Drug Delivery

ISSN: (Print) (Online) Journal homepage: www.tandfonline.com/journals/idrd20

Tailoring and optimization of a honey-based

nanoemulgel loaded with an itraconazole–thyme
oil nanoemulsion for oral candidiasis

Amal M. Sindi, Waleed Y. Rizg, Muhammad Khalid Khan, Hala M. Alkhalidi,

Waleed S. Alharbi, Fahad Y. Sabei, Eman Alfayez, Hanaa Alkharobi,
Mohammed Korayem, Mohammed Majrashi, Majed Alharbi, Mohammed
Alissa, Awaji Y. Safhi, Abdulmajeed M. Jali & Khaled M. Hosny

To cite this article: Amal M. Sindi, Waleed Y. Rizg, Muhammad Khalid Khan, Hala M. Alkhalidi,
Waleed S. Alharbi, Fahad Y. Sabei, Eman Alfayez, Hanaa Alkharobi, Mohammed Korayem,
Mohammed Majrashi, Majed Alharbi, Mohammed Alissa, Awaji Y. Safhi, Abdulmajeed M. Jali &
Khaled M. Hosny (2023) Tailoring and optimization of a honey-based nanoemulgel loaded with
an itraconazole–thyme oil nanoemulsion for oral candidiasis, Drug Delivery, 30:1, 2173337,
DOI: 10.1080/10717544.2023.2173337

To link to this article: https://doi.org/10.1080/10717544.2023.2173337

© 2023 The Author(s). Published by Informa Published online: 27 Jan 2023.

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Drug Delivery
2023, VOL. 30, NO. 1, 2173337
https://doi.org/10.1080/10717544.2023.2173337

RESEARCH ARTICLE

Tailoring and optimization of a honey-based nanoemulgel loaded

with an itraconazole–thyme oil nanoemulsion for oral candidiasis
Amal M. Sindia , Waleed Y. Rizgb , Muhammad Khalid Khanc, Hala M. Alkhalidid, Waleed S. Alharbib,
Fahad Y. Sabeie, Eman Alfayezf, Hanaa Alkharobif, Mohammed Korayemg, Mohammed Majrashih,
Majed Alharbii, Mohammed Alissaj, Awaji Y. Safhie, Abdulmajeed M. Jalik and Khaled M. Hosnyb
a
Department of Oral Diagnostic Sciences, Faculty of Dentistry, King Abdulaziz University, Jeddah, Saudi Arabia; bDepartment of
Pharmaceutics, Faculty of Pharmacy, King Abdulaziz University, Jeddah21589, Saudi Arabia; cDepartment of Biochemical Materials,
Beautsway commercial foundation, Cairo, Egypt; dDepartment of Clinical Pharmacy, Faculty of Pharmacy, King Abdulaziz University, Jeddah,
21589, Saudi Arabia; eDepartment of Pharmaceutics, College of Pharmacy, Jazan University, Jazan45142, Saudi Arabia; fDepartment of Oral
Biology, Faculty of Dentistry, King Abdulaziz University, Jeddah, Saudi Arabia; gPreventive Dental Sciences Department, Faculty of Dentistry,
Albaha University, Albaha, Saudi Arabia; hDepartment of Pharmacology, College of Medicine, University of Jeddah, Jeddah, 23890, Saudi
Arabia; iDepartment of Pharmaceutical Chemistry, Faculty of Pharmacy, King Abdulaziz University, Jeddah, Saudi Arabia; jDepartment of
Medical Laboratory Sciences, College of Applied Medical Sciences, Prince Sattam bin Abdulaziz University, Al-Kharj, 11942, Saudi Arabia;
k
Department of Pharmacology and Toxicology, College of Pharmacy, Jazan University, Saudi Arabia

