الفهرس 
General IntroductionOnly 14 pages are availabe for public view
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Abstract
Summary
Nano-carriers are promising drug carrier systems being
developed to deliver the drugs to the target organ. These
systems have proven a great success in the treatment of several
diseases. Many drug carrier systems have been prepared and
successfully utilized in the treatment of skin fungal infections,
among which are liposomes, niosomes, transferosomes,
ethosomes, solid lipid nanoparticles (SLNs) and nanostructured
lipid carriers (NLCs).
Lipid nanocapsules (LNCs) are lipid nanoparticles with a
structure that is a hybrid between polymeric nanoparticles and
liposomes. Till current date, only one paper was published on
the use of LNCs in treatment of skin fungal infections.
Luliconazole is a novel broad spectrum antifungal drug used
topically for fungal infections such as Tinea and Onychomycosis.
Therefore, the aim of this work was to study the possibilities for
using essential oils having antifungal activity (Eucalyptus oil and
orange oil) for LNCs preparation instead of using medium chain
triglyceride (Miglyol®812) and incorporation of luliconazole in
those LNCs in an attempt to modulate its therapeutic efficacy in
diseases such as tinea pedis.
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The work in this thesis was divided into three chapters
Chapter I: Preparation and characterization of Lipid
nanocapsules using essential oils with antifungal activity
The work in this chapter dealt with the formulation, and
characterization of essential antifungal oil LNCs. LNCs were
prepared by phase inversion method, and characterized for their
morphology using transmission electron microscopy, particle
size, zeta potential and in vitro antimycotic activity. Stability of
eucalyptus oil LNCs and orange oil LNCs stored at 4˚C was
tested through monitoring the changes in particle size and zeta
potential for six months.
The work in this chapter included the following:
1- Preparation of LNCs by phase inversion temperature
method. LNCs were prepared using different natural oils
with reported antifungal activity (eucalyptus oil and
orange oil) instead of using the most commonly used
medium chain triglyceride (Miglyol®812).
2- A ternary diagram was used to plot all possible mixtures,
composed of distilled water, hydrophilic surfactant
(Kolliphor®HS15) and the oil (eucalyptus oil or orange
oil)
3- characterization of the prepared LNCs was performed
through the following studies:
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193
a) Particle size, PDI and zeta potential analysis of freshly
prepared LNCs formulations were carried out using zetasizer.
b) Samples of LNCs were examined for morphology by
transmission electron microscopy.
c) Physical stability study on all LNCs formulations was
conducted over a storage period of six months at 4° C by the
assessment of the effect of storage on the particle size, charge
and PDI of the nanoparticles.
d) Viscosity determination of selected LNCs (O36 and
E35) formulation was carried out using Brookfield rheometer
(DV3T).
e)The electron micrographs of formulae O36 and E35
showed spherical shaped particles with clear core and shell
morphology
f) The in-vitro antimycotic activity of selected LNCs
formulations (O36), (E35) was evaluated and compared to a
standard LNCs formula prepared using Miglyol®812 (M36).
The results of this work revealed the following:
1- Successfully prepared LNCs using essential
antifungal oils (eucalyptus oil and orange oil).
2- Eucalyptus oil LNCs particle size ranged from 60-402
nm.
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194
3- Orange oil LNCs particle size ranged from 26-145
nm.
4- All prepared LNCs were charged, with charges
ranging from (-12 to -34 mV).
5- Eucalyptus oil and orange oil LNCs displayed good
storage properties as manifested by slight changes in
particle size, polydispersity index and zeta potential
values.
6- Transmission electron microscopy of LNCs displayed
spherical shaped particles with clear core and shell
morphology.
7- orange oil based LNCs (O36) was found to follow the
typical Newtonian behavior where the viscosity was
constant and there was a linear relationship between
the shear stress and shear rate
8- Eucalyptus based LNCs (E35) was found to follow a
shear thinning or pseudo plastic behavior where a
gradual decrease in viscosity occurred upon
increasing the shear rate
9- The highest antifungal activity was observed in E35
containing eucalyptus oil (35%w/w) having inhibition
zone (3.56 ± 0.12 cm), then O36 containing orange oil
(40%w/w) having inhibition zone (2.6 ± 0.1 cm)
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195
compared to M36 containing medium chain
triglyceride (Miglyol®812) which did not show any
antifungal properties.
Chapter II: Preparation, characterization, in-vitro and exvivo
evaluation of luliconazole loaded lipid nanocapsules
(LNCs)
The work in this chapter was concerned with the loading
luliconazole in the selected eucalyptus oil LNCs and orange oil
LNCs. characterization of the prepared formulae was done
through the following studies: particle size, zeta potential, invitro
release, in-vitro antimycotic activity and skin deposition
experiments. Stability study on the LNCs formulations stored at
4˚C was conducted as well.
The work in this chapter included the following:
1- Loading luliconazole in the selected eucalyptus oil LNCs.
2- Loading luliconazole in the selected orange oil LNCs.
3- characterization of the prepared LNCs was done through
the following studies:
a) Particle size and zeta potential analysis of freshly
prepared luliconazole LNCs was carried out using
Zetasizer.
