J. DRUG DEL. SCI. TECH.

, 22 (4) 347-352 2012

Novel formulation and clinical evaluation

of nalidixic acid ointment in impetigo
S.S. Tous1, A.M. El Sayed1, M.G. Abd El Mohsen1, E.M.K. Youssef2, M.N. Agban3, M.F. Boushra1*

Pharmaceutics Department, Faculty of Pharmacy, Assiut University, 71526 Assiut, Egypt

1

2
Dermatology, Venerology & Andrology Department, Faculty of Medicine, Assiut University, 71526 Assiut, Egypt
3
Microbiology and Immunology Department, Faculty of Medicine, Assiut University, 71526 Assiut, Egypt
*Correspondence: mariamceutics84@yahoo.com

The aim of this work was to formulate and evaluate nalidixic acid for the first time in different topical ointment bases. Among different oint-
ment formulations, hydrocarbon base achieved the lowest release, while water-soluble base achieved the highest release. The presence of both
isopropanol as cosolvent and nalidixic acid-sodium benzoate solid dispersion in formula (W8) enhanced both the release and the antibacterial
activity of nalidixic acid compared to ointment formula (W5) containing neither of them. A stability study was also performed where no significant
change in pH or drug content was observed in all stored formulations (W5 and W8). Stability was further checked by thin-layer chromatography
(TLC). After clinical application in impetigo, it was found that the presence of isopropanol and nalidixic acid-sodium benzoate solid dispersion
in ointment formulation (W8 ) caused a significant reduction in the mean time of healing (only four days) in impetigo patients.

Key words: Nalidixic acid – Ointment – Release – Stability – Impetigo.

The delivery of drugs through the skin has long been a promising (Egypt). Isopropyl myristate was obtained from Merck Schichardt
concept because of ease of access, large surface area and noninvasive, (Germany) and Span 60 from Fluka Chemical, Buch (Switzerland).
painless nature of the treatment, whether the desired bioavailability
is systemic or local [1]. It is also simple to terminate the therapy if 2. Methodology
any adverse or undesired effect occurs [2]. Topical application of 2.1. Preparation of nalidixic acid ointments
antimicrobial agents is a useful tool for the therapy of skin infections. In all preparations, nalidixic acid was sieved through mesh no.
Firstly, it can avoid unnecessary exposure to gut flora which may 100. Then the calculated amount of the drug was incorporated by the
exert selection for resistances. Secondly, it is expected that the high mechanical method, after congealing of the base, in order to obtain
local drug concentration in topical application will be able overwhelm 1 % ( w/w) of nalidixic acid topical formulations.
many mutational resistances. Finally, topical application is less likely
to cause side effects compared to systemic therapy [3]. However, the 2.1.1. Preparation of hydrocarbon (H) ointment base
emergence of resistant microorganisms has led to failure in the treat- The composition of different hydrocarbon ointment bases is
ment of different skin diseases by the available antimicrobial agents shown in Table I. Hydrocarbon ointment bases were prepared by fu-
[4], therefore the search for new topical antibacterial agents is a must. sion method [7] where white soft paraffin (formula H1) and beeswax
The discovery of nalidixic acid or 1-ethyl-1,4 dihydro-7-methyl-4 (formula H2) were melted on a water bath. Liquid paraffin (formulae
oxo-1,8 naphthyridine-3 carboxylic acid in 1962, and its introduction H3 and H4) was then added to the melt and stirred till congealing.
into clinical use in 1967, marked the beginning of five decades of
quinolone development and use [5]. Nalidixic acid is available in the 2.1.2. Preparation of absorption (A) ointment base
market in the oral tablet and oral suspension form. The usual adult Table II demonstrates the composition of different absorption
dose is 4 g daily by mouth in 4 divided doses for at least 7 days in
acute infection [6]. Since no topical formulations of nalidixic acid are Table I - Composition of different hydrocarbon (H) ointment bases
containing 1 % (w/w) nalidixic acid.
yet available in the market, it is of tremendous benefit to formulate it
topically and make benefit of its antibacterial activity in the treatment Ingredient Percent of ingredient of each formulation
of skin infections like impetigo. (w/w)
The aim of this work was to formulate nalidixic acid for the first H1 H2 H3 H4
time in different ointment bases and evaluate its in vitro release char-
White soft paraffin 100 95 85 75
acteristics and antibacterial activity. Then the selected formulations Beeswax - 5 5 5
were further investigated for their clinical efficacy in patients with Liquid paraffin - - 10 20
impetigo.
Table II - Composition of different absorption (A) ointment bases con-
I. EXPERIMENTAL taining 1 % (w/w) nalidixic acid.
1. Materials
Nalidixic acid was kindly provided by Memphis Co. (Egypt). Ingredient Percent of ingredient of each formulation
(w/w)
Standard cellophane membrane (molecular cut of range = 12000) was
purchased from Sigma Chem. Co. (United States). White soft paraffin, A1 A2 A3
liquid paraffin, wool fat, hard paraffin, beeswax, sodium carboxymethy Wool fat 5 6 10
cellulose (Na-CMC), cetostearyl alcohol, stearic acid, potassium hy- Hard paraffin 5 - 5
droxide, sodium lauryl sulfate, Tween 80, n-butyl alcohol, and borax Cetostearyl alcohol 5 0.5 5
were obtained from Adwic, El-Nasr Pharmaceutical Chemicals Co. White soft paraffin to 100 100 100

