Pharmacology & Pharmacy, 2011, 2, 289-298
doi:10.4236/pp.2011.24037 Published Online October 2011 (http://www.SciRP.org/journal/pp)
289
Design and Development of an in Vitro Assay for
Evaluation of Solid Vaginal Dosage Forms
Jyoti Gupta1, Jason Qihai Tao2, Sanjay Garg1, Raida Al-Kassas1*
1School of Pharmacy, Faculty of Medical and Health Sciences, The University of Auckland, Auckland, New Zealand; 2International
Partnership for Microbicides, Silver Springs, USA.
Received February 27th, 2011; revised April 11th, 2011; accepted July 20th, 2011.
ABSTRACT
Vaginal dosage forms are seen as a viable option for empowering women to protect themselves from the risk of HIV
transmission. Because of limited research in the field, there is a lack of suitable dissolution methods established for
determination of drug release from vaginal formulations inside the vaginal tract. The main aim of this study was to de-
velop a simple, reliable and reproducible in vitro release method for evaluation of solid vaginal dosage forms (VDFs)
which was hoped to exhibit a close in vitro-in vivo correlation. Dapivirine, a drug being developed as a microbicide
and a well estab lished marketed a nti fungal d rug, Clotrimazole were used as model drugs. Two doses (0.5 mg and 1.25
mg) of Dapivirine were prepared as novel rapidly disintegrating, bioadhesive tablets. Clotrimazole 100 mg, prepared in
house as conventional release tablets and commercially available Canesten (Clotrimazole tablet 100 mg) were used.
The in vitro drug release testing of these tablets was carried out using a designed system which consisted of modified
USP dissolution Appa ratus II in conjunction with Enhancer cell (as sample holder) in 150 ml capacity flasks instead of
the standard 900 ml flasks. The suitability of the system was investigated for variable parameters such as formulation
types, drug concentration, stirring speeds, media volume and comparison of in house product with marketed product.
The method was successfully optimized at a volume of 100 ml and a low speed of 25 rpm at pH 4 and was found sensi-
tive enough to distinguish between formulation s and evaluate products of different streng ths. A linear drug release pro -
file (R2 = 0.99) was obtained in case of Dapivirine, indicating that drug release is controlled by diffusion. The devel-
oped dissolu tion system has a potential to exhibit a good in vitro-in vivo correlation in addition to carrying out routine
dissolution tests for solid VDFs.
Keywords: Vaginal Solid Dosage Forms, Enhancer Cell, Dapivirine, Clotrimazole
1. Introduction
Administration of drugs in the vagina is believed to be as
old as pharmacotherapy, first written documents dating
from 19th century BC [1]. Vaginal drug delivery presents
several advantages including, ease of administration,
possibility of self-administration, hepatic first pass-effect
bypass, low systemic drug exposure and increased per-
meability for some drugs when compared to the oral or
other routes [1]. Vaginal dosage forms available around
the world include creams, gels, tablets, capsules, pessaries,
foams, ointments, films, tampons, rings and douches.
While majority of vaginal drugs so far have been in the
form of gels, there is a growing interest in alternative
dosage forms such as tablets, rings, and films. Conven-
tional vaginal dosage forms are associated with limita-
tions of poor retention, leakage and messiness causing
inconvenience to users, leading to poor subject/patient
compliance and loss of therapeutic efficacy [2]. In recent
years vaginal bioadhesive tablets have been developed as
a new type of controlled release form for the treatment of
both topical and systemic diseases. The greatest advan-
tages of such bioadhesive tablets are the release of drug
at a controlled rate and the possibility of maintaining
them in the vagina for extended periods of time [3].
Dissolution testing is routinely used in Quality Control
(QC) and Research and Development (R & D). To date
none of the official compendia [4-6] have included a
standard method for evaluation of release pattern from
vaginal preparations. Modified USP apparatus [7-10] and
various other methods [11-13] have been employed by
researchers to study the release of drugs from vaginal
formulations. In these studies a standard dissolution ap-
paratus with large volume of media has been employed

Design and Development of an in Vitro Assay for Evaluation of Solid Vaginal Dosage Forms 290
which does not simulate the actual physiological condi-
tions of the vagina, characterized by small volume of
fluid. Volume plays a significant role in the rate of drug
released and absorbed. In addition, other parameters such
as speed of agitation and pH should be considered in
order to assure close simulation of in vivo situation.
