Development and Characterization of Ketoconazole Loaded Organogel for Topical Drug Delivery

Inventi Rapid: NDDS Vol. 2015, Issue 3

[ISSN 0976-3791] 2015 pndds 16335 © Inventi Journals (P) Ltd

Published on Web 24/06/2015, www.inventi.in

RESEARCH ARTICLE

INTRODUCTION In the last few years, delivery of lipophilic drug in the form

of gel have gained a lot of interest to most research

scientist due to its certain advantages like ease of

formulation, non toxicity and optimum permeable capacity

through skin. It is a challenge to enhance the permeation

ability of drugs using drug delivery systems with high drug-

loading capacity and at the same time low skin irritation via

topical administration. However, the poor penetration and

drug loading capacity of common products such as cream,

gel or patch limited their topical therapeutic effects. Hence,

organogel was found worth exploring in topical uptake of

any of such drugs.

The transdermal route of drug administration is

definitely one of the potential routes for local and systemic

delivery. This route provides a controlled diffusion of drugs

into the systemic circulation, breaks many barriers in

medical therapy like the need of assistance, intermediate

dosing and uncomfortable administration and improves

patient compliance. In general, gel-based products may be

categorized either as hydrogels or organogels depending

on the polarity of the external liquid component. Water is

the external liquid component of hydrogels while

organogels were formulated using non-polar solvents such

as hexane, isopropyl myristate, sunflower oil, corn oil or

others. The success of dermatological or transdermal drug

delivery systems depends on the ability of the drug to

penetrate into and/or permeate through skin in sufficient

quantities to achieve therapeutic levels. the use of

organogel based products is increasing due to their easy

method of preparation and inherent long-term stability. [1]

Depending on the mechanism of the formationof the three

dimensional gel skeleton, the organogels areconsidered as

1Department of Pharmaceutics, MET’s Institute of Pharmacy, Bhujbal

Knowledge City, Adgaon, Nashik-422003, Maharashtra, India.

E-mail: [email protected]

*Corresponding author

fluid-filled structures and solid-fiber based gels. [2] They are

thermoreversible and have the ability to accommodate

both hydrophilic and hydrophobic compounds within the

gel structure. This property has also widened the scope of

the organogels uses as controlled drug delivery systems

which can be taken via several routes of administration.

The gelators which compose the major skeleton of an

organogel are generally amphiphilic substances, such as

sorbitan monostearate (Sp60) and sorbitan monopalmitate

(Sp40). [3]

A gel may be defined as a semi-solid formulation having

an external solvent phase, apolar (organogel) or polar

(hydrogel) immobilized within the spaces available of a

three dimensional networked structure. [4] Organogels are

gels based on non-aqueous liquids, which have been

mentioned in various Pharmacopoeias as useful topical

deliveries for lipophilic drugs. Organogel not only exert a

local effect but also are capable of achieving systemic effect

through percutaneous absorption, when their lipophilic

nature and occlusive effect are potentiated by the presence

of a penetration enhances. [5]

Organogel systems are applied topically when the active

agent is oil-soluble or penetration into the deeper skin

layer is required. Surfactants act as penetration enhancers

that alter the membrane bilayer structure and thus reduce

the diffusion barrier and enable the drug to penetrate

deeply into skin. [6] Since the discovery of simple gelator

molecules, organogels have attracted increasing attention.

These novel formulations can be used in small quantities

without further additives, resulting in more biocompatible

products. [7] A wide variety of organogels have been

developed by researchers and classified based on the

nature of the organogelators, such as LO, gelatin-stabilized

organogels, limonene GP1/PG organogel, non-ionic

surfactant based organogels and polyethylene organogels. [8]

Organogels have been studied to have many

applications in pharmaceuticals, nutraceuticals, cosmetics,

food and so on. The scope of organogels further increases,

Development and Characterization of Ketoconazole Loaded Organogel for Topical Drug Delivery