ABSTRACT ARTICLE HISTORY

The use of essential oil–based nanoemulsions (NEs) has been the subject of extensive research Received 21 November 2022
on a variety of conditions affecting the oral cavity. NEs are delivery methods that improve the Revised 1 January 2023
solubility and distribution of lipid medicines to the intended areas. Because of their antibacterial Accepted 2 January 2023
and antifungal properties, itraconazole and thyme oil–based self-nanoemulsifying drug delivery KEYWORDS
systems (ItZ-ThO-SNEDDS) were created to protect oral health against oral microorganisms. The Sustainability of natural
ItZ-ThO-SNEDDS were created utilizing an extreme verices mixture design, and varying concentrations resources; Honey-based gel;
of ThO (10% and 25%), labrasol (40% and 70%), and transcutol (20% and 40%) were used. The Itraconazole; thyme oil; design
ItZ-ThO-SNEDDS had droplet sizes of less than 250 nm, a drug-loading efficiency of up to 64%, of experiments; oral
and a fungal growth inhibition zone of up to 20 mm. The accepted design was used to obtain microbiota
the ideal formulation, which contained ThO in the amount of 0.18 g/ml, labrasol 0.62 g/ml, and
transcutol 0.2 g/ml. The best ItZ-ThO-SNEDDS formulation was incorporated into a honey-based
gel, which demonstrated improved release of ItZ in vitro and improved transbuccal permeation
ex vivo. In addition, when compared with various formulations tested in rats, the optimized loaded
emulgel decreased the ulcer index. This study therefore demonstrated that the ItZ-ThO-SNEDDS
could offer an effective defense against oral diseases caused by microbial infections.

1. Introduction Dental clearance prior to medical procedures such as cardiac

surgery, cancer/bisphosphonate therapy, and radiation ther-
For overall health and a high quality of life, oral health is
apy reduces the systemic and oral side effects of these pro-
crucial. In 2012, the World Health Organization defined oral
cedures (Rajan et al., 2014).
health as the absence of mouth or facial pain, oral infections
Free-living eukaryotic organisms known as fungi can be
or sores, and other disorders that restrict a person’s ability
to engage in with his daily and psychosocial activities (World molds (filamentous fungi), yeasts (round fungi), or a blend
Health Organization, 2012). Oral diseases continue to be a of both (dimorphic fungi) (Terezhalmy & Huber, 2011). One
neglected area of global health despite the significant social of the common fungal infections of the oral mucus mem-
and financial costs they impose on many nations (FDI World branes is oral candidiasis (Prasanna, 2012). It is caused by
Dental Federation, 2015). Most adults have tooth decay, and the yeast Candida albicans, which is a fairly normal compo-
15% to 20% of middle-aged adults have serious gum disease nent of the oral microbiota; it is found in 30% to 50% of
(Buset et al., 2016). Poor dental health may be linked to such people. As people age, the rate of carriage rises (Dangi et al.,
diseases as diabetes, heart disease, stroke, pneumonia, and 2010). In 60% of patients over 60 years of age with dental
other respiratory illnesses, according to the literature (Silva problems, C. albicans is recovered from their mouths (Parihar,
et al., 2015). Another typical reason for headaches and ear 2011). In addition to C. albicans, there are several other spe-
and facial pain is disorders of the jaw (Katz et al., 2010). cies of Candida that can be found in the oral environment,

CONTACT Khaled M. Hosny [email protected] Department of Pharmaceutics, Faculty of Pharmacy, King Abdulaziz University, Jeddah, Saudi
Arabia
© 2023 The Author(s). Published by Informa UK Limited, trading as Taylor & Francis Group.
This is an Open Access article distributed under the terms of the Creative Commons Attribution License (http://creativecommons.org/licenses/by/4.0/), which permits unrestricted
use, distribution, and reproduction in any medium, provided the original work is properly cited.
2 A. M. SINDI ET AL.