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196
b) Physical stability study on luliconazole based LNCs was
conducted over a storage period of four months at 4° C
by the assessment of the effect of storage on the particle
size, charge and PDI of the nanoparticles.
c) Viscosity of luliconazole loaded LNCs were carried out
using Brookfield rheometer (DV3T).
d) Samples of luliconazole loaded LNCs were examined
for morphology by transmission electron microscopy.
e) In-vitro release study was performed on the selected
luliconazole loaded LNCs formulae.
f) Ex-vivo deposition/permeation of the selected
luliconazole loaded LNCs formulae was carried out
using Franz diffusion apparatus and compared with the
oil (eucalyptus oil and orange oil) solution of the drug.
g) The in-vitro antimycotic activity of selected LNCs
formulations (O36l), (E35l) was evaluated and compared
to a standard LNCs formula prepared using Miglyol®812
(M36l).
The results of this work revealed the following:
1- Luliconzole orange oil LNCs and luliconazole eucalyptus
oil LNCs were successfully prepared using phase
inversion temperature method.
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2- Luliconazole loaded LNCs particle size ranged from 40 -
174 nm.
3- All of the prepared luliconazole loaded LNCs were
negatively charged, with zeta potential values ranging
from (-14 to -38 mV).
4- Eucalyptus oil luliconazole LNCs and orange oil
luliconazole LNCs displayed the best storage properties
as manifested by the slight changes in particle size,
polydispersity index and zeta potential values.
5- The formulations containing eucalyptus oil as oily phase
revealed significant increase in viscosity values (above
20cp) compared to the formulations containing orange
oil as oily phase (near to 1cp) (p<0.05).
6- Transmission electron microscopy of luliconazole loaded
LNCs displayed spherical shaped particles with clear
core and shell morphology.
7- The in-vitro drug release results from eucalyptus oil
formulae showed that the highest % luliconazole released
after 20 hours was from E26l (75.18±0.76) and the
lowest % luliconazole released after 20 hours was from
E24l (14.91±1.65).
8- The in-vitro drug release results from orange oil
formulae showed that the highest % luliconazole
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released after 20 hours was from O36l (59.04±3.40) and
the lowest % luliconazole released after 20 hours was
from O23l (39.29±3.09).
9- Ex vivo skin deposition experiments demonstrated the
high potential of the selected LNCs formulae (E35l and
O25l) in accumulating the drug into the deeper epidermal
and dermal layers of the skin, and hence they were
selected for further clinical studies compared to a
standard LNCs formula prepared using Miglyol®812
(M36l).
10- The highest antifungal activity was observed in E35l
containing eucalyptus oil (35%w/w) having inhibition
zone (4.16 ± 0.05 cm), followed by O25l containing
orange oil (20%w/w) having inhibition zone (3.4 ± 0.1
cm). The lowest antifungal activity was for M25l
having inhibition zone (2.43 ± 0.11 cm).
Chapter III: In-vivo and clinical studies of selected
luliconazole loaded lipid nanocapsules (LNCs)
In this chapter, the therapeutic efficacy of luliconazole
eucalyptus oil LNCs and luliconazole orange oil LNCs was
done through the following studies: in-vivo animal model using
candida albicans for induction of fungal infection and also the
selected formulae were clinically tested on patients suffering
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199
from tinea pedis, by satisfaction and efficacy report filled by
the patients under the doctor supervision.
The work in this chapter included the following:
1- In-vivo study was carried on 32 male albino rats to evaluate
the antifungal effect of selected luliconazole eucalyptus oil
LNCs, luliconazole orange oil LNCs and Miglyol ®812
luliconazole LNCs for 7 days and compared to blank
formulae. This study included:
a) Fungal burden scoring: 10 blocks from the same animal
were implanted onto one plate. Every block was
inspected visually for candida albicans growth. The
skin blocks yielding fungal growth were regarded as
culture positive. Culture negative skin blocks had no
growth around any side of the skin block.
b) The histopathological study: through staining skin
tissue samples with hematoxylin and eosin and Periodic
Acid-Schiff.
2- The clinical study was conducted on fourteen patients
suffering from tinea pedis fungal lesion. Dermatological
examination was performed to detect the type of fungal
infections. Fungal scraping using 10% potassium hydroxide
(KOH) solution offers an inexpensive, rapid, and useful
technique for the daily practice of clinicians and
mycologists managing patients with clinically suspected
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200
tinea pedis. The patients were photographed before
treatment with follow up photographing every week up to 2
weeks or complete improvement.
The results of this work revealed the following:
1- Collectively the results of the fungal burden scoring and
histopathological study revealed that candida caused skin
damage in epidermis and dermis with diffuse inflammatory
edema and mononuclear inflammatory cells infiltration. On
the topical administration of E35l as antifungal treatment
containing luliconazole (antifungal drug) and eucalyptus oil
(essential antifungal oil), the results showed that E35l
treated candidiasis and its induced inflammation more than
E35, O25l, O25, M25l and M25
2- As could be delineated from the results, patients receiving
medicated eucalyptus oil LNCs (E35l) displayed
significantly better clinical therapeutic outcome compared to
plain eucalyptus oil LNCs (E35), medicated orange oil
LNCs (O25l) and medicated miglyol® 812 LNCs (M25l).
This could be attributed to the synergistic effect of essential
antifungal oil (eucalyptus oil) and luliconazole along with
the higher skin deposition (no flux rate) potential of the
medicated eucalyptus oil LNCs (E35l).