Moreshwar P Patil1*, Ganesh P Shinde1, Sanjay J Kshirsagar1, Durgesh R Parakh1

Abstract: The aim of the present investigation was to formulate and evaluate stable ketoconazole organogel preparation to

increase the solubility of ketoconazole and release the drug for prolonged period of time. Ketoconazole was dissolved in clove

oil. The required amount of the tween 80 and PEG 400 was added to the clove oil containing propyl paraben. Subsequently,

water containing methyl paraben was added drop-by-drop to the organogelator solution with constant stirring on magnetic

stirrer until there was a formation of clear microemulsion. Carbopol 934 was used as a gelling phase, slowly mixed with

microemulsion in 1:1 ratio with constant and uniform stirring to get milky white homogeneous organogel. Formulated

organogels were evaluated for their physical appearance, pH, viscosity, globule size, drug content, spreadability, extrudability,

in-vitro and ex-vivo drug release, antifungal activity and stability. Organogel carrying ketoconazole showed good physical

appearance, acceptable skin pH (6 - 6.8), non-newtonian pseudoplastic system, drug content (99.68±0.19), globule size (572

nm), spreadability (21.67±0.034 gm.cm/sec), good extrudability, in-vitro release (98.75±0.32 %), ex-vivo release (73.45±0.86 %)

for optimized batch. Skin irritation study did not showed any irritation reaction and possess a good anti-fungal activity. The

optimized batch OG3 showed better drug release as compared to marketed cream. Similarly ex-vivo release of formulation

showed better drug release through mice skin as compared with marketed cream (KT CURE). The formulations followed zero

order kinetic model followed by higuchi mechanism. Thus, results of the current study clearly indicated a promising potential of

the ketoconazole organogel as an alternative to the conventional dosage form.

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RESEARCH ARTICLE

since; the topical route becomes one of the convenient

methods of drug delivery. Since, it is easy to manufacture

the commercialization process may also become cost

effective. Moreover it could also be able to cure chronic

diseases like osteoarthritis, when appropriate analgesic

drugs are incorporated. Now-a-days exposure to UV rays is

giving way to skin cancer in people. Organogels can become

a viable alternative, if anticancer agents are delivered

through them which have not been tried till date. [9]

Fungal infection is a common infection which affects

two third of population among the world. Many antifungal

drugs are available for treatment of fungal infection like

Miconazole, Itraconazole etc. Organogels, 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 delivery. For topical delivery microemulsion is

incorporated in Carbool 934 gel base to prolong the local

contact to the skin.

Ketoconazole is BCS Class II drug. Ketoconazole is orally

or topically active antifungal agent with a broad spectrum

of activity. It is effective against several fungal strains such

as Candida albicans which are responsible for topical

candidiasis. Ketoconazole shows very low water solubility

and having log P value 4.35 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 semisolid

preparations are widely accepted over solid and liquid

dosage forms. Hence, limitation in formulating oral dosage

forms. Such drugs pose a challenge in development of

topical drug delivery system. This study was aimed to

formulate and evaluate stable organogel formulations

containing ketoconazole. Organogel are composed of clove

oil as oil phase, PEG 400 and Tween 80 as organogelator

and aqueous phase in appropriate ratio, carbopol as

consistency modifier, methyl and propyl paraben as

preservatives. Delivery of drugs using these organogel

through skin increases the local/systemic delivery by

different mechanisms that make them suitable vehicles for

the delivery of anti-fungal agents. To achieve these

objectives, the organogel was evaluated for the influence of

pH, rheological properties, gel sol transition study,

spreadability, in-vitro drug release, globule size,

extrudability, drug content, ex-vivo release and skin

irritation study. The anti-fungal activity of selected

ketoconazole containing formulation using modified disc

diffusion method had been evaluated and compared with

marketed cream formulation (KT CURE cream).

MATERIALS AND METHODS Ketoconazole was kindly gifted by Holden Medical

Laboratories Ltd., Sinnar (Nashik), India. Carbopol 934,

PEG 400, Tween 80, clove oil, methyl paraben, propyl

paraben, sodium hydroxide, potassium dihydrogen

phosphate and methanol were supplied by Thomas Baker

Mumbai, India. All the chemicals used during study were of

analytical reagent grade.

Solubility Determination of Ketoconazole Determination of appropriate oil, surfactant and

cosurfactant that had good solubilizing capacity of

ketoconazole and thus could be used as the oil phase and

organogelator in organogel, the solubility of ketoconazole

in various oils, suractants and cosurfactants were

measured.