347
J. DRUG DEL. SCI. TECH., 22 (4) 347-352 2012
S.S. Tous, A.M. El Sayed, M.G. Abd El Mohsen, E.M.K. Youssef, M.N. Agban, M.F. Boushra
Table III - Composition of different water soluble (W) ointment bases
containing 1% (w/w) nalidixic acid.
Ingredient Percent of ingredient of each formulation
(w/w)
W1 W2 W3 W4 W5
PEG 4000 60 50 40 47.5 40
PEG 400 10 20 30 47.5 60
Water 20 20 20 - -
Isopropyl myristate 10 10 10 - -
Cetostearyl alcohol - - - 5 -
ointment bases. All ingredients of the base were melted, mixed and
triturated till congealing.
2.1.3. Preparation of water soluble (W) ointment base
The ingredients used in the preparation of water soluble ointment
base are given in Table III. The ointments were prepared by mixing
accurately weighed PEG 400, PEG 6000 (formula W5) and cetostearyl
alcohol (formula W4) and heating to melting (about 50 °C). The water
and isopropyl myristate (formulae W1, W2 and W3) were previously
heated to about 37-40  °C and then added under magnetic stirring (Gal-
lenkamp, United Kingdom) to the melt [8]. Stirring was continued on
cooling till obtaining a homogenous ointment.
2.2. Evaluation of different nalidixic acid topical preparations
2.2.1. Cosmetic and aesthetic criteria
Physical appearance of different formulations was observed visu-
ally while spreadability was observed by spreading 1 g of formulation
on a clean even glass surface [9].
2.2.2. pH measurement
A digital pH meter (Jenway 3505, United Kingdom) was used to
determine the pH of the prepared formulations [10, 11]. The results
are the mean of three determinations.
2.2.3. Viscosity measurement
The viscosity of the different formulations was determined using
Brookfield DV-ІІІ Viscometer [12]. (Brookfield Engineering Labo-
ratories, Inc., United States). T-bar spindle was used to measure the
viscosity and was rotated at 25 rpm. The average reading of three
determinations was recorded.
2.2.4. Determination of drug content
An accurately weighed amount of each formulation was dissolved
in 5 mL 0.1 N NaOH, filtered using a nylon membrane filter disk
(0.45 μm). Then they were suitably diluted and assayed spectrophoto-
metrically (Jenway 6305, United Kingdom) for drug content at λmax
(257 nm) against a similarly treated blank.
2.3. In vitro release study
2.3.1. In vitro release of nalidixic acid from different topical
formulations
The in vitro release of the drug from different formulations was
studied by dialysis method using cellophane membrane under sink
conditions. Of each formulation, 500 mg was placed on a circular area
of cellophane membrane previously soaked in the receptor medium
for at least 30 min. The loaded membrane was then firmly stretched
using a rubber band over one of the open ends of a glass tube with a
surface area of 1 cm2. The tube was then immersed in 250 mL beaker
containing 25 mL of phosphate buffer pH 7.4 serving as a receptor
media. The receptor medium was kept at 32 °C to reflect the usual
skin temperature [13] in a thermostatically controlled water bath
(Labortechik, Germany) adjusted at 25 stroke/min. At predetermined
time intervals of 30, 60, 90, 120, 150 and 180 min, aliquots of 2 mL
Novel formulation and clinical evaluation of nalidixic acid ointment in impetigo
Table IV - Composition of nalidixic acid water soluble (W) ointment
formulations containing cosolvent and/or solid dispersion.
Ingredient Percent of ingredients of each formulation
(w/w)
W6 W7 W8
Nalidixic acid 1 - -
Nalidixic acid-sodi- - equivalent to equivalent to
um benzoate solid 1 % drug 1 % drug
dispersion (1:8)
Isopropanol:water 1 - 1
30% (v/v)
Water soluble oint- 100 100 100
ment base (W5) to
were withdrawn from the receptor media and immediately replaced by
equal volume of fresh phosphate buffer kept at the same temperature.
Aliquots at different time intervals were assayed spectrophotometri-
cally at (257 nm) for drug content against a suitable blank and the
cumulative amount of drug released was calculated. The results are
the mean of three determinations.
2.3.2. Effect of cosolvency and solid dispersion on nalidixic acid
release
In a previous work [14], nalidixic acid solubility was carefully
investigated. Among different cosolvents, it was found that 30 %
(v/v) isopropanol as a cosolvent gave the highest rise in nalidixic
acid solubility. For solid dispersion, nalidixic acid-sodium benzoate
solid dispersion in ratio 1:8 had enhanced nalidixic acid solubility best
therefore they were both selected for incorporation into the selected
formulations.
Nalidixic acid solid dispersion was prepared by common solvent
method in which both nalidixic acid and sodium benzoate (1:8 ratio)
were dissolved in a minimal amount of 25 %(v/v) ammonia solution
under continuous stirring. The solvent was then allowed to evaporate
at room temperature and the mass obtained was then crushed, pulver-
ized, and sifted through mesh No. 100(76) [15].
Among all prepared ointment formulations, formula (W5) gave the
highest release. Therefore, it was reformulated, as shown in Table IV,
to investigate the effect of both cosolvency and solid dispersion on
nalidixic acid release.
2.4. Antibacterial activity study
The antibacterial activity of the selected nalidixic acid ointment
formulations against various strains of microorganisms was evalu-
ated by standard cup plate method [16]. The antibacterial activity
was evaluated by measuring the diameter of zones of inhibition (in
mm). The inhibition zone diameters were measured with the help of
zone reader and the results are the mean of three determinations [16].
Bacillus subtilis, Staphylococcus aureus, Pseudomonas aeruginosa
and Proteus mirabilis were used in this study. The influences of both
cosolvency and solid dispersion on the antibacterial activity of the
drug in the selected formulations were also investigated.
2.5. Stability study
Selected ointment formulations were stored at room temperature
for six months [17]. Physical evaluation of the samples’ stability
was carried out by visual inspection. Stability was evaluated by pH
measurements and spectrophotometric analysis of the drug content
[17, 18]. The effects of both cosolvency and solid dispersion on the
stability of the selected formulations were also investigated. Thin
layer chromatography (TLC) technique was also used to study the
stability of nalidixic acid [19]. Nalidixic acid was extracted from
stored formulations at the end of storage interval (6 months) using 0.1
348
Novel formulation and clinical evaluation of nalidixic acid ointment in impetigo J. DRUG DEL. SCI. TECH., 22 (4) 347-352 2012
S.S. Tous, A.M. El Sayed, M.G. Abd El Mohsen, E.M.K. Youssef, M.N. Agban, M.F. Boushra

