ISSN-2321-0966 (Print) ISSN-2321-0974 (Online)

PHARMAGENE
Vol: 1 Issue:2

Research Article

www.genesisjournals.org

Formulation and Evaluation of Microemulsion Based Gel of Itraconazole

Parag Patel* , Mansi A Monpara, S N Mandal, Nikita Patel, Rajesh KS.
Department of Pharmaceutics, Parul Institute of Pharmacy, Li mda, Vadodara, Gujarat.

Abstract:

The aim of the present study was to investigate the potential of a microemulsion formulation for topical delivery of Itraconazole.
Using pseudo-ternary phase diagram microemulsion area was selected and according to that ratio of oil, surfactant and
cosurfactant microemulsion was optimized. The prepared microemulsion was subjected for various tests like size distribution
study, zeta potential, conductance, % transmittance and centrifugation. The optimized microemuls ion was then incorporated in
the 1% carbopol 934 gel in ratio of 1:1. The prepared microoemulsion based gel was subjected to various tests like extrudability,
spreadability, drug content, In-vitro drug release and rheological study and in-vitro antifungal activity against C.albicans.

Key-words: M icroemulsion, Itraconazole, Antifungal activity

Received on 15-05-2013 Modified on 15-06-2013 Accepted on 23-06-2013

INTRODUCTION
Fungal infection is a common infection which affects two-
third of population among the world. 1 M any antifungal
drugs are available for treatment of fungal infection like
M iconazole, Ketoconazole etc. Itraconazole is orally or
topically active antifungal agent with a broad spectrum of
activity. It is effective against several fungal strains such as
Candida albicans and Candida topicalis, which are
responsible for topical candidiasis 2. Itraconazole shows
very low water solubility and having log P value 6.5 which
indicate high permeability through membrane and it is
beneficial for topical delivery. Systemic side effects can be
overcome by its topical delivery. For topical delivery semi-
solid preparations are widely accepted over solid and liquid
dosage forms 3. M icroemulsions, which are optically
isotropic and thermodynamically stable systems of water,
oil, surfactant, and co-surfactant, have been studied as drug
delivery systems because of their capacity to solubilize
poorly water-soluble drugs as well as their enhancement of
topical and systemic availability. It helps to solubilize the
lipophilic drug moiety and it shows rapid and efficient
penetration to the skin. So it is beneficial for topical drug
*Address for correspondence:
M r. Parag P atel,
E-mail: p4pharmacy@yahoo.co.in
Contact: +919825566115
delivery.4 For topical delivery microemulsion is
incorporated in Carbool 934 gel base to prolong the local
contact to the skin.
MATERIALS AND METHODS
Materials: Itraconazole was kindly gifted by Alembic
pharma, Baroda. Tween 80 was purchased from Chemdyes
corporation and Ethanol from Lobachemie. All other
chemicals used were of AR grade.
Methods: Drug dissolved in the oleic acid. Then add
Tween 80 and Ethanol as surfactant and Co-surfactant
respectively. M ixed well all of them. Then water was added
drop by drop and mixed well on magnetic stirrer.
Transparent microemulsion was formed. Now for
preparation of Gel base, Carbopol 934 was suspended in
water and hydrated it for overnight. pH was adjusted
around 6 to 6.5 using Triethanolamine. For preparation of
M icroemulsion based gel; M icroemulsion and 1% Carbopol
934 gel were mixed in the ratio of 1:1.
S olubility of itraconazole 5
Solubility of itraconazole was determined in various oils
like castor oil, light liquid paraffin, capmul M CM , oleic