Determination of Drug Solubility in Different Oils and Surfactants The solubility of ketoconazole in various oils (rice bran oil,

sunflower oil, clove oil, soyabean oil), surfactants (Labrasol,

Tween 80, Tween 20, Span 80) and cosurfactants (PEG 400,

PEG 200, Propylene Glycol) were determined by adding an

excess amount of drug to 3 ml of selected oils, surfactants

and cosurfactants separately in 10 ml capacity stopper vials

and mixed using a vortex mixer (Remi motor, Mumbai,

India). The mixture vials were then shaken for 48 hrs at

40±0.5°C using magnetic stirrer. Further the mixtures were

kept aside for 24 hrs at room temperature to reach

equilibrium. The equilibrated samples were then

centrifuged at 3000 rpm for 20 min. The supernatant was

taken and filtered through a 0.45 μm membrane filter. The

filtrates were diluted with methanol and ketoconazole

concentration was subsequently quantified by U.V.

spectrophotometer at 242 nm. [10]

Construction of Pseudo-ternary Phase Diagrams On the basis of solubility studies, the clove oil was selected

as the oil phase. Tween 80 and PEG 400 were selected as

surfactant and cosurfactant. Distilled water used as

aqueous phase. The pseudo-ternary phase diagrams were

constructed using water titration method to determine the

micro emulsion region and to detect the possibility of

making microemulsions with different possible

compositions of oil and surfactants. The ratios of surfactant

and cosurfactant (Tween 80:PEG 400) were chosen as 1:1,

1:2, 1:3, 2:1 and 3:1. Such mixtures were prepared. These

mixtures were mixed with the oil phase to give the weight

ratios of 90:10, 80:20, 70:30, 60:40, 50:50, 40:60, 30:70,

20:80 and 10:90. Water was added drop by drop and

stirred using a magnetic stirrer until a homogeneous

dispersion or solution was obtained. After each addition,

the system was examined for the physical appearance. The

end point of the titration was the point where the solution

becomes cloudy or turbid. Pseudo ternary plots were

constructed using Chemix School Software, trial version 3.6

(Oslo, Norway) and microemulsions were prepared by

ternary phase diagram. [11-12, 10]

Formulation Development of Organogel 1. Selection of Ratio of Surfactant and Co-surfactant for Formulation of Organogel The five ratios like 1:1, 1:2, 1:3, 2:1 and 3:1 were prepared

and plotted with pseudo ternary phase diagram by using

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RESEARCH ARTICLE

Chemix school software v3.6 trial version. The pseudo

ternary phase diagram showing larger region was selected

for formulation and development of organogel.

2. Preparation of Organogel The eight organogel formulations were prepared and

formulations that containing composition having surfactant

ratio 3:1 was taken for the preparation of organogel.

Composition of different formulation of organogel is shown

in Table 1.

Accurate quantity of ketoconazole was weighed as per

the formula. Propyl paraben was dissolved in oil. Tween 80

and PEG 400 were mixed thoroughly in the proportion of

3:1 ratio (w/w) to obtain the surfactant cosurfactant

mixture (Smix), which was used as gelator. Specified amount

of the Smix was added to the clove oil (CO) containing propyl

paraben; Ketoconazole was added to the mixtures of oil,

surfactant mixture with appropriate percentages as

described in Table 1. The above mixture was stirred on

magnetic stirrer for 20 min. Subsequently, water

containing methyl paraben was added drop-by-drop to the

gelator solution (GS) using a burette with constant stirring

on magnetic stirrer until there was a formation of clear

microemulsion. Carbopol 934 used as gelling phase added

into microemulsion phase. Thus organogel were obtained

spontaneously on stirring the mixture. Depending on the

composition of the GS-water mixture, the system either

formed gelled structures or remained as microemulsion. A

ternary plot depicting the proportions of Smix, water and CO

was prepared to figure out the compositions, which formed

organogels. The final concentration of ketoconazole in

organogels was maintained at 2 % (w/w).

Evaluation of Organogel 1. Physical appearance The formulated ketoconazole organogel formulations were

observed visually for their color, homogeneity and

consistency, presence of any clog and sudden change in

viscosity. [13]

2. pH The pH of all formulations was measured by digital pH

meter (Systronic, Mumbai, India). The pH of organogel is

also required to measure because change in pH may affect

the zeta potential and finally affect the stability of products.