N NaOH. The extracts were then filtered and diluted with methanol. Table V - pH, viscosity and drug content of different nalidixic acid oint-
Two microlitre volumes of each solution were spotted on TLC plates ment bases.
as well as 2 μL of adequate freshly prepared solution of stock nalidixic Formulation pH Viscosity Drug content
acid. Chromatograms were developed using methanol:n-butyl alco- code (cps) (% w/w)
hol:25 % ammonia [20]. The plates were dried at room temperature H1 5.5 ± 0.12 62300 ± 10 96.21 ± 0.65
and viewed under UV (at λ = 257 nm). The values of the retention H2 5.8 ± 0.11 71600 ± 15 97.53 ± 0.22
factor (RF) of the analyzed solutions were calculated as the mean H3 5.8 ± 0.09 56900 ± 12 98.23 ± 0.34
of three determinations and compared to that of nalidixic acid stock H4 5.7 ± 0.8 44060 ± 11 98.34 ± 0.45
solution [20]. A1 6.1 ± 0.13 139030 ± 23 96.65 ± 0.57
A2 6.3 ± 0.14 146020 ± 14 95.92 ± 0.67
2.6. Clinical application in impetigo A3 6.8 ± 0.11 150400 ± 20 96.46 ± 0.43
2.6.1. Patients W1 6.4 ± 0.15 76050 ± 12 97.31 ± 0.78
The protocol was approved by the Medical Ethics Committee of W2 6.3 ± 0.11 72030 ± 11 98.24 ± 0.56
the Faculty of Medicine, Assiut University. The clinical study was W3 6.7 ± 0.08 61000 ± 13 97.45 ± 0.59
initiated on 40 patients in the Department of Dermatology, Assiut W4 6.9 ± 0.12 55100 ± 15 98.52 ± 0.53
W5 7.2 ± 0.09 52050 ± 14 98.93 ± 0.61
University Hospital. However, only 30 patients followed-up regularly.
W6 7.2 ± 0.16 51556 ± 11 98.5 ± 0.66
Both male and female patients were involved in the clinical study,
W7 7.1 ± 0.18 51100 ± 20 98.92 ± 0.58
the age of whom ranged from 2 years to 12 years old. The diagnosis W8 7.1 ± 014 50300 ± 20 98.95 ± 0.54
of impetigo was mainly a clinical one, but further confirmation was
done by microbiological investigation of skin swap.
visual appearance and odor, acceptable cosmetic criteria, suitable
2.6.2. Study design consistency and ease of spreadability.
A randomized, placebo controlled, double blind study was per-
formed. The patients were divided into three groups; each consisted of 1.2. pH measurement
ten patients, as follows: Group I: patients received ointment containing The pH of different nalidixic acid ointment bases was determined
1  % nalidixic acid (W5). Group II: patients received ointment contain- immediately after preparation (Table V). It was found that the pH of
ing isopropanol and nalidixic acid-sodium benzoate solid dispersion different formulations lied in the range of 5.5-7.2 (near the normal
equivalent to 1 % drug (W8). Group ІІІ: patients received placebo pH of the skin) and therefore no skin irritation is expected upon the
ointment. application of any of them.