There are several systems used in the industry to study
the release of drugs from various pharmaceutical dosage
forms including vaginal formulations. These include USP
apparatus 4 (Flow through cell), Franz cells and Enhan-
cer cells [14,15]. Flow through cell could be used for
solid and semi solid vaginal formulations; however the
equipment is quite expensive and not available in many
laboratories. On the other hand, Franz cell has been pro-
posed to be one of the USP standard apparatus for topi-
cal/transdermal formulations. However, it is mostly suit-
able for semi-solid formulations and requires highly
trained technician to operate it and a membrane to be a
supportive physical barrier. The release of the drug from
the formulation, in most instances, depends on the type
of membrane to be used and its diffusion through it. En-
hancer cell has also been used to evaluate release profiles
of semi-solid formulations. The system consists of a
paddle over enhancer cell with a small dissolution vessel.
To date, most of the membranes used in this method
were synthetic membranes or filters. This system utilizes
the existing USP apparatus and is thus easy to set up,
portable and reproducible.
In the present study, we have employed Enhancer cell
system in conjunction with USP dissolution apparatus II
and 150 ml capacity flasks to replace the 900 ml flasks
used in the standard apparatus. The key difference of our
design is “mesh membrane” which replaced the synthetic
membranes most commonly used in past. The mesh
membrane of adequate pore size holds the formulation in
place and provide physical barrier which prevents the
formulation from being agitated too much while allowing
the drug to migrate or release freely from the formulation
matrix to the media and allowing some contact/exchange
of the media and the sample in the cavity. This system
adopts physiological parameters of the vagina (volume of
fluid, speed of agitation and pH), in an attempt to obtain
a standard quality control procedure for evaluation of
vaginal tablets, to minimize variables in results and to
achieve a close approximation to in vivo situation.
The aims of our design and the study were to develop
a simple, reliable and reproducible in vitro release
method for evaluation of solid vaginal dosage forms
(VDFs) which mimicked in vivo vaginal application of a
tablet dosage and was hoped to exhibit an adequate in
vitro-in vivo correlation for future developments.
The drugs selected for the study were Dapivirine and
Clotrimazole. Dapivirine, also known as TMC120, is a
substituted diarylpyrimidine derivative and a non nucleo-
side reverse transcriptase inhibitor (NNRTI) being de-
veloped as a vaginal microbicide for prevention of HIV
transmission [16]. Vaginal tablets of Dapivirine were
prepared to disintegrate rapidly in the presence of small
volume of fluid and form a bioadhesive gel from which
the drug is slowly released over a period of time (there is
no evidence yet for this bioadhesive tablet to be a con-
trolled release formulation).
Clotrimazole is a chlorinated imidazole derivative wide-
ly prescribed for the treatment of vulvovaginal candidiasis
VVC. Vaginal tablets of Clotrimazole were prepared in
house as conventional release tablets.
2. Materials and Methods
2.1. Materials
Dapivirine was obtained as a gift from International Part-
nership for Microbicides (IPM). Clotrimazole was also
received as a gift from Zhejiang Chemicals Import and
Export Corporation, China. Canesten tablets (Clotrima-
zole tablets 100 mg) were procured from a local phar-
macy in New Zealand. The directly compressible lactose
was procured from Fonterra (New Zealand), microcrys-
talline cellulose (MCC) from Asahi Kasei Corp (Japan),
magnesium stearate from Acros Organic (USA), sodium
starch glycollate from Spectrum, USA. Ammonium ace-
tate was purchased from BDH Laboratory (England),
sodium phosphate dibasic from Riedel-de Haen (Ger-
many), sodium phosphate monobasic, acetonitrile and
methanol from Merck (Germany). Sodium lauryl sul-
phate was purchased from Serva Electrophoresis and
tween 80 from Sigma Aldrich (Germany).
2.2. Tablet Preparation
Dapivirine was prepared in two different doses (0.05%
and 0.125%) as novel rapidly disintegrating, long acting
bioadhesive tablets. Dapivirine being a low dose drug
was mixed geometrically with polymer, diluents, su-
perdisintegrant and acid buffering agents to ensure a
homogeneous mixing of the drug with the excipients.