The pH meter was calibrated before each use with standard

pH 4 and 7 buffer solutions. The electrode was immersed in

organogels and readings were recorded on pH meter. The

measurement was performed at ambient temperature and

in triplicate. [14-15, 5]

3. Rheological Study Rheology is an important parameter as it affects the

spreadibility and adherence of the transdermal

formulations to the skin surface. The rheogram of

organogel was obtained at 25±1°C with a Brookfield Digital

viscometer-LV DV E (Brookfield, Massachusetts, USA). The

viscosity was measured by using spindle S-64. The viscosity

of all formulations was measured at different rotational

speed (rpm) i.e. at 4, 5, 6 and 10. [13, 16, 5]

4. Spreadability The spreadability of the formulation was determined using

an apparatus suggested by Mutimer. The apparatus

consisted of two glass slides (7.5 × 2.5 cm), some of which

was fixed onto the wooden board and the other was

movable, tied to a thread which passed over a pulley

carrying a weight. 1 gm of formulation was placed between

the two glass slides. 50 gm weight was allowed to rest on

the upper slide for 1 to 2 minutes to expel the entrapped

air between the slides and to provide a uniform film of the

formulation. The weight was removed and the top slide

was subjected to a pull obtained by attaching 45 gm weight

over the pulley. The time required for moving slide to

travel premarked distance i.e. 7.5 cm was noted. The

readings obtained were indications of relative spreadability

of different formulations. Spreadability is expressed in

terms of time in seconds taken by two slides to slip off from

jellified organogel and placed in between the slides under

the direction of certain load. Lesser the time taken for

separation of two slides, better the spreadability. [13, 17]

It is calculated by using the formula:

S = M. L / T

Where, M = Weight tied to upper slide, L = Length of

glass slides, T = Time taken to separate the slides.

5. Extrudability Test (Tube Test) Extrudability test is based upon the determination of

weight required to extrude 0.5 cm ribbon of organogel in

10 sec. from lacquered collapsible aluminum tube. The

extrudability value was calculated using following formula:

Table 1: Composition of Organogel Formulations

Ingredient (% w/w) OG 1 OG 2 OG 3 OG 4 OG 5 OG 6 OG 7 OG 8

Ketoconazole 2 2 2 2 2 2 2 2

Clove oil 20 10 15 20 10 15 15 10

Tween 80 33.75 33 37.5 33 41.25 30 45 45

PEG 400 11.25 17 12.5 17 13.75 25 15 15

Carbopol 934 1 1.5 0.5 0.5 1 0.5 1 1.5

Methyl paraben 0.03 0.03 0.03 0.03 0.03 0.03 0.03 0.03

Propyl paraben 0.01 0.01 0.01 0.01 0.01 0.01 0.01 0.01

Water 35 40 35 30 35 30 25 30

Triethanolamine q.s q.s q.s q.s q.s q.s q.s q.s

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Extrudability = Weight applied to extrude organogel from

tube (gm) / Area (cm2). [18, 19]

6. Globule Size and its Distribution in Organogel Globule size and distribution was determined by Malvern

Zetasizer. About 1.0 gm sample was dissolved in double

distilled water and agitated to get homogeneous

dispersion. Sample was injected to photocell of zetasizer.

Mean globule diameter and distribution was obtained. [20]

7. Drug Content Determination Drug concentration in organogel was measured by UV

spectrophotometer. 1 g of the prepared gel was

dissolved in 100 ml of methanol. The solution was

sonicated to dissolve the drug in methanol. About 1 ml of

solution was withdrawn and further diluted to 10 ml.

Then absorbance was measured at 242 nm in UV/VIS

spectrophotometer. The % drug content was calculated

using the equation, which was obtained by linear

regression analysis of calibration curve of drug in

methanol. [15, 20]

8. In-vitro Diffusion Study In-vitro diffusion was carried out by modified Franz

diffusion cell. A glass cylinder with both ends open, 10 cm

height, 3.7 cm outer diameter and 3.1 cm inner diameter

was used as diffusion cell. An egg membrane (soaked in

phosphate buffer 24 hours before use) was fixed to one end

of the cylinder with the aid of an adhesive. About 1gm of

organogel was taken in the cell (donor compartment) and

cell was immersed in a beaker containing 500 ml of

phosphate buffer (pH 6.8) as receptor compartment. The

entire surface of the cell was in contact with the receptor

compartment which was agitated using magnetic stirrer

and a temperature of 37±1°C was maintained. Sample of 5

ml of the receptor compartment was removed at 1 hour

interval of time over a period 8 hours with same amount

replaced to maintain sink condition. The sample was

analyzed at 226 nm against blank using UV

Spectrophotometer. Amount of ketoconazole released at

various time intervals was calculated with the help of

calibration curve with phosphate buffer (pH 6.8) and

plotted against time. [14-15, 18] 9. Ex-vivo Diffusion Study The optimized formulation and the formulations giving

better in-vitro drug diffusion rate were selected for the ex-vivo diffusion study. The wistar mice weighing average

175±25 g were shaved at abdominal region. After

anaesthesia to the rats, the abdominal skin was removed

surgically from the animal and adhering subcutaneous fat

was carefully cleaned. The dermal side of the skin was kept

in contact with phosphate buffer, pH 6.8 for 2 hrs before

start of study. Organogel formulation, 1 gm was placed on

the membrane and dipped it into receptor medium and

maintained the temperature at 37±1°C, aliquots of 5 ml

were withdrawn at different time intervals and same

volume of buffer was added to maintain sink conditions.