including Candida guilliermondii, Candida krusei, Candida gla- source of glucose for the thriving cellular components in
brata, Candida parapsilosis, Candida pseudotropicalis, and wounds (Molan, 2002). Honey’s water activity is lower than
Candida tropicalis (Dangi et al., 2010). 0.91 aw, and this inhibits and limits bacterial development
Itraconazole (ItZ) is a triazole antifungal that can be taken on injured sites and promotes fluid movement that causes
orally and has a propitious pharmacokinetic profile. It has debris and necrotic tissue to be sloughed from the wound
been shown to have a wide range of efficacy (Chong & (Gleiter et al., 2006; Sundoro et al., 2012).
Sullivan, 2007). ItZ has been demonstrated in noncomparative A breakthrough idea called nanomedicine has shown great
clinical trials to be highly effective when delivered once or promise for delivering prospective medications with low sol-
twice daily for a variety of superficial and more consequential ubility and bioavailability (Salem et al., 2022; Hussein et al.,
‘deep’ fungal ailments (DiMasi et al., 2003). This drug’s original 2022; Ali et al., 2021; Alhakamy et al., 2022). A nanoemulsion
purpose was to act as a broad-spectrum antifungal drug by (NE) is a delivery system that can improve solubility and
inhibiting 14-α-lanosterol demethylase (14LDM), an enzyme stability (Rizg et al., 2021). Widely researched as drug delivery
that causes ergosterol to be produced in fungi and choles- platforms, NEs are made up of nanosized oil droplets stabi-
terol to be produced in humans (Okabayashi et al., 2009). It lized by a combination of surfactants and co-surfactants
is used to prevent immunosuppressive diseases, as well as (Hosny et al., 2021). Using EO–based NEs to combat oral
treat fungal infections such aspergillosis, candidiasis, and bacteria is a viable strategy (Hosny et al., 2021). Gels are
histoplasmosis (Chang et al., 2017). ItZ has uncommon side relatively new dosage forms often generated when significant
effects such as neutropenia, liver failure, and heart failure quantities of aqueous, hydroalcoholic, or nonpolar vehicles
but is generally considered to be safe (Girois et al., 2006). are mixed in a three-dimensional (3D) network of polymeric
There are numerous members of the genus Thymus of the components (Ali et al., 2021). However, they are less effective
family Lamiaceae. These plants are frequently employed for at delivering hydrophobic medications (Wong & Dodou,
culinary, cosmetic, and therapeutic purposes (Ghasemi et al., 2017). Emulgels, a hybrid of emulsions and gels, were there-
2020). The most recognized plant ingredient in the industrial fore a viable alternative that would allow introducing hydro-
pharmacy sector is thyme, which is derived from Thymus phobic medications topically (Talat et al., 2021). These dosage
vulgaris L. and Thymus zygis L. (Kosakowska et al., 2020). The forms are well liked because they combine the benefits of
chemical constituents of thyme essential oil (EO) include both emulsions and gels, like regulated drug release of emul-
monoterpene alcohols, phenol derivatives, ketones, alde- sions and excellent thermodynamic stability of gels (Laffleur
hydes, ethers, and esters, among other chemical groups & Keckeis, 2020).