2.6.3. Treatment strategy 1.3. Viscosity measurement

All patients were instructed to apply the formulation twice daily The rheological properties are thought to play an important role in
for 7-14 days. Follow-up and close supervision was done whenever the release of drug from different formulations. Therefore, the viscosity
possible during the treatment course in order to assess any signs of of different freshly prepared nalidixic acid topical formulations was
healing. determined and listed in Table V.

2.6.4. Clinical assessment 1.4. Determination of drug content

Efficacy was assessed by clinical investigation of the patients,
counts of lesions and crusts, and microbiological isolation and iden-
tification of bacteria at baseline and at the end (day 7 or 14) of the
study. Colored photographs were taken before and after treatment in
order to assess the degree of response to the therapy. Clinical outcome
was scored as excellent, good or poor and patients were classified into
groups according the mean time of healing.
2.6.5. Patient compliance
Patients were monitored for any adverse effects during the treat-
ment course. Any problem in drug application was also reported in
order to ensure the highest degree of patient comfort and compliance.
3.7. Statistical analysis
Experimental results were expressed as mean ± SD. Student’s
t-test and two-way analysis of variance (ANOVA) were applied to
check significant differences in the obtained results. Differences were
considered to be statistically significant at p < 0.05 (where p < 0.05:
mild significance, p < 0.01: moderate significance and p < 0.001: high
significance). The statistical software used for analysis was GraphPad
Prism Software Version 5.0 (GraphPad Prism Software, San Diego,
CA, United States) [21].
ІІ. RESULTS AND DISCUSSION
1. Evaluation of different nalidixic acid topical
preparations
1.1. Cosmetic and aesthetic criteria
All prepared nalidixic acid ointment formulations showed good
The drug content of different nalidixic acid topical formulations
(Table V) had ranged from 96.21 to 98.95 (% w/w). This high drug
content indicates that drug is uniformly distributed in different prepared
formulations.
2. In vitro release study
2.1. In vitro release study of nalidixic acid from different
ointment bases
Nalidixic acid release from different hydrocarbon (H) ointment
bases is illustrated in Figure 1. The release can be arranged as follows
H4 > H3 > H1 > H2. It is obvious that the release pattern was depend-
ent on the composition and the viscosity of the hydrocarbon ointment
base where the presence of beeswax in formula (H2) significantly (P
< 0.001) decreased the release of drug compared to formula (H1).
While the addition of liquid paraffin in formulae (H3) and (H4) sig-
nificantly (P < 0.001) increased the release of nalidixic acid. Formula
(H4) containing 20 % (w/w) liquid paraffin showed the highest release
among different hydrocarbon ointment bases. This may be due to the
resultant decrease in viscosity upon the addition of liquid paraffin to
the base leading to an increase in drug diffusion. Formula (H2), with
the highest viscosity among all prepared formulations, achieved the
lowest release.
The release of nalidixic acid from different absorption (A) bases
(formulae A1, A2, A3) is shown in Figure 2. Both illustrate that no
significant difference existed between the release of drug from dif-
ferent absorption ointment bases.
Figure 3 shows the release of nalidixic acid from water soluble
(W) ointment bases (formulae W1, W2, W3, W4, and W5). The dif-