The mixing was carried out in a pilot scale cube mixer
attached to a multipurpose unit operator (Erweka, Ger-
many). The detailed composition and manufacturing
procedure for Dapivirine cannot be disclosed due to In-
tellectual Property reasons.
Clotrimazole 100 mg conventional vaginal tablets were
prepared as follows: Clotrimazole, passed through 60
mesh, was mixed with MCC (passed through 30 mesh) in
the cube mixer for 30 minutes. Directly compressible
grade lactose was passed through 30 mesh and added to
the mix of Clotrimazole and MCC and mixed for one
hour. Magnesium stearate and sodium starch glycollate

Design and Development of an in Vitro Assay for Evaluation of Solid Vaginal Dosage Forms291
after passing through 44 mesh were added to the above
blend and mixed for additional 10 minutes.
The tablets of Dapivirine and Clotrimazole were pre-
pared by direct compression technique and compressed
using a single punch tablet press (Cadmach, India) with
almond shaped (23 × 13 mm) punch. The average weight
of Dapivirine and Clotrimazole tablets were set as 1.0 gm.
These tablets were characterized for various parameters
including hardness, thickness, moisture content, disinte-
gration time and drug content.
2.3. Methods of Analysis
The HPLC methods were developed for Dapivirine and
Clotrimazole and validated for linearity, specificity, ac-
curacy and precision according to ICH guidelines (ICH).
Dapivirine was analysed using a Waters Symmetry Sheild
RP18 column, 250 mm × 4.6 mm (Waters, USA) and
acetonitrile and 5 mM Ammonium acetate buffer (pH 4.5)
in the ratio of 80:20 v/v as mobile phase with a detection
wavelength of 286 nm. Clotrimazole was analyzed using
a Waters Nova Pack C18 column, 150 × 3.9 mm (Waters,
USA) and 5 mM sodium dibasic phosphate buffer and
methanol in a composition of 25:75 v/v as mobile phase at
a wavelength of 262 nm.
2.4. Solubility Studies
Solubility studies of Dapivirine and Clotrimazole were
carried out by shaking an excess amount of each drug
with 3 ml of the following dissolution medium simulated
vaginal fluid, sodium phosphate buffer of pH 4 and 1%
Tween, sodium phosphate buffer pH 4 and 0.5% sodium
lauryl sulphate, sodium phosphate buffer pH4 and 1%
sodium lauryl sulphate, sodium phosphate buffer pH 4
and methanol (ratio 75:25), sodium phosphate buffer
pH4 and methanol (ratio 60:40), sodium phosphate
buffer pH4 and methanol (ratio 40:60). The samples were
incubated in a 37˚C water bath (JEIO TECH, Model:
BS-06/11/21/31, Korea) at 80 rpm for 24 hours. After 24
hours, the samples were centrifuged in Eppendorf cen-
trifuge (Model 5810 R, Germany) at 10,000 rpm for 10
minutes. The supernatants were collected and filtered
through 0.45 µm filters and injected into HPLC for ana-
lysis.
2.5. Dissolution Apparatus Design
A USP 8-Flask/8-pindle SR8-Plus (Hanson Research,
USA) dissolution tester was used. The USP dissolution
Aperture was modified with a “small volume” adapter kit
(p/n 72-800-721, Hanson Research), including 150 ml
flat-bottom flasks and mini paddles. The Enhancer Cell
(p/n 12-4000, VanKel, USA) as shown in Figure 1, con-
sisted of a cap, a washer-membrane, an O-ring, and an
adjustable drug reservoir. Specified volume of the de-
gassed in vitro release testing (IVRT) medium was placed
into each of the six vessels and the medium was equili-
brated to 37˚C ± 0.5˚C. Each tablet was placed in the
Enhancer cell cavity. Residual volume of the cell cavity
was filled with the medium (~2 ml). The cell cavity was
immediately covered with nylon mesh, O-ring and se-
cured with the cap. Each Enhancer cell was gently slid
into each of the vessel and the apparatus was operated at
the specified rotation speed. At each sampling point in-
dicated in Table 1, a syringe was fitted to the stainless
steel sampling probe to withdraw 2 ml of solution from a
zone midway between the surface of the medium and the
top of the paddle, not less than 1 cm from the vessel wall.
2 ml of the dissolution medium was replaced back to each
vessel after every sampling point. The sample solution
was filtered through a 13 mm × 0.45 μm nylon filter and
collected into HPLC vial for analysis.