(Bendif et al., 2020). The two isomeric phenolic monoterpenes In order to gather the most data with the fewest exper-
thymol (2-isopropyl-5-methylphenol) and carvacrol (2-methyl- iments, and provide an explanation for any experimental
5-(1-methyethyl) phenol) make up most of the thyme EOs errors, statistical experimental designs were developed (Gregg
(Sharifi-Rad et al., 2018; Nagoor Meeran et al., 2017). Stetsko Manager, 1986). Utilizing these designs has the added
For centuries, people have used the thyme plant and its benefit of requiring that formulations be made carefully and
EOs to treat infections related to the upper respiratory tract, that statistical laws be followed. This means that scientists
the symptoms of bronchitis, pruritus brought on by derma- must be precise when setting up the experimental goals and
titis, bruises, and sprains (Kohlert et al., 2002). These days, it the procedures for meeting those goals. Additionally, the
is frequently used in dental medicine as a disinfectant and design can forecast the behavior of the ideal formulation
as an expectorant for coughs brought on by colds (Nilima (Hosny et al., 2020). Because statistical designs frequently
et al., 2013). It has antibacterial effects on both Gram-positive lead to an optimum formulation, they can be a cost-effective
and Gram-negative bacteria, as well as antifungal, antispasm, method of study (Alkhalidi et al., 2018). In order to create a
anti-inflammatory, antioxidant, and antiviral activities against novel formulation for the successful treatment of oro-mucosal
human rhinoviruses, influenza viruses, and herpes simplex ulcers brought on by oral candidiasis, the current study tar-
virus type I (Walther et al., 2020). There have been no cases geted the development of a honey-based nanoemulgel con-
of toxicity when the oil is taken at levels regularly used, and taining an itraconazole and thyme oil–based nanoemulsion
therefore it is generally regarded as safe (Eva et al., 2018; (ItZ-ThO-NE) and evaluated its antibacterial and antifungal
Salehi et al., 2018). efficacy and calculated its ulcer index.
Several antimicrobial medicines have lost their efficacy for
treating wounds owing to the development of
treatment-resistant microbes (Guo & Dipietro, 2010). The use 2. Materials and methods
of honey for wounds is one of the alternative medicines that
2.1. Materials
have been sought out more widely used in clinical settings
for the treatment of wounds (Molan, 2001). Honey’s inherent The beauts-way Commercial Co., Ltd. (Jeddah, Saudi Arabia)
qualities and active ingredients are essential for the process provided the ItZ and ThO. The Saudi Arabian Japanese
of healing in wounds (Molan & Rhodes, 2015). Natural honey Pharmaceutical Company Ltd generously donated the PVP
has a jelly-like viscosity and forms a superficial film covering and PEG (SAJA, Jeddah, Saudi Arabia). Agar and Laureth-20
wounds and lesions and prevents bacteria and dehydration were acquired from the Sigma Water Import and Export Co.,
(Langemo et al., 2009; Sell et al., 2012). Because of its high Ltd. (Shaanxi, China). Tween and Cremophor EL, as well as
sugar content, honey produces a strong osmotic gradient isopropyl alcohol and glycerin, were procured from Sigma
that draws fluid up the subdermal tissue and provides a Aldrich (St. Louis, Missouri, USA). Labrasol and transcutol
 Drug Delivery 3