32
Microemulsion Based Gel of Itraconazole
acid,Iso propyl myristate, coconut oil, surfactants like
tween 60, tween 80, tween 40 and cosurfactants like
Isobutyl alcohol, ethanol, acconan and Isopropyl alcohol.
Excess of Itraconazole was added to 5 ml each, of oils,
surfactants, and co-surfactants in screw capped tubes and
shaken on orbital flask shaker at 100 RPM for 48 hours at
ambient temperature. The suspension was centrifuged at
5000 RPM and clear supernatant liquid was decanted and
filtered through Whatmann filter paper. The solubility of
Itraconazole was estimated by UV-visible
spectrophotometer.
Pseudo-ternary phase diagram5
The microemulsion region was determined by constructing
pseudo-ternary phase diagrams which are prepared by
using CHEM IX software. Water titration method was used
for its determination. These diagrams help to determine oil,
surfactant and co-surfactant ratio. Different mixtures of
surfactant to co-surfactants were prepared and the ratios
were fixed to 2:1, 3:1 and 4:1. These mixtures (S/CoS)
were mixed with oil phase to give weight ratio of 4.5:0.5,
4:1, 3.5:1.5, 3:2, 2.5:2.5, 2:3, 1.5:3.5, 1:4 and 0.5:4.5, water
was added drop by drop and stirred using magnetic stirrer
until homogeneous dispersion or solution was obtained.
The end point of the titration was the point in which the
solution becomes cloudy or turbid. The quantity of water
required to make the mixture turbid was noted.
Characterization of Microemulsion
Droplet size measurements: Size analysis of
microemulsion was carried out by dynamic light scattering
with zetasizer (M alvern instruments ltd., M alvern, U.K).
The polydispersity index of the formulation was
determined by the same instrument.
Zeta potential measurements: Zeta potential for
microemulsion was determined using zetasizer (M alvern
instrument ltd., UK).
Dilution test: It is confirmatory test of microemulsion to
know which type of microemulsion was formed. The
prepared optimized microemulsion was diluted with water
(as external phase).
Conductivity measurement6: The measurement of
electrical conductivity gives the quantitative idea of the
solubilization of water phase in the selected mixture
containing oil phase, surfactant and cosurfactant. It also
gives the idea about the types of microemulsion. The oil,
surfactant and co surfactant concentration is selected as per
optimized formulation. Then the water phase was added
drop wise to the mixture of the oil and amphiphiles and
electrical conductivity of formulated samples was measured
using a conductometer at ambient temperature.
Transmission electron microscopy: Oil globules of
M icroemulsion were visualized using TEM , Phillips
PHARMAGENE Vol: 1, Issue: 2
Technai-20 electron microscope (Philips, Holland) with an
accelerated voltage of 20 - 200 kV. The samples were
negatively stained with a 1% aqueous solution of
phosphotungstic acid (PTA). M icroemulsion was dried on a
carbon coated copper grid. After drying, the samples were
viewed under microscope 80 kV.
Characterization of microemulsion based gel
Physical appearance: The prepared Itraconazole gellified
microemulsion formulations were inspected for their color,
homogeneity, consistency and pH. The pH values of 1%
aqueous solutions of the prepared Gellified microemulsion
were measured by a pH meter.
S preadability measurement7: To determine the
spredability of M icroemulsion based gel, 0.5 g of gel was
placed within a circle of 1 cm diameter pre-marked on a
glass plate, over which a second glass plate was placed. A
weight of 500 g was allowed to rest on the upper glass plate
for 5 min. The increase in the diameter due to gel spreading
was noted.
Rheological study: The viscosity of M icroemulsion based
gel formulation was determined at 37ºC using a brook field
viscometer (Brookfield DV‐E viscometer). ). 62 no spindle
was used and rpm set at 12.
Drug content determination: Itraconazole content in
microemulsion based gel was measured by dissolving
known quantity of microemulsion based gel in solvent
(methanol) by Sonication. Absorbance was measured after
suitable dilution at 280 nm using UV/VIS
spectrophotometer (UV‐1700 CE, Shimadzu Corporation,
Japan).
Extru dability (Tube Test)8: It is a usual empirical test to
measure the force required to extrude the material from
tube. The method adopted for evaluating micoemulsion
based gel formulation for extrudability is based upon the
quantity in percentage of gel and gel extruded from
aluminum collapsible tube on application of weight in
grams required to extrude at least 0.5 cm ribbon of emulgel
in 10 seconds. M ore quantity extruded better is
extrudability.
In Vitro release study: Franz diffusion cell (with effective
diffusion area 3.14 cm2 and 15.5 ml cell volume) was used
for the drug release studies. M icroemulsion based gel (1 g)
was applied onto the surface of egg membrane evenly. The
egg membrane was clamped between the donor and the
receptor chamber of diffusion cell. The receptor chamber
was filled with freshly prepared PBS (pH 5.5) solution. The
receptor chamber was stirred by magnetic stirrer. The
samples collected at suitable time interval. Samples were
analyzed for drug content by UV visible spectrophotometer
at 280 nm after appropriate dilutions. The cumulative
amount of drug released across the egg membrane was
determined as a function of time.
33
Microemulsion Based Gel of Itraconazole