The method was optimized by studying different vari-
ables:
2.5.1. Selecti o n of Dru gs
The two drugs used in this study were Dapivirine and
Clotrimazole. The selection of drugs was based on two
parameters:
1) Developmental Stage of the drugs (Novel vs Estab-
lished).
Figure 1. Enh a ncer cell.
Table 1. Dissolution conditions.
Media volume 100 ml, 150 ml
Temperature 37.0˚C ± 0.5˚C
Paddle Height 2.5 cm (Distance from paddle to membrane)
Paddle Rotation Speed 100, 75, 50 and 25 rpm
Sampling volume 2 ml with replacement
Sampling Time Point 0.25, 0.5, 1, 2, 3, 4, 6, 8 hours

Design and Development of an in Vitro Assay for Evaluation of Solid Vaginal Dosage Forms 292
Dapivirine is an investigational compound, being de-
veloped as a potential vaginal microbicide for the pre-
vention of HIV transmission in women. Clotrimazole is a
broad spectrum antimycotic agent widely used for vagi-
nal candidiasis. It is a well established marketed product
available in various dosage forms.
2) Formulation Design.
Dapivirine tablets have been prepared as novel rapidly
disintegrating, bioadhesive tablets expected to be retained
in the vaginal cavity for prolonged periods whereas,
Clotrimazole tablets were prepared as conventional re-
lease tablets.
2.5.2. In Vitro Rel ease Testing Media
Selection of the receptor medium was based on the solu-
bility and stability of the drug in that medium. The re-
ceptor medium must provide adequate stability and sink
conditions to ensure that the drug has sufficient solubility
in the receptor medium such that it does not affect the
release rate of the drug from the matrix. The vaginal pH
of healthy women of reproductive age is acidic (pH 4 - 5)
[17]. Hence sodium phosphate buffer (pH 4.0) was se-
lected as the media. Dapivirine and Clotrimazole being
poorly water soluble, solubility of these drugs were con-
ducted in the selected media with different concentra-
tions of surfactants and co solvents.
2.5.3. Medi a Vol ume
Two volumes were selected to study this parameter—100
ml and 150 ml. The drug selected for this study was
Dapivirine 0.5 mg tablets.
2.5.4. Stirri ng Speed
The tablets are not expected to undergo much agitation in
the vagina due to weak contractions occurring in that
area of the body. Hence, release studies of tablets were
conducted at low speed (25 rpm) and the results were
compared with the release profiles obtained at higher
speeds (50, 75, 100) to determine if speed can have a
significant effect on release rate of drugs. The drug se-
lected for this study was Dapivirine 0.5 mg tablets.
2.5.5. Dose Variability
The two doses of Dapivirine tablets (0.5 and 1.25 mg)
were used to study the effect of concentration on the re-
lease properties of the drug. The tablets were prepared
with the same manufacturing procedure, having similar
formulation characteristics.
2.5.6. Comp arison of Prepared and M arketed
Clotrimazole Tablets
The drug release profile of Clotrimazole tablet 100 mg
prepared in house was compared with that of commercial
Canesten (Clotrimazole tablet 100 mg) tablet, in order to
establish the ability of the method to detect differences in
the formulation characteristics.
2.6. Kinetics of Drug Release
To describe the kinetics of drug release from Dapivirine
(0.5 mg and 1.25 mg) and Clotrimazole formulations,
dissolution data obtained from drug release studies was
fitted to various mathematical models such as zero-order,
first order and Higuchi model. Korsmeyer-Peppas model
was also applied to analyze the mechanism of drug re-
lease.
3. Results and Discussion
A well designed dissolution testing is important not only
as a method for evaluation of a dosage form but also as a
potential tool to predict the in vivo drug release behavior
of the formulation. Currently, no official compendia spec-
ify a test to study the release pattern for vaginal formula-
tions. The main aim of this study was to develop an in
vitro dissolution procedure which is close to the physio-
logical conditions of vagina thus to be able to give some
predictive estimate of the in vivo drug release character-
istics.
3.1. Tablet Properties
Tablets of Dapivirine and Clotrimazole were prepared
successfully and complied with USP in respect to drug
content (98% - 102%) and having the physical character-
istics as described in Table 2. Dapivirine tablets, being
fast dissolving, disintegrated within 60 seconds.