349
J. DRUG DEL. SCI. TECH., 22 (4) 347-352 2012 Novel formulation and clinical evaluation of nalidixic acid ointment in impetigo
S.S. Tous, A.M. El Sayed, M.G. Abd El Mohsen, E.M.K. Youssef, M.N. Agban, M.F. Boushra

Cumulative amount released (µg)

Cumulative amount released (µg)

140 W5
2000 W6
120 1800 W7
1600 W8
100 1400
80 1200
1000
H1
60 800
H2
40 600
H3
400
20 H4
200
0 0
0 30 60 90 120 150 180 0 30 60 90 120 150 180 210
Time (min) Time (min)

Figure 1 - In vitro release of nalidixic acid from different hydrocarbon Figure 4 - In vitro release of nalidixic acid from water soluble (W) oint-
(H) ointment bases. ment bases containing cosolvent and or solid dispersion.

isopropanol. It displayed significantly (P < 0.001) higher release

Cumulative amount released (µg)

compared to formula (W5) containing no cosolvent. These results

140

120

100 are expected as cosolvents simply function to produce a saturated

or nearly saturated solution of the active ingredients and thereby
A1
80
A2

maximizing the thermodynamic activity of the penetrant [24]. It was

A3
60

40

20 reported that the addition of 10 % (v/v) of ethanol to emulsion as well

0
0 30 60 90 120 150 180
as oleaginous ointment bases of oxolinic acid increased its release
mainly because of the solubilizing effect [23] as it is expected that
Time (min)

Figure 2 - In vitro release of nalidixic acid from different absorption increasing the solubility of the drug in the vehicle plays an important
(A) ointment base. role in its release promotion[26]. Further confirmation was obtained by
Aukunuru et al. [27] who used propylene glycol and PEG to enhance
the solubility of ibuprofen in ointment bases and accordingly enhance
the release of the drug.
Cumulative amount released (µg)

900
800
700
It is well known that improvement of drug release from ointment,
creams and gels is one of the most important applications of solid dis-
600 W1
500 W2
W3

persion technique [28]. Therefore, formula (W7) contained nalidixic

400 W4
300 W5

200
100
0
0 30 60 90
Time (min)
Figure 3 - In vitro release of nalidixic acid from different water soluble
(W) ointment bases.
ference between the released amount of drug from these bases was
statistically significant (P < 0.001) and can be arranged as follows:
W5 > W4 > W3 > W2 > W1. Drug solubility appeared to be mainly
related to PEG content in the ointment [8]. Hence formula (W5),
containing the highest percent of PEG 400, showed the best release
among different water-soluble ointment bases. In addition, increasing
the concentration of PEG 400 in the formulation resulted in a decrease
in the viscosity values and therefore increased the release of the drug
from the base. These results are in agreement with those obtained by
Ismail et al. [22] who found that increasing the percentage of PEG
400 in water-soluble vehicle increased the release and attributed that
to the decrease in the viscosity values of the base.
Among different ointment bases, hydrocarbon base achieved
the lowest release.This can be attributed to the hydrophobic nature
and the low water solubility of the drug which favors its retention in
the base rather than passing into the aqueous solution [23]. On the
contrary, water-soluble base achieved the highest release. This may
be due to rapid dissolution of PEG and, consequently, giving more
chance for drug to be released [24]. The diffusion of water through
the cellophane membrane and formation of water-polyethylene glycol
solution resulting in an increase of drug solubility may also explain
these results [25].
These results are further confirmed with those obtained with
oxolinic acid, a quinolone drug, where the best release was provided
by the water-soluble PEG base and its lowest release was exhibited
by the oleaginous base [23].
2.2. Effect of cosolvency and solid dispersion on nalidixic acid
release
The effect of cosolvency alone on nalidixic acid release (Figure 4)
is illustrated in formula (W6) which contained 1 mL of 30 % (v/v)
acid- sodium benzoate solid dispersion (1:8) instead of the drug itself
120 150 180 210 showed significantly (P < 0.001) greater release pattern compared to
formula (W5) containing the drug alone.
These results are in agreement with improved drug release found
with topical formulations containing ketoconazole in the form of a
solid dispersion with polyvinyl pyrollidone compared to the drug itself
[29]. Also in vitro release of aceclofenac solid dispersion incorporated
gels was greatly improved when compared to that of aceclofenac incor-
porated gels [30]. This may be ascribed to enhancing drug solubility
and increasing the overall amount of drug released from the topical
formulation [29].
The combined effect of both cosolvency and solid dispersion
on nalidixic acid release was investigated in formula (W8) which
demonstrated a higher release compared to formulae (W6 and W7).
This may be attributed to the greater solubilizing effect achieved by
combining both cosolvency and solid dispersion. Muela et al. [31]
obtained similar results where the combination of solid dispersions
with cosolvents increased the water solubility of thiabendazole to a
larger extent than each method separately.
3. Antibacterial activity study
The antibacterial activity of selected nalidixic acid ointment
formulation (W5) was assessed using agar plate diffusion method.
The inhibition zone diameters (mm) of different nalidixic acid topical
preparations are illustrated in Table VI. It is obvious that the obtained
results are in agreement with those of the in vitro release study. In
other words, formula (W5), which achieved a higher release rate,
also showed a larger inhibition zone. Similar results were obtained
by Malipeddi et al. [11] who studied the formulation of norfloxacin in
different ointment and gel bases and found a direct correlation between
the in vitro release of the drug and the inhibition zone diameter. It is
worth mentioning that the presence of cosolvency and solid dispersion
in formula (W8) had a larger inhibition zone and greater antibacterial
activity compared to formulae (W5) containing neither of them.
4. Stability study
The selected formulations (W5 and W8) were stored for 6 months
at room temperature, inspected visually and assayed monthly for drug