The release profile data of prepared organogel (OG3)

formulation was compared with the marketed cream (KT

CURE cream).

10. Skin irritation study Skin irritation Study performed by using of the Hen’s Egg

Test – Chorioallantoic Membrane (HET- CAM) test method.

a. CAM Preparation Select fresh (not older than 7 days), clean, fertile 50-60 g

White Leghorn chicken eggs. Place eggs in an incubator

with a rotating tray. Incubate eggs at 38.3±0.2°C and

58±2% relative humidity in a forced-air incubator. Hand

rotate eggs five times per day until day 8. Candle the eggs

on incubation day 8 and remove any nonviable or defective

eggs. Remove eggs from the incubator on day 9 for use in

the assay. Discard any nonviable or defective eggs. Mark

the air cell of the egg. Cut the section marked as the air cell

with a rotating dentist saw blade and then pare it off.

Moisten the inner membrane with 0.9% NaCl. Place the egg

into the incubator for a maximum of 30 minutes. Remove

the egg from the incubator and decant the 0.9% NaCl

solution. Carefully remove the inner membrane with

forceps.

b. Procedure

Apply Organogel onto the CAM surface.

c. Observation Observe the reactions on the CAM over a period of 300

seconds. Endpoints that should be observed are:

i. Hemorrhage (bleeding from the vessels)

ii. Vascular lysis (blood vessel disintegration)

iii. Coagulation (intra- and extra-vascular protein

denaturation). [20, 21]

11. Anti-fungal Study a. Disc Diffusion Assay i. Preparation of Standard Solutions Ketoconazole was used as the reference standard drug

which was prepared in DMSO.

ii. Test Organism Used Candida albicans MTCC 227 iii. Preparation of Culture Medium Sabouraud dextrose agar was used as culture medium for

antifungal activity.

iv. Preparation of Inoculum This was sterilized by autoclaving at 15 lbs/sq. inch

pressure for 15 min. After sterilization a lapful of organism

was transferred from mother culture to this sterilized SDA

medium. Here Pour plate technique was utilized.

v. Sterilization of Apparatus Petri dishes, glass syringe, test tubes, conical flask were

sterilized by autoclaving at 15 lbs/sq. inch for 15 min.

vi. Preparation of Petri Dishes

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After transferring inoculated medium to the Petri dishes it

was allowed to solidify. After the solidification of the media

the sterile filter discs which were dipped in drug solution

were placed on it. DMSO was used as the control. The plates

were kept in refrigerator at 8°-10°C for proper diffusion

into the media. Then the plates were transferred to the

Biological Oxygen Demand (BOD) incubator and

maintained at 37°±3°C for 24–48 hrs. After incubation the

petri plates were observed for the zone of inhibition. The

diameter of zone of inhibition was reported in mm. [23, 22, 13]

12. Stability Studies The prepared ketoconazole organogel formulations were

stored away from light in collapsible tube at 40°C and 75%

RH for 3 months. After storage the samples were tested for

their physical appearance, pH, % drug release, viscosity

and % drug content. [22, 15] RESULTS AND DISCUSSION Selection of Oils, Surfactants for Formulation Study

Oil, surfactant and cosurfactant were selected on the basis

of results of solubility study obtained. Surfactants like

Tween 80, cosurfactant like PEG 400 and oil like clove oil

had showed higher solubilizing capacity for the drug, hence

selected for further study.

Physical Appearance The prepared organogels were evaluated for physical

appearance, homogeneity, consistency and change in

viscosity. The results indicated that the organogels were

creamy white in colour, homogenous without grittiness;

having semisolid consistency without change in viscosity

for required period. This confirms the stability of

organogels.

pH Determination Since, topical systems are directly applied on the skin; their

pH should be compatible with the skin pH. An acidic or

basic pH causes skin irritation or disruption of the skin

structure. The pH of all formulations were found to be

between 6-6.8 which is acceptable for skin preparations

summarized in Table 2.