were generously provided by Gattefosse (Saint-Priest, France), Table 1. Upper and lower limits of investigated factors combined with
measured responses and their constraints.
and the honey was acquired from the Hali Health Company
(Ras Al Khaimah, UAE). The other compounds that were uti- Levels
lized in this experiment were all of analytical grade. Purified Studies factors low high
water was used in the experiments. A: ThO % 10.0 25.0
B: Labrasol % 40.0 70.0
C: Transcutol % 20.0 40.0
Measured responses Constrains
2.2. Methods Droplet size (Y1) minimize
ItZ loading capability (Y2) Maximize
2.2.1. Solubility studies Inhibition Zone (Y3) Maximize
To determine the variable self-emulsifying regions that may
be used to construct ThO-based NEs containing ItZ, the drug
solubility was evaluated in a variety of surfactants and factors were noted and optimized utilizing the Design of
co-surfactants (Hosny et al., 2021). Experiments.
To determine the drug’s solubility in each specific solution,
excess quantities of ItZ were dissolved in 3 ml of the surfac-
2.2.4. Visual screening of ItZ-ThO-SNEDDS
tants, namely, Tween, labrasol, Laureth-20, and Cremophor
Visual inspection was used to assess the effectiveness of the
EL, as well as the co-surfactants transcutol, isopropyl alcohol,
production of the ItZ-ThO-SNEDDS in terms of their clarity
glycerin, HCO-20, and propylene glycol. The combinations
and stability; this included defects such as coalescence or
were kept in vials for 3 days at 25 ± 2 °C in a water bath. After
the shattering of spontaneously generated NEs (Hosny
the required balance was achieved, the mixtures were pre-
et al., 2021).
cipitated by centrifugation for 20 minutes at 4000 rpm to
remove any extra medication. In order to determine the ItZ
content of the supernatants, a high-performance liquid chro- 2.2.5. Assessment of the nanoemulsions’ droplet sizes
matography (HPLC) method using a C18 HS (250 × 4.6 mm) Double-distilled deionized water, 400 ml, was used to dilute
column was employed (Thimmaraju et al., 2012). Acetonitrile 100 ml of each formulation, and the blend was mixed using
and double-distilled water were combined in this procedure’s a vortex mixer for 5 minutes. Aliquots of 200 µl were taken
mobile phase in a ratio of 90:10 v/v at a flow rate of 1.0 ml/ from the diluted samples to determine the droplet size and
min. A UV detector set at 263 nm was used for the detection. polydispersity index (PDI) using a Zetatrac and a dynamic
It was discovered that ItZ had a retention time of 7.5 minutes. light-scattering approach (Microtrac, Inc., Montgomeryville,
ItZ also had a linearity range of 4 to 60 µg/ml and a linear Pennsylvania, USA) (Hosny et al., 2021).
regression coefficient of 0.991.
2.2.6. Analysis of ItZ-ThO-SNEDDS’ drug-loading
2.2.2. Itz-ThO emulsifying regions’ tenacity in particular efficiency
surfactants and co-surfactants By independently dispersing a known excess quantity of ItZ
In a review to determine the lower and higher limits for in 1 g of a plain SNEDDS, the ItZ loaded in each SNEDDS
every ingredient of the self-nanoemulsifying drug delivery mixture was calculated. The mixtures were placed in vials
systems (SNEDDS), which were chosen based on solubility, and kept in a shaking water bath at 25 °C for 24 hours. The
the self-emulsifying regions were located using pseudoter- blends were precipitated by centrifugation for approximately
nary phase diagrams. When diluted with water and gently 15 minutes at 4500 rpm after reaching equilibrium. The pre-
stirred within those limits, blends of ItZ, ThO, the chosen cipitates were gathered, thoroughly cleaned, and then dis-
surfactant (labrasol), and the chosen co-surfactant (transcutol) solved in methanol. The amount of ItZ was calculated using
spontaneously created a pellucid emulsion with globules in the previously described HPLC procedure after the drug had
the nanometer range. When the droplet size is greater than been removed. The following mathematical formula was used
1 nm but smaller than 1 m, the emulsion area in the phase to calculate the drug-loading capacity (Hosny et al., 2021):
area can be recognized as an NE region. The ItZ-loaded NEs
were then tested. Product sdrug content  mg 
Drugloadingefficiency  100 Eqn. (1)
Total product weight  mg 
2.2.3. Design of ItZ-ThO self-nanoemulsifying
formulations and optimization of experiments 2.2.7. Valuation of growth inhibition zone against
Because of its great effectiveness in evaluating the emulsions Candida albicans
and projecting the best solutions, an extreme vertices mix- The disk diffusion method was used to assess the formula-
ture experimental design (Table 1) was used for the creation tion’s antifungal effectiveness. To prepare C. albicans strains
and selection of the best ItZ-ThO-SNEDDS. To ascertain the for inoculation and conduct an in vitro antifungal evaluation
impacts of the ThO level (A), labrasol level (B), and transcutol utilizing the fungus strains, the strains were grown on
level (C) in 17 formulations, a mixed design based on extreme Sabouraud dextrose agar medium (Hosny et al., 2021).
vertices was used. The prepared SNEDDS’ globule size (Y1),
drug-loading efficiency (Y 2), and growth inhibition of 2.2.7.1. Inoculum development:. Five standard C. albicans
C. albicans (Y3) were the dependent responses. The response strain colonies were chosen, suspended in 2 ml of normal
4 A. M. SINDI ET AL.