3.2. Solubility Studies
Selection of IVRT medium was based on physiological
pH and solubility of the drug in that medium. In order to
have good sink conditions, it is important to select a me-
dia in which the solubility of the drug is at least 3 to 10
times its saturation solubility. Solubility of Dapivirine
and Clotrimazole was first determined in simulated
vaginal fluid (SVF) prepared according to Owen and
Katz [18]. However, low solubility results were observed
due to hydrophobic nature of the drugs. Hence, the solu-
bility of the drugs in phosphate buffer (pH 4) containing
various surfactants and alcohol as co-solvents at different
concentration was evaluated (Table 3).
3.3. The Design
Vaginal cavity usually contains very small volume of
fluid [18]. The cavity of the enhance cell creates and
mimics the situation when the formulation is placed in-
side the vaginal cavity as the tablet is allowed to disinte-
grate in small amount of fluid (2 ml) inside the enhancer
cell. Artificial membranes (synthetic and semi-synthetic)
have been recommended for i vitro quality control testing n

Design and Development of an in Vitro Assay for Evaluation of Solid Vaginal Dosage Forms 293
Table 2. Physical characteristics of Dapivirine 0.5 mg and 1.25 mg and Clotrimazole 100 mg tablets.
Parameters Dapivirine 0.5 mg Dapivirine 1.25 mg Clotrimazole 100 mg
Average weight (g) 1.0175 ± 0.003 1.010 ± 0.002 1.009 ± 0.12
Hardness (horizontal) (Kp) 11.028 ± 0.12 11.25 ± 0.42 14.015 ± 0.19
Hardness (vertical) (Kp) 7.93 ± 0.33 8.07 ± 0.50 11.39 ± 0.28
Thickness (mm) 5.36 ± 0.04 5.39 ± 0.01 4.95 ± 0.07
Friability (% w/w) 0.65 0.57 0.72
Disintegration time (min) 0.52 ± 0.05 0.58 ± 0.03 6.19 ± 0.10
Moisture content (% w/w) 2.54 ± 0.04 2.51 ± 0.02 1.25 ± 0.02
Table 3. Solubility data of Dapivirine and Clotrimazole in SVF and phosphate buffe r c ontaining surfactants and co-solvents.
Media Concentration of Dapivirine (µg/ml) Concentration of Clotrimazole (µg/ml)
Simulated Vaginal Fluid 0.53 21.30
Sodium Phosphate Buffer (pH 4.0) + 0.5% Tween 80 21.19 22.06
Sodium Phosphate Buffer (pH 4.0)+ 1.0% Tween 80 40.21 112.30
Sodium Phosphate Buffer (pH 4.0) + 0.5% SLS 36.82 494.27
Sodium Phosphate Buffer (pH 4.0) + 1.0% SLS 123.51 1338.60
Sodium Phosphate Buffer (pH 4.0):Methanol (75:25) 0.85 16.82
Sodium Phosphate Buffer (pH 4.0):Methanol (60:40) 7.18 102.08
Sodium Phosphate Buffer (pH 4.0):Methanol (40:60) 112.70 2323.11
due to the variability involved with biological mem-
branes. The membranes should be permeable to the drug
and its pore size and thickness should not affect the drug
release. The membrane selected should provide an inert
holding surface for the test formulation, and not act as a
barrier as vaginal microbicidal formulations are not de-
signed for systemic absorption but expected to act topi-
cally. The Nylon Mesh used during this entire experi-
ment served only as a holder first for the placed tablet
and then for the gel-matrix obtained after disintegration
of the tablet in the sample compartment in contact with
the receptor solution. The mesh size was small enough to
hold disintegrated tablet inside the sample cavity but still
permitted the receptor solution to easily circulate from
the 150ml dissolution vessel into the sample cavity of the
Enhancer cell, thus facilitating homogenous distribution
of the drug released from the formulation.