Rheological Study Rheological behaviour of the organogel indicated that the

systems were showed non-newtonian shear thinning

pseudo plastic type of flow, i.e. shear thinning in nature

showing decrease in viscosity at the increasing shear rates.

The gelator molecules involved in the formation of the

gelled structures via fluid-filled microstructures. As the

applied shear is increased, the hydrophobic interactions

are not able to keep the fluid-filled microstructures

together. This results in the transition of the system from

the gelled phase to the free-flowing liquid phase, marked

by the disruption of the 3-dimensional networked

structures. An increase in the concentration of carbopol

934 and water (0.5 to 1.5%) were expected to show

increase in viscosity. All formulations exhibited shear

thinning properties. The results are depicted in Table 3. Spreadability Study One of the essential criteria’s for an organogel that it

possesses good spreadability. A more viscous formulation

would have poor spreadability. Spreadability is a term

Table 2: pH of Different Organogel Formulations

Formulations pH (mean±SD)* OG 1 6.13±0.55

OG 2 6.01±0.44

OG 3 6.75±0.06

OG 4 5.68±0.25

OG 5 6.27±0.38

OG 6 6.07±0.45

OG 7 6.54±0.01

OG 8 5.98±0.23 *(n=3)

Table 3: Viscosity of Different Organogel Formulations

RPM Viscosity (cps)

OG 1 OG 2 OG 3 OG 4 4 89000 100200 46900 53180

5 79400 86700 43200 47710

6 68980 76700 40190 41300

10 49200 54650 31790 30430

RPM Viscosity (cps)

OG 5 OG 6 OG 7 OG 8 4 56200 61900 78900 108300

5 50850 52100 68300 90150

6 42340 45590 60390 79980

10 34220 36770 43610 58320

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expressed to denote the extent of area to which gel readily

and spreads on application to the skin. The therapeutic

efficacy of a formulation also depends upon its spreading

value. The spreadability of different organogel formulations

is shown in the Table 4.

From the result obtained it was observed that the OG3

formulation shows the more spreading coefficient as

compared to other formulations shown in Table 4. OG3

formulation gives the spreading coefficient ±gm.cm/sec

which may be due to presence of optimum concentration of

gelling agent.

Extrudability Study Extrudability of the organogel is depending upon the

viscosity of that organogel. Less viscid the organogel, lesser

the force required to remove it from tube, thus shows

better extrudability. The extrudability of formulated

organogels is showed in Table 5.

From the obtained data it can be concluded that, the

formulations OG2 and OG8 were contained highest amount

of carbopol 934 i.e.1.5%, so it showed lowest extrudability

due to highest viscosity among all formulations. OG6 and

OG3 had showed best extrudability than other formulations

Table 4: Spreadability of all Organogel Formulations

Formulations Spreadability (gm.cm/sec) (mean±SD)* OG 1 7.88±0.12

OG 2 5.42±1.9

OG 3 21.67± 0.034

OG 4 17.33±0.85

OG 5 13.68±1.67

OG 6 12.38±0.59

OG 7 9.29±1.08

OG 8 4.91±0.77

Marketed cream 18.57±0.39 *(n=3)

Table 5: Extrudability of Different Organogel Formulations

Formulations Extrudabiility OG 1 ++

OG 2 +

OG 3 +++

OG 4 ++

OG 5 ++

OG 6 +++

OG 7 ++

OG 8 + (+ = fair, ++ = good and +++ = excellent)

Table 6: Globule Size of Optimized Organogel Formulation

Formulation Globule Size (nm)

OG 3 572

Figure 1: Globule size distribution in OG 3 organogel formulation

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as it contained carbopol 934 in 0.5%. Other formulations

also showed good extrudability due to less composition of

gelling agent.

Globule Size and its Distribution in Organogel From result we can conclude that globule size in all

organogel formulation decreases when the concentration of

surfactant and co-surfactant decrease. As compared with

globule size of micro emulsion with organogel there is

slight increase in globule size of organogel may be due to

effect of addition of gelling agent into it, due to gelling

agent might have led to the entrapment of oil globule in its

network thus, causing slight increase the interfacial tension

between oil water phases. Globule size distribution of

optimized formulation showed in Table 6 and Figure 1.