Antifungal activity:9 Antifungal activity of formulation
was checked by cup-plate method. A definite volume of the
Candida albicans suspension (inoculum) was poured into
the sterilized Sabouraud dextrose agar media (cooled at
40ºC) and mixed thoroughly. About 20 ml of this
suspension was poured aseptically in the petri plates and
kept till the solidification. The surface of agar plates was
pierced using a sterile cork borer. The prepared wells were
filled with equal volume of itraconazole containing
microemulsion, itraconazole containing microemulsion
based gel, microemulsion without itraconazole and. After
incubation for 18 to 24 hours at 25°C, the fungal growth
was observed and the zone of inhibitions were measured
using antibiotic zone reader.
RES ULT AND DIS CUSS ION
Excipient selection: All selected oils, surfactants and
cosurfactants followed under GRAS (Generally Regarded
As Safe). Selection of oil based upon solubility of
itraconazole in oil. Itraconazole shows better solubility in
oleic acid so this oil keeps itraconazole in solubilized form.
Safety is a major determining factor in choosing a
surfactant, as a large amount of surfactants may cause skin
irritation. Non-ionic surfactants are less toxic than ionic
surfactants. High HLB value surfactants form stable o/w
microemulsion. So tween 80 was selected as surfactant
which having HLB value 15. M oreover for better stability
of microemulsion secondary stabilizer added called co-
surfactant. It fills gap between surfactant molecule and
provide flexibility to the film. The co-surfactant selected
for the study was ethanol which has an HLB value of 4.2.
The suitable combination of high and low HLB surfactant
and cosurfatant shows better stability of microemulsion.
Carbopol 934 is good gelling agent and shows good gelling
property after hydration. Carbopol 934 form gel at neutral
pH. So to neutralize negative charge of carboxylic acid,
Triethaolamine added.
Size measurement and Zeta potential: Globule size of
optimized microemulsion was found to be 142.2 nm. PDI is
a measure of particle homogeneity and it varies from 0.0 to
1.0. If PDI value closer to 0.0 which indicates narrow size
distribution of the formulation. PDI of optimized
microemulsion was found to be 0.422 which indicates
prepared microemulsion is monodisperse which remains
stable and will not convert to the macroemulsion. Results
of globule size and PDI are shown in figure 2.
The magnitude of the zeta potential gives an indication of
the potential stability of the colloidal system. If all the
particles in suspension have a large negative or positive
zeta potential then they will tend to repel each other and
there will be no tendency for the particles to come together.
However, if the particles have low zeta potential values
then particles coming together and flocculating. The
general dividing line between stable and unstable
suspensions is generally taken at either +30 or -30 mV.
Particles with zeta potentials more positive than +30 mV or
more negative than -30 mV are normally considered stable.
There will be no force to prevent the zeta potential was
found to be -26.7 which shows good stability of
microemulsion. Result of zeta potential is given in figure 3.

Pseudo-ternary phase diagram:

Based on solubility study oleic acid selected as oil, tween
80 selected as surfactant and Ethanol selected as co-
surfactant. In pseudo-ternary phase diagram one axis
represent oil phase, other rep resent the aqueous phase and
third represent fixed weight ratio of surfactant and co-
surfactant. Here coloured area in the figure 1 refers to the
microemulsion region while the outside area shows
multiphase turbid regions. From pseudo-ternary phase
diagram of different ratio of Tween 80 & ethanol it was
found that best microemulsion zone was found for ratio 4:1.
So this oil, surfactant and co-surfactant ratio was used in
preparation of microemulsion. These phase diagrams are
given in figure 1.
Figure 1: Pseudo-ternary phase diagrams

34
PHARMAGENE Vol: 1, Issue: 2
Microemulsion Based Gel of Itraconazole

Dilution test: Upon dilution with water no phase

separation was observed. So, it proved that prepared
microemulsion is o/w type.

Conductivity measurement: This is one type of

confirmatory test of microemulsion to know the type of
microemulsion formed. When up to 5% of water added
shows no significant change in the conductivity because oil
concentration is more than water concentration so
resistance is more. When 5% to 45% water added
conductivity increase significantly shows bicontinuous
region of microemulsion but above 45% of water no Figure 5 : In-vitro release profile of Itraconazole
significant change in conductivity. Conductivity remain formulation
constsnt upon further addition of water which shows that
o/w type microemulsion formed. This is shown in figure 4. TEM analysis: TEM image is indicative of globule size of
M E. Figure 20 indicates that oil globules of optimized
microemulsion were spherical in shape and size range in
between 30-50 nm. TEM image of optimized
microemulsion is shown in figure 5.