3.4. In Vitro Drug Release Studies
3.4.1. Selectio n of Media
Dapivirine and Clotrimazole are poorly water soluble
drugs. This necessitated the use of media with surfactant
or organic solvent. Based on the solubility data of the
drugs (Table 3), the two media selected to study the re-
lease profile were sodium phosphate buffer containing 1)
1% SLS and 2) 60% methanol. The release study was
conducted at a paddle rotation speed of 100 rpm and me-
dia volume was 100 ml. Figures 2 and 3 show compara-
tive release profiles of Dapivirine and Clotrimazole, re-
spectively in two media. Dapivirine release profile from
surfactant medium showed a sharp release in 15 minutes,
forming a plateau after 6 hours. However, profile in
aqueous organic medium demonstrated a slow and linear
rise in the drug release during the 8 hours study. This can
be explained by higher solubility of Dapivirine in surfac-
tant media (123.5 µg/ml) as compared to aqueous or-
ganic media (112.7 µg/ml). Due to high probability of air
bubbles with the use of surfactant media, aqueous or-
ganic media which shows a profile closer to the expected
in vivo drug release behavior was selected as the media
of choice. Previous reports [19,20] on Dapivirine have
used aqueous organic media for drug release studies. On
the other hand, Clotrimazole release in aqueous organic
media was higher as compared to that in surfactant media,
which is attributed to the drug’s higher solubility in
queous organic media (2323 µg/ml). a

Design and Development of an in Vitro Assay for Evaluation of Solid Vaginal Dosage Forms 294
Figure 2. Dapivirine release profile in aqueous organic and surfactant media.
Figure 3. Clotrimazole release profile in aqueous organic and surfactant media.
3.4.2. Medi a Vol ume
Figure 4 shows the effect of the media volume on the
release of Dapivirine tablets. The study was conducted
using 100 ml and 150 ml of dissolution media. As can be
seen from the Figure 4, the percentage drug release in
the two volumes (30.6% in 100 ml and 27.4% in 150 ml
after 8 hours) was not markedly different from each other;
100 ml was selected as the volume for further studies.
Vaginal cavity usually contains very small volume of
fluid [18]. The lowest volume in 150 ml flask which
could be selected was 100 ml. During conducting the
dissolution studies, it is important that the sink condition
is maintained. These parameters were considered in se-
lection of the dissolution volume in our study. After 8
hours, Dapivirine release in 100 ml of media was ap-
proximately 30% (1.6 µg/ml), indicating that the sink
conditions were maintained as the saturation solubility of
Dapivirine in the receptor medium was 10 times higher
than the maximum concentration detected during the 8
hours of drug release study. Thus with the sink condi-
tions present, it is more likely that dissolution results will
reflect the properties of the dosage form [6].
3.4.3. Stirri ng Speed
In order to simulate the physiological conditions of weak
contractions in vagina, release rate of Dapivirine tablet
was studied at the lowest allowable speed of rotation in a
standard USP apparatus-25 rpm. The percent drug re-
lease at 2 and 6 hours were compared for different speeds
of rotation (Table 4). As can be seen from the table,
maximum drug release was observed with 100 rpm rota-
tional speed. This can be attributed to the fact that an
increase in agitation leads to a reduction in the thickness
of the diffusion layer formed at the interface between the
receptor and the membrane and facilitates better mixing
[14]. The release at 100 rpm was approximately 1.5 times
higher than at 25 rpm. However, the tablets at 25 rpm
showed a comparable drug release rate to 50 or 75 rpm
and were closer to physiological conditions of weak con-
tractions; therefore 25 rpm was selected as the speed of
choice.
3.4.4. Comp arison of Prepared and M arketed
Clotrimazole Tablets
This study was conducted to compare the release profiles
of marketed Canesten tablets with the prepared Clotri-
mazole tablets. The drug release profiles in Figure 5
shows an initial higher release rate (5.5%) of the drug
from Canesten as compared to that from in house Clotri-
mazole tablets (1.0%). This e due to the variation may b

Design and Development of an in Vitro Assay for Evaluation of Solid Vaginal Dosage Forms295
Figure 4. Comparison of Dapivirine release in different media volumes.
Figure 5. Release profile comparison of different formulations of Clotrimazole.
Table 4. Comparison of Dapivirine release after 2 and 6
hours at different speeds of rotation.
Paddle Speed
(rpm)
Release in 2 hours
(%)
Release in 6 hours
(%)
25 14.7 23.6
50 20.2 29.9
75 18.6 27.8
100 21.6 36.0
in the disintegration time of the two products. The disin-
tegration time of the two formulations when measured
using standard BP Disintegration apparatus, was found to
be 6 - 7 minutes for prepared Clotrimazole tablets and 45
seconds for Canesten. However, after a period of time the
release profiles of the marketed Canesten and the pre-
pared clotrimazole tablets became comparable. Thus, it is
evident that the method is sensitive enough to differenti-
ate variability in the method of manufacturing.