Drug Content Determination The drug content of all formulations was found to be in the

range of 97% to 99% given in Table 7. Hence uniformity of

drug content was found satisfactory and was within

Pharmacopoeial limits i.e. 98.0 to 102.0 % of ketoconazole

according to I.P. 2014. [24]

In-vitro Diffusion Study In-vitro release profiles of ketoconazole from its various

organogel formulations are represented in Table 8. It was

Table 7: Drug Content of Different Organogel Formulations

Formulations Drug Content (%)(mean±SD)* OG 1 98.05±0.07

OG 2 97.84±0.13

OG 3 99.68±0.19

OG 4 98.99±0.12

OG 5 98.78±0.07

OG 6 98.57±0.07

OG 7 98.27±0.07

OG 8 97.97±0.13 *(n=3)

Table 8: Cumulative Amount of Ketoconazole Diffused (%) from Organogel Formulations using Modified Diffusion Cell

Time (Hrs.) % Cumulative Drug Release (mean±SD)* OG1 OG2 OG3 OG4

1 10.94±1.07 10.15±0.56 15.63±0.25 13.75±0.3

2 17.24±0.88 16.18±1.02 28.44±1.92 34.47±0.6

3 23.76±0.48 21.53±0.87 40.25±0.85 42.29±0.4

4 30.12±0.42 26.65±2.06 53.54±1.04 49.74±0.3

5 41.13±0.98 32.06±3.21 63.95±0.87 60.55±1.7

6 53.41±2.69 44.84±2.06 77.68±3.01 72.69±0.4

7 62.11±3.02 52.37±1.38 86.65±2.15 81.09±1.8

8 73.07±3.15 68.01±0.19 98.75±0.32 86.40±1.0

Time (Hrs.)

% Cumulative Drug Release (mean±SD)* OG5 OG6 OG7 OG8 M. Cream

1 16.01±1.16 17.50±1.49 18.58±0.17 10.25±0.54 11.11±2.22

2 25.61±1.06 33.57±1.01 28.6±0.66 16.99±0.67 16.11±2.56

3 34.88±1.1 43.09±3.37 40.03±1.59 23.89±2.7 25.59±2.15

4 41.94±1.05 53.40±1.77 53.26±1.88 31.33±1.51 32.8±5.69

5 49.3±0.4 59.48±0.67 65.67±1.82 42.44±1.42 42.32±4.98

6 58.43±1.1 70.48±2.10 72.71±3.04 52.51±0.57 52.63±4.66

7 66.24±1.1 77.11±4.08 81.63±3.81 60.11±0.94 57.51±2.95

8 80.09±0.7 87.34±0.38 90.92±2.50 77.05±2.70 65.94±3.30

Table 9: Ex-vivo Drug Release from Organogel Formulation and Marketed Cream

Time (hr.) % Cumulative Drug Release (mean±SD)* OG 3 Marketed Cream

1 9.35±1.01 8.34±0.5

2 12.08±0.6 9.32±0.2

3 18.81±0.26 13.63±1.2

4 27.01±0.9 19.26±0.7

5 36.93±1.7 24.09±0.65

6 53.62±0.43 29.98±1.87

7 64.26±0.32 46±0.12

8 73.45 ± 0.4 54.44±0.8

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RESEARCH ARTICLE

observed that all formulations were become swelled at the

end of experiment due to penetration of diffusing media

into gel matrix which cause breaking of gel matrix and thus

release of drug. The higher drug release was observed with

formulations OG3 and OG6. This may be due to presence of

minimum amount of gelling agent i.e. carbopol 934. The

minimum amount of carbopol 934 cause less viscous

formulation as compared to other organogel formulations,

leads to less packed gel matrix which easily get breaked,

thus higher release of drug. The formulations OG3 and OG6

showed 98.75±0.32% and 87.34±0.38% cumulative drug

release at the end of 8 hrs. respectively and these are

shown in Table 8. Initial burst release followed by control

release was observed for all formulations. When data was

plotted according to zero-order kinetics linear plots were

obtained. Highest regression coefficient values were

obtained for zero-order the values ranging from 0.965 to

0.989, suggesting the mechanism of release from all the

formulations followed Zero-order kinetics.

