
agent. Emulsion with only one emulsifying agent such as test tube 7 is unstable
if compare with other tubes which has 2 emulsifying agent. Generally, usage of
two emulsifying agents will form a very more stable emulsion. Mixtures of
emulsifying agents are able to form a stable complex condensed film at the
oil/water interface. This high viscosity
film is sufficiently flexible to permit distortion of the droplets, resisted
rupture, and gave a lower interfacial tension.
However, there are some errors made
during the experiment which leads to the inaccuracy of some results. For example parallax error when measuring water, oil and
srfactant, errors when observing the phase separation in the test tube and time
taken for separation to occur.
2.
Compare the physical appearance of the mineral oil emulsions produced and give
comments. What is Sudan III test?
Compare the colour dispersion in emulsions produced and give comments.
For mineral oil:
|
|
Before homogenation
|
After homogenation
|
|
Texture
|
Smooth, cloudy,
non-homogenous
|
More smooth, milky,
homogenous
|
|
Consistency
|
Bad(less viscous)
|
Better(more viscous)
|
|
Physical oily degree
|
More oily
|
Less oily
|
|
Globule size
|
big
|
Small
|
|
Color dispersion
|
less uniform(less red
spot)
|
uniform(more red spot)
|
Sudan
stain test is a useful screening test for steatorrhea. Feces mixed with Sudan III or
Sudan IV stain are examined microscopically for detection of undigested (direct
test) or digested (indirect test) fats that appear as red-stained globules.
Sudan solution is used to
show the shape and physical characteristic of oily emulsion. It can show the
emulsion whether is oily-in-water emulsion or water-in-oil emulsion by
comparing the amount of the globules in red color and the colorless globules.
Sudan solution is red in color, and is dissolved in the oily phase in the
emulsion. This causes the oily globules red in color.
A w/o preparation will have a
greasy texture and often exhibits a higher apparent viscosity than o/w
emulsions. After homogenization, the w/o emulsion is converted to o/w emulsion.
The oily phase will be broken down into small globules. The texture of the
mineral oil occurs in a smoother and homogenous state. The color dispersion is
more uniform. This shows that the globule is dispersed equally. Besides, the
globule size is significantly smaller than that before homogenization. The
homogenization process makes the oily globules disperse evenly and becomes more
stable in the aqueous phase.
3. Plot and explain:
(1) Graphs of emulsion viscosity before and
after temperature cycle against various
amounts of mineral oil added.
GROUP 8 mineral oil = 30 ml
|
Reading
|
Viscosity (cP)
|
Mean
± SD
|
||
|
1
|
2
|
3
|
||
|
Before
temperature cycle
|
240
|
100
|
180
|
243.571± 57.35
|
|
After
temperature cycle
|
320
|
240
|
340
|
352.915± 43.20
|
|
Difference
(%)
|
30.983%
|
|||
.
|
Mineral oil (ml)
|
Viscosity average (cP)
x ± SD
|
Difference in viscosity (%)
|
|
|
Before
|
After
|
||
|
20
|
294.62± 241.19
|
36 ± 0
|
87.78%
|
|
25
|
453.95 ± 44.72
|
86.67 ± 1.25
|
80.91%
|
|
30
|
364.9 ± 121.33
|
324.2 ± 28.71
|
11.15%
|
From the graph, viscosity of different amount of
Mineral oil present in the emulsion has not much difference between the before
and after temperature cycle, the most significant difference are the viscosity
of 30ml mineral-oil-containing emulsion after the temperature cycle. Emulsion
with 20 ml of Turpentine oil has the lowest viscosity compared to the emulsion
with other volume of turpentine oil.
After
undergoing temperature cycle, the viscosity of all the emulsion increased.
Emulsion with 30 ml of mineral oil exerts the greatest increase in the
viscosity. By theory, an o/w emulsion stabilized by non-ionic emulgents will,
on heating, invert to form a w/o product. This is because as the temperature
increases, the HLB value of a non-ionic surfactant will decrease as it becomes
more hydrophobic. At the temperature at which the emulgent has equal
hydrophilic and hydrophobic tendencies (the phase inversion temperature), the
emulsion will invert. The higher viscosity of the emulsion after temperature
cycle shows that the emulsion formed becoming w/o emulsions. w/o emulsions
generally has the higher viscosity compared to o/w emulsions. Phase inversion
has occurred.