saline, and thoroughly mixed. The 0.6-meq McFarland powder and ThO as an oil were used to make the honey gel
standards were used to preserve the uniform turbidity of for F3; that is, a SNEDDS was not used. The nanoemulgel for
the microbial suspension. To obtain the grass culture, streaks F4 was created without adding honey. The F5 honey gel,
were produced on the Sabouraud dextrose agar media by manufactured in the same way as the F3, did not contain
inserting a sterilized cotton swab into the prepared fungal ThO, and the F6, prepared in the same way as the F4, did
suspension. not contain ItZ powder. The produced gels were evaluated
for the following characteristics.
2.2.7.2. Disk diffusion technique:. To prepare the test and
positive control samples, sterile filter paper disks measuring
2.2.10.1. In-vitro dissolution of the hydrogel formulations. In
5 mm in diameter were submerged in 100 µl of the ItZ-ThO-
this study, the USP Dissolution Tester (Apparatus I) was used.
SNEDDS and ethyl alcohol (99.9%), respectively. The paper
Instead of using baskets, glass cylindrical tubes measuring
disks were inserted into seeded Sabouraud dextrose agar
2.7 cm in diameter and 10 cm in length were attached to
media after being soaked. The infected plates were incubated
the spinning shafts and securely sealed with semipermeable
for 48 hours at 24 ± 2 °C before they were examined for each
membranes of 100 µm in pore size. These tubes contained the
zone of inhibition.
formulations to be assessed. The tested formulations (F2 to
F5) each weighed 10 g and contained 50 mg of ItZ. Each glass
2.2.8. Optimization of ItZ-ThO-SNEDDS tube containing one of the formulations was submerged in
For the analysis of variance (ANOVA) of the resulting models, 50 ml of phosphate buffered saline (PBS, pH 6.8). The medium
the F-ratio, p-value, and degrees of freedom were computed was agitated at a speed of 25 rpm while the release study
was conducted at 37 ± 0.5 °C.
for the investigated parameters and their interactions. The
Over 1 hour, samples were taken out of the dissolved mix-
responses were utilized to choose the model that would be
ture at various intervals. Using the previously described HPLC
most suitable for the data based on the results. The p-values
procedure, the absorbance was measured to determine how
below .05 showed that the tested model predictors were
much ItZ had been released (Hosny et al., 2021).
significant. The model fitness was further evaluated using
the coefficient of variation (CV%) values, determination cor-
relations, anticipated R2 value, and adapted R2 value. In this 2.2.10.2. Ex-vivo transmucosal penetration test of the
hydrogel formulations. Ex-vivo tests were conducted on
way, the NEs with the smallest globule size and highest
the ItZ aqueous suspension and various manufactured
drug-loading efficacy were identified. formulations (F2 to F5), each of which contained 5 mg/ml of
ItZ. An automated Franz diffusion cell (MicroettePlus, Scilogex
2.2.9. Choosing the optimized ItZ-ThO-SNEDDS LLC, Rocky Hill, Connecticut, USA) was used as the apparatus,
The development and evaluation of the formulations involved and sheep buccal mucosal membrane obtained from a nearby
local butcher was used as the model penetration membrane.
evaluating the drug-loading capability and globule size. The
The Franz diffusion cell’s donor and receptor compartments
formulations were then examined for their ex-vivo perme- were adequately separated by the processed sheep
ation, in-vitro drug release, and ulcer index. buccal mucosa. The exposed surface area of the receptor
compartment covered by the membrane was 1.75 cm2. The
2.2.10. Preparation of honey-based nanoemulgel loaded medium was agitated at a rate of 400 to 450 rpm while being
kept at a temperature of 37 ± 0.5 °C in the drawing chamber,
with ItZ-ThO-NE
which encompassed 8 ml of PBS (pH 6.8).
An optimized ItZ-ThO-SNEDDS–loaded honey nanoemulgel
At regular intervals, precise aliquots were automatically
was created by combining 5% (v/v) honey with 10% (w/v)
removed, and the previously described HPLC method was
PVP, 1% (w/v) agar solution, and 2% (v/v) PEG. An aqueous
used to quantify the ItZ content. A graph was made to show
PVP solution made by dispersing 10 g of PVP in 60 ml of
how long it took for the ItZ to penetrate the Q per cm2 of
distilled water was used to create the simple honey gel (i.e.
the membrane; this finding let researchers better understand
formulation F1), which was allowed to sit at room tempera-
how the drug was dispersed throughout the mucosa. From
ture (25 °C) overnight. To create the agar blend, 1 g of agar
the acquired diffusion data, significant parameters such as
was solubilized in 20 ml of distilled water, heated until clear,
the Jss (steady-state flow), Pc (permeability coefficient), EF
and stirred constantly before 2 ml of PEG was added. Fifteen
(enhancement factor), and D (diffusion coefficient) were
ml of honey was introduced when the mixture’s temperature
determined. Plots were made of the comparative dissemina-
dropped below 45 °C after the homogeneous mixture had
tion patterns for various formulations. The following equation
been in a sonicator water bath for an hour at 37 °C. The same
(2) was used to determine the percentage of ItZ that pene-
procedures were followed in the preparation of an optimized
trated the membrane and the overall amount of ItZ distrib-
honey nanoemulgel (i.e. formulation F2) that was loaded with
uted throughout the receptor chamber (Hosny et al., 2021):
the optimized ItZ-ThO-SNEDDS, but 10 g of PVP was dissolved
in 60 ml of the NE created by dissolving 10 ml of the ItZ-ThO-
Lp
SNEDDS in 50 ml of distilled water and stirring for 5 minutes Percentage permeated  100  Eqn. (2)
LT
before adding PVP. The remaining ingredients were then
added as in the previously prepared plain honey gel. where LT was the initial concentration of ItZ in the donor
Additionally, four different hydrogel formulations were chamber and Lp was the quantity of ItZ that was transferred
prepared for comparison (formulations F3 to F6). ItZ as a into the receptor compartment.
 Drug Delivery 5

2.2.10.3. Assessment of the growth