350
Novel formulation and clinical evaluation of nalidixic acid ointment in impetigo J. DRUG DEL. SCI. TECH., 22 (4) 347-352 2012
S.S. Tous, A.M. El Sayed, M.G. Abd El Mohsen, E.M.K. Youssef, M.N. Agban, M.F. Boushra

Table VI - The inhibition zone diameter of selected nalidixic acid topical 5. Clinical application in impetigo
formulations. The clinical outcome of some treated patients is demonstrated in
Formula- Inhibition zone diameter (mm) Table VII. The results of the table can be statistically analyzed accord-
tion code Microorganism
ing to the mean time of healing (days) of each formulation.
The presence of cosolvent and solid dispersion in ointment for-
S. aureus P. mirabilis P. aerugi- B. subtilis mulations (W8) caused a significant reduction in the mean time of
nosa
healing of patients compared to those patients treated with formulations
W5 15 ± 0.45 23 ± 1.00 22 ± 1.12 24 ± 1,16 containing neither of them (W5).
W6 19 ± 1.12 28 ± 2.00 29 ± 1.53 27 ± .98 Accordingly, the effectiveness of the tested formulations can be
W7 27 ± 1.62 28 ± 2.23 30 ± 2.13 30 ± 1.13 classified according to the mean time of healing as shown in Table VIII.
W8 30 ± 2.12 32 ± 1.55 33 ± 2.43 35 ± 2.56 All the previously mentioned data demonstrate generally the ef-
fectiveness of nalidixic acid topical formulations and specially that of
ointment containing isopropanol and nalidixic acid-sodium benzoate
solid dispersion in the treatment of impetigo. It is worth mentioning
that none of the tested formulations engendered adverse effects in any
of the patients. In addition, the short period of treatment and the ease
of application increased patient compliance and adherence to treat-
ment. Colored photographs (Figure 6) were taken for patient before
(e) and after treatment with nalidixic acid in order to assess the degree
(d) of response to the therapy.
(c)
(b)
(a)

Figure 5 - TLC chromatogram showing: (a) RF value of nalidixic acid

stock solution; (b) RF value of nalidixic acid extracted from fresh ointment
(W5); (c) RF value of nalidixic acid extracted from stored ointment (W5);
(d) RF value of nalidixic acid extracted from fresh ointment (W8); (e)
RF value of nalidixic acid extracted from stored ointment (W8). *Each Figure 6 - Left: multiple honey crusts of 8-year-old female child with
spot is represented by three lines. scalp impetigo (before treatment). Right: the same child after treatment
for 7 days with ointment containing isopropanol and nalidixic acid-sodium
benzoate solid dispersion (W8).
content and pH. No significant change in pH or drug content was ob-
served and all stored formulations retained acceptable cosmetic and Table VIII - Time of healing after treatment with different tested for-
aesthetic criteria. The stability of the stored formulations was further mulations.
checked by TLC. The chromatogram (Figure 5) showed no significant
Formulation Time of healing Mean time of
difference between RF values of nalidixic acid extracted from stored (day) healing (day)
formulations and that of nalidixic acid stock solution (54 mm) as
reported by Hopkala and Kowalczuk [19].This strongly indicated that Ointment containing drug alone 7-14 12.4
(W5)
nalidixic acid is stable in the selected formulation under the storage
conditions. All the obtained results had strongly recommended the Ointment with cosolvent* and 7-10 8.8
selection of water soluble ointment base (W5 and W8) for clinical solid dispersion** (W8)
application on patients with impetigo. *Isopropanol. **Nalidixic-acid sodium benzoate solid dispersion.