Physical appearance: The prepared Itraconazole

microemulsion based gel formulation was yellow viscous
clear gel preparation with a smooth and homogeneous
appearance. The pH values of prepared formulation
ranged from 6.5 to 6.8, which are acceptable because adult
human skin pH is 5.5-6.4 so skin irritation can be avoided.

Refractive index: Refractive index of optimized

Figure 2: Size measurement
microemulsion based gel was found to be 1.35, measured
by Abbe’s refractometer which was same as refractive
index of water so it indicated that prepared gel was
isotropic in nature.

S preadability measurement: For topical preparation

spreadability is one of the important parameter.The
spreadability is important for ease of application and better
patient compliance. Prepared M E based gel shows increase
in diameter during spreadability measurement was 6.2 cm
which showed good spreadability of the formulation.

Rheological S tudy: Viscosity of final formulation was

Figure 3: Zeta potential measurement found to be 2.6±0.8 cP which indicates that formulation is
good enough to adhere to the skin so ultimately increase the
local contact for longer time, penetration of drug across the
skin and systemic absorption of drug can be avoided.

Drug Content Determination:Drug content of prepared

microemulsion based gel was found to be 93.34% which
indicates better drug loading capacity of the formulation
and required dose of drug was available for the
pharmacological action.

Extru dability measurement: It is important test for

Figure 4: Conductivity measurement semisolid preparation. Pressure required for extrude the

35
PHARMAGENE Vol: 1, Issue: 2
Microemulsion Based Gel of Itraconazole
more quantity of semisolid preparation is depend on the
viscosity and consistency of the formulation. M ore quantity
of microemulsion based gel extrude at little applied
pressure on tube which shows better patient compliance.
In-Vitro Drug Release Study: In-vitro drug release is
important tool to predict that how much amount of the drug
was available at site of application. Final formulation
shows 78% of drug release. In M E preparation more
quantity of surfactant used which having low solubility of
Figure 5: TEM image of microemulsion
ACKNOWLEDGEMENTS :
Authors are thankful to Alembic Pharma, Vadodara
(Gujrat, India) for providing free gift sample of
itraconazole and also thankful to Parul Institute of
Pharmacy, Vadodara for providing all required facilities for
completion of research work.
REFERENCES :
1. Jain A, Jain S and Rawat S.Emerging fungal
infections among children: A review on its clinical
manifestations, diagnosis, and prevention. J of
Pharm & Bioallied sci. 2010; 314-320
2. Deveda P, Jain A, Vyas N et al.Gellified emulsion for
sustain delivery of Itraconazole for topical fungal
diseases.Int. J. of Pharm. and Pharm. Sci. 2010; 104-
112
3. Sailesh.Topical Drug Delivery Systems: A
Review”.Pharmaceutical information & blog. 2008
4. Rao Y, Sree K and Chowdary K. M icroemulsions: A
Novel drug carrier system. Int J Drug Delivery Tech.
2009; 2, 39-41
PHARMAGENE Vol: 1, Issue: 2
drug. So it delayed the drug release due to more
partitioning between internal and external components.
Antifungal activity: M icroemulsion based gel without
drug shows no zone of inhibition. But microemulsion based
gel with itraconazole shows 28.3 mm zone of inhibition and
marketed preparation shows 22.8 mm zone of inhibition.
This showed that prepared microemulsion based gel of
itraconazole is better than marketed formulation.
5. Shah R, M agdum C , Wadkar K et al.Fluconazole
Topical M icroemulsion: Preparation and
Evaluation.Research J. Pharm. and Tech. 2009;2, 353-
357
6. Dash M , Chandrasekaran N, M ukherjeeanti A,
“Bacterial activity of sunflower oil microemulsion,
Int. J. Pharm. and Pharm. Sci. 2010 , 2 ,123-128
7. Bachhavi Y and Patravalei V .Formulation of
meloxicam gel for topical application:In vitro and in
vivo evaluation. Acta Pharm. 2010; 60, 153–163
8. Barakat N, Fouad E and Elmedany A.Formulation
Design of Indomethacin-Loaded Nanoemulsion For
Transdermal Delivery. Pharm. Analytica Acta.
2011;1-8.
9. Sarkar B, Patel R, Bhadoriya U .Antimicrobial
Activity of Some Novel Pyrazoline Derivatives. J
Advanced Pharm Edu & Res .2011; 243-250
36