3.4.5. Dose Variability
This parameter was based on release studies conducted
on two different doses of Dapivirine (0.05% and 0.125%),
in order to assess the sensitivity of the developed disso-
lution method (Figure 6). The dissolution conditions
were as reported in Table 1. From Figure 6, it can be
seen that the shape of the two release profiles is similar
showing a higher release from the higher dose, indicating
the capability of the method to differentiate the variabil-
ity in dose.
3.5. Comparison of Release Profile from
Enhancer Cell and Standard USP
Dissolution Method
The drug release study was conducted using the standard
dissolution method described in official compendia [4-6]
for solid oral dosage forms. The release profile from
standard method was compared with that from Enhancer
cell method. Dapivirine 0.5 mg tablet was employed in
this study and 500 ml of 0.1 N HCL was used as media.
As can be seen from Figure 7, Enhancer cell method
shows a slow release as only 20% of drug is released
within 2 hours. On the other hand, the release rate of
drug studied in standard dissolution apparatus is high as
80% of drug is released within 2 hours.
With the standard USP dissolution method, employing

Design and Development of an in Vitro Assay for Evaluation of Solid Vaginal Dosage Forms 296
Figure 6. Comparison of Dapivirine release from two doses (0.5 mg and 1.25 mg).
Figure 7. Comparative release profile of Dapivirine from standard and enhancer cell method at 50 rpm.
500 ml of media and 50 rpm rotational speed, the rapidly
disintegrating tablets of Dapivirine dispersed quickly in
the large media volume giving a solubility profile of the
drug rather than predicting its release characteristics.
These tablets were designed to disperse quickly and form
a gel in small amount of fluid in the vagina. Enhancer
cell has the advantage of containing the tablet in a pocket
wherein it is allowed to disperse in small volume (2 ml)
of media and form a bioadhesive gel from which the drug
is released slowly. Comparative release studies of Da-
pivirine from Enhancer cell and standard method at 50
rpm clearly show that the shape of the two profiles is
same. The only difference is the rate of release of the
drug which corresponds to the difference in the volume.
3.6. Kinetics and Mechanism of Drug Release
The release data of Dapivirine 0.5 mg and 1.25 mg were
fitted best to Higuchi’s model (R2 = 0.989 and 0.992 re-
spectively) as indicated in Figures 8 and 9. This suggests
that the drug release is controlled by diffusion. When the
release data was fitted to the model of Korsmeyer-Peppas
(Table 5), the release exponent n value was found to be
close to 0.5 for both the doses thus indicating Fickian
diffusion. The drug release data of Clotrimazole tablets
was best fitted to zero-order model with an R2 value of
0.9957. This was confirmed when the data was applied to
Korsmeyer Peppas model. The value of release exponent
n was found to be close to 1.0 (Table 5) indicating zero-
order release.
4. Conclusions
A dissolution system was designed and optimized for
evaluation of vaginal tablets. The system enabled con-
tainment of the drug matrix in the cell to avoid complete
distribution and dissolution of the drug in the medium.
Drug release studies were successfully conducted on
Dapivirine and Clotrimazole tablets. The suitability of
the system was investigated for various parameters like
different formulation design, dose variability, different
stirring speeds, media volume and comparison of pre-
pared and marketed product. The Enhancer cell method
was found to be sensitive enough to distinguish between
formulations and demonstrated the ability to detect
products of different strengts. In conclusion, a simple, h

Design and Development of an in Vitro Assay for Evaluation of Solid Vaginal Dosage Forms297
Figure 8. Dapivirine 0.5 mg release per cm2 versus square root of time.
Figure 9. Dapivirine 1.25 mg release per cm2 versus square root of time.
Table 5. Model fitting using Korsmeyer-Peppas equation.
Product R2 n k
Dapivirine 0.5 mg 0.9272 0.6907 0.4090
Dapivirine 1.25 mg 0.9161 0.4742 1.6420
Clotrimazole 100 mg 0.9624 0.9122 0.0850
reliable, and reproducible dissolution method has been
developed for solid vaginal dosage forms. This method
can be used to support formulation development for early
stage drug product development and potentially for qual-
ity control and in vivo performance analysis of the final
product.
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