Table VII - Clinical assessment of some representative impetigo patients treated with different formulations.
Patient groups Patient Duration of treat- Outcome
treated with differ- ment (days)
Age (years) Gender Impetigo type Severity
ent formulations
Group I: 2 F Non-bullous Severe 10 Good
ointment contain- 10 F Non-bullous Severe 14 Excellent
ing drug alone 7 F Non-bullous Mild 14 Excellent
(W5) 6 F Non-bullous Severe 10 Good
4 M Non-bullous Severe 14 Good
Group II: 3 F Non-bullous Severe 10 Excellent
ointment contain- 8 F Non-bullous Severe 7 Excellent
ing cosolvent* and 10 F Non-bullous Severe 10 Good
drug solid disper- 7 M Non-bullous Moderate 7 Excellent
sion** (W8) 2 M Non-bullous Severe 10 Excellent
Group III: 4 F Non-bullous Mild >14 No improvement
placebo ointments 12 F Non-bullous Mild >14 No improvement
and gels 11 M Non-bullous Mild >14 No improvement
9 M Non-bullous Mild >14 No improvement
6 M Non-bullous Mild >14 No improvement
*Isopropanol. **Nalidixic-acid sodium benzoate solid dispersion.

351
J. DRUG DEL. SCI. TECH., 22 (4) 347-352 2012
S.S. Tous, A.M. El Sayed, M.G. Abd El Mohsen, E.M.K. Youssef, M.N. Agban, M.F. Boushra
REFRENCES
1. Daniels R., Knie U.- Galenics of dermal products, Vehicles
properties and drug release. - JDDG, 5, 367-381, 2007.
2. Özgüney I.S., Karasulu H.Y., Kantarcı G., Sözer S., Güneri T.,
Ertan G. - Transdermal delivery of diclofenac sodium through
rat skin from various formulations. - AAPS Pharm. Sci. Tech.,
7, 1-7, 2006.
3. Yamakawa T., Mitsuyama J., Hayashi K. - In vitro and in vivo
antibacterial activity of T-3912, a novel non fluorinated topical
quinolone. - J. Antimicrob. Chemother., 49, 455-465, 2002.
4. Silver L.L., Bostian K.A. - Discovery and development of new
antibiotics: the problem of antibiotic resistance. - Antimicrobial
Agents Chemother., 37, 377-383, 1993.
5. Emmerson A.M., Jones A.M. - The quinolones: decades of
development and use. - J. Antimicrob. Chemother., 51, 13-20,
2003.
6. Sweetman S.C. - Naldixic acid. - In: Martindale, The Complete
Drug Reference, London, Pharmaceutical Press, 36th ed., 2010,
p. 303-305.
7. Guad R.S., Gupta G.D. (Eds.) - Practical Pharmaceutics. - 1st
ed., CBS Publishers, New Delhi, 2002.
8. Mura P., Nassini C., Proietti D., Manderioli A., CortiStirring
P. - Influence of vehicle composition variations on the in vitro
and ex vivo clonazepam diffusion from hydrophilic ointment
bases. - Pharm. Acta Helv., 71, 147-154, 1996.
9. Kesavanarayan K.S., Nappinnai M., Ilavarasan R. - Topical
dosage form of valdecoxib: preparation and pharmacological
evaluation. - Acta Pharm., 57, 199-209, 2007.
10. Pandey A., Jagtap J.V., Patil A.A., Joshi R.N., Kuchekarthe B.S.
- Formulation and evaluation of anti-bacterial and anti-fungal
activity of a herbal ointment containing Aloe vera, Azadirachta
indica and Curcuma longa. - J. Chem. Pharm. Res., 2, 182-186,
2010.
11. Malipeddi V.R., Dua K., Sara U.V.S., Malipeddi H, Agrawal
A. - Comparative evaluation of transdermal formulations of
norfloxacin with silver sulfadiazine cream, USP, for burn bound
healing property. - J. Burns Wounds, 5, 26-31, 2006.
12. Şenyiğit T., Sonvico F., Barbieri S., Özer Ö., Santi P., Colombo
P. - Lecithin/chitosan nanoparticles of clobetasol-17-propionate
capable of accumulation in pig skin. - J. Control. Release, 142,
368-373, 2010.
13. Siewert M., Dressman J., Brown C.K., Shah V.P. - FIP/AAPS
Guidelines to Dissolution/in vitro Release Testing of Novel/
Special Dosage Forms. - AAPS PharmSciTech, 4, 1-10, 2003.