According to theory, supposed the viscosity
difference before and after temperature cycle is increased as the amount of oil
present in the emulsion 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. Therefore the graph obtained supposed to have an
increasing curve. But as what this graph obtained, the result is bias from what
that is expected. Experimental error might have occur such as during the
preparation of emulsion, the amount of materials used was not in the exact
proportion, eg: excipients and active ingredients. Another error is of the
viscometer, continue using the machine without washing everytime finish
measuring viscosity will lead to the inaccuracy of the data.
4. Plot graph of separated
phase ratio formed from the centrifugation process versus the different amount
of Mineral Oil. Explain.
|
Mineral oil (ml)
|
Separation phase ratio (x ± SD)
|
|
20
|
0.639 ± 0.020
|
|
25
|
0.714 ± 0.066
|
|
30
|
0.446 ± 0.218
|
Graph of separated phase ratio formed from the centrifugation process
versus the different amount of Mineral oil.
Based on the theory, the separation phase ratio
should be increasing with the increasing of the mineral oil contained in the
formulation. From the data obtained, the formulation with 20 ml and 25ml
mineral oil has the increasing value of separation phase ratio, the formulation
with highest value of separation phase ratio will produce the most unstable
emulsion. Thus to prepare an emulsion with maximal stability and homogenous
condition, the separation phase ratio shall as minimal as possible.
The
graph supposed to be curve increasing from 20 ml oil to the highest with 30 ml
oil usage. However, the separation phase ratio for 30ml is the lowest in this
experiment. Thus we can conclude that error has occurred in this part of experiment.
The
error occurred may be due to some careless mistake made when the experiment was conducted. This
maybe due to the homogenous process was not done properly. Other than that, the
emulsion volume of each test tube is maybe not equal when undergo the
centrifugation process. So it may affect the centrifugation process and lead to
the inaccuracy of the result. In addition, the quality of acacia is different
for each group, so it may cause the emulsion produced not equally.
5. What is the function of each ingredient used in
the emulsion preparation? How can the different amount of ingredients influence
the physical characteristics and the stability of the emulsion?
|
Ingredient
|
Function
|
|
Mineral oil
|
The oily phase in the o/w emulsion.
|
|
Acacia
|
Emulsifying agent which reduces the interfacial
tension and maintain the separation of the droplets in the dispersed phase.
|
|
Syrup
|
Increase the viscosity of the
emulsion and acts as sweetening agent.
|
|
Vanillin
|
As flavoring agent.
|
|
Alcohol
|
As antimicrobial agent.
|
|
Distilled water
|
As the aqueous phase in the o/w emulsion.
|
Amount of the mineral oil (oily
phase) and the distilled water (aqueous phase) used is important to determine
the type of emulsion formed, whether o/w or w/o emulsion. The volume of the
dispersed phase should not be more than the volume of the continuous phase. Or
else, phase inversion will occur.
Acacia which acts as the emulsifying
agent should be used in appropriate amount according to the HLB value. If the
amount used is less than which is required, the emulsion formed is not uniform
due to the large interfacial tension between the dispersed phase and the
continuous phase. Then the separation of phase will occur.
Syrup will affect
the viscosity of the emulsion formed as it is a viscous liquid. Suitable amount
of syrup should be used to give suitable viscosity to the emulsion formed.
Viscosity of the emulsion will affect the physical stability and the
rheological characteristic of the emulsion.
Alcohol which acts
as the antimicrobial agent should not be used in large amount to reduce
toxicity.
CONCLUSION
The
effect of HLB of surfactant on emulsion stability was determined and physical
effects. Appropriate amount emulsifying agent should be added to an emulsion to
achieve HLB value required by the oily phase in order to produce stabile
emulsion. The stability of the emulsion formulation due to usage of different
emulsifier agents was identified.
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,
Viscosity Increase 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. Pharmaceutics, The Science of Dosage Form Design, Michael Aulton, 3rd
Edition
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