Ex-vivo Diffusion Study Ex-vivo diffusion study was performed using fresh mice

skin. The diffusion study was performed on both standard

marketed ketoconazole cream and formulated

ketoconazole organogel (OG3). The % drug release of both

marketed gel and organogel are given in Table 9. The

standard ketoconazole gel showed 54.44±0.8% permeation

of ketoconazole through mice skin at the end of 8 hrs,

whereas formulated organogel i.e. OG3 showed permeation

of ketoconazole was 73.45±0.4 at 8 hrs. The OG3 batch

contains carbopol 934 in the concentration of 0.5% w/w, at

this concentration carbopol 934 possess lower viscosity

and thus imparts in better diffusion of drug through loosely

bound network of gel, provide good release of drug and

thus increasing permeation. Both of these figures indicated

that formulated organogel gives higher flux and permeation

as compared to standard marketed gel. The enhanced

permeation from the organogel due to the smaller droplet

size of the micro emulsion, ketoconazole directly diffuse

Figure 2: Positive control

Figure 3: Negative control

Figure 4: Test substance

Figure 5: Zone of inhibition for optimized formulation OG 3

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RESEARCH ARTICLE

from the droplets to the stratum corneum without micro

emulsion fusion to the stratum corneum and subsequent

permeation enhancement.

Skin Irritation Study

1. HET CAM Test Incubated Hen’s Eggs were subjected for HET CAM Test

and observed for either coagulation or hemorrhage or

haemolysis after 300 seconds of application of test

substance. Results are discussed in Figure 2-4 and Table

10. Coagulation was observed within 9 seconds of

application in case of positive control while no coagulation

or any viable reaction was observed in case of Negative

control and Test substance. Following photographs shows

the Observations for HET CAM test.

Anti-fungal Study

Figure 5 and 6 shows that the zone diameter of standard

ketoconazole and organogel sample are comparable. This

shows that the organogel optimized batch OG3 is more

effective in inhibiting the growth of the Candida albicans

species. Results are shown in Table 11.

Stability Study Short term accelerated stability study was performed at

40°C and 75% RH for 3 months. After the period of 3

months the organogel (OG3) formulation was tested for its

physical appearance, pH, drug content, viscosity and drug

release. After performing tests organogel formulation was

found to be creamy white and homogeneous. The pH,

viscosity, drug content and drug release of formulation was

found to be 6.75±0.06, 46900 cps, 99.68±0.19% and

98.75±0.32% respectively. Table 12 indicating the

prepared formulation was stable for the storage period.

CONCLUSION The current study deals with the development of Tween

80-PEG 400 based organogels. Amongst all formulations,

organogel prepared with oil (15%), Smix (50%), carbopol

934 (0.5%) and water (35%) were better with respect to

overall product qualities. When organogel was compared

with the marketed gel, drug release from the organogel was

found to be increased and prolonged. The formulations

followed zero ordered kinetic model of drug release which

involves control release. In-vitro diffusion studies indicated

that the organogels may be used as matrix for controlled

delivery systems. After performing anti-fungal study, it

was found that the anti-fungal activity of formulated

organogel (OG3) was better than standard marketed

ketoconazole cream. The formulated organogel showed no

irritation after performing skin irritation study using HET-

CAM Test. The stability of the organogels was found to be

dependent on the Smix and water proportions. The results

Figure 6: Zone of inhibition for placebo, marketed and optimized OG 3 formulation

Table 10: Results of HET CAM Test

Test Negative Control (0.9% NaCl)

Positive Control (0.1N NaOH)

Test Substance Score

(Optimized OG 3 Batch)0 9 0

Table 11: Zone of Inhibition of Ketoconazole against Candida albicans

S. No. Formulation Concentration (μg/ml) Zone of Inhibition against Candida albicans 1 Standard 50 17 mm

2 Test 50 18 mm

3 Placebo 0 6 mm

Table 12: Stability Study of Optimized Organogel Formulation

Formulation Appearance pH (mean±SD)* % Drug Content (mean±SD)*

% Drug Release (mean±SD)*

OG 3 White, creamy and homogeneous 6.75± 0.06 99.68 ± 0.19 98.75±0.32 *(n=3)

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RESEARCH ARTICLE

indicated that the developed matrices may be used as a

vehicle for controlled delivery system. Thus, results of the

current study clearly indicated a promising potential of the

ketoconazole organogel as an alternative to the

conventional dosage form. However, further clinical studies

are needed to assess the utility of this system. By

considering all above points it was concluded that, the

objective of the present research study can be achieved

successfully.

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Cite this article as: Moreshwar P Patil, Ganesh P Shinde,

Sanjay J Kshirsagar et al. Development and

Characterization of Ketoconazole Loaded Organogel for

Topical Drug Delivery. Inventi Rapid: NDDS, 2015(3):1-

10, 2015.

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Development and Characterization of Ketoconazole Loaded Organogel for Topical Drug Delivery

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