14. Tous S.S., El Sayed A.M., Abd El Mohsen M.G., Agban M.N.,
Boushra M.F. - Enhancement of nalidixic acid solubility by co-
solvency and solid disoersion. - The Seventh Assiut University
International Pharmaceutical Sciences Conference, Assiut,
17-18 March 2010, p. 125-126.
15. Chowdary K.P.R., Rao P. V. - Biopharmaceutical studies on
solid dispersions of nalidixic acid in modified starches. - Drug
Dev. Ind. Pharm., 20, 3015-3022, 1994.
16. Rajalkshmi G., Damodharan V., Bhai C.V.KV., Janardhanreddy
Novel formulation and clinical evaluation of nalidixic acid ointment in impetigo
R.P. - Formulation and evaluation of clotrimazole and ichtham-
mol ointment. - Int. J. Pharm. Biol. Sci., 1, 7-16, 2010.
17. Tas C., O’zkan Y., Savaser A., Baykara T. - In vitro release
studies of chlorpheniramine maleate from gels prepared by
different cellulose derivatives. - Il Farmaco, 58, 605-611, 2003.
18. Rupal J., Kaushal J., SettMallikarjuna S.C., Dipti P. - Preparation
and evaluation of topical gel of valdecoxib. - Int. J. Pharm. Sci.
Drug Res., 2, 51-54, 2010.
19. Belal F., Al-Majed A. A., Al-Obaid A.M. - Methods of analysis of
4-quinolone antibacterials. - Talanta, 50, 765-786, 1999.
20. Hopkala H., Kowalczuk D. - Thin layer chromatographic analysis
of nalidixic acid, and oxolinc acid and cinoxacin. - Acta Polon.
Pharm. Drug Research, 56, 11-15, 1999.
21. Khamanga S.M., Parfitt N., Nyamuzhiwa T., Haidula H.,Walker
R.B. - The evaluation of eudragit microcapsules manufactured
by solvent evaporation using USP apparatus 1. - Dissolution
Technologies, 16, 15-21, 2009.
22. Ismail S., Mohamed A. A., Abd El Mohsen M.G. - In vitro release
of sulconazole nitrate from ointment bases. - Bull. Pharm. Sci.,
Assiut university, 13, 115-123, 1990.
23. El Faham T.H., Shawky S., Abd El Meguid E. - Evaluation of
the release and efficacy of two new antibacterial agents from
different ointment bases. - Pharmazie, 44, 211-214, 1989.
24. El Faham T.H., Safwat S.M. - Comparative study of the release
of antiinfammatory activity of indomethacin from topical formula-
tions. - Die Pharmazeutische Industrie, 45, 82-86, 1992.
25. Ezzedeen F.W., Shihab F.A., Husain E.J. - Percutaneous diffu-
sion of cefalexin, sulfamethoxazole and diphenhydramine from
ointments. - Pharmazie, 45, 512-514, 1990.
26. Hoa H.O., Chena L.C., Linb H.M., Sheua M.T. - Penetration
enhancement by menthol combined with a solubilization effect
in a mixed solvent system. - J. Control. Release, 51, 301-311
1998.
27. Aukunuru J., Chinnala K.M., Guduri V. - Development of a
novel transdermal ibuprofen ointment. - Curr. Trend. Biotechnol.
Pharm., 3, 97-104, 2009.
28. Arunachalam A., Karthikeyan M., Konam K., Prasad P.H., Se-
thuraman S., Ashutoshkumar S. - Solid dispersions: a review.
- Curr. Pharm. Res., 1, 82-90, 2010.
29. Balata G., Mahdi M., Abu Bakera R. - Improvement of solubility
and dissolution properties of ketoconazole by solid dispersions
and inclusion complexes. - Asian J. Pharma Sci., 5, 1-12, 2010.
30. Aejaz A., Azmail K., Saaullah S., Mohsen A.A. - Formulation and
in vitro evaluation of aceclofenac solid dispersion incorporated
gels. - Int. J. Appl. Pharm., 2, 7-12, 2010.
31. Muela S., Escalera B., Pena M.A., Bustamante P. - Influence of
temperature on the solubilization of thiabendazole by combined
action of solid dispersions and co-solvents. - Int. J. Pharm., 384,
93-99, 2010.
Manuscript
Received 24 October 2011, accepted for publication 3 January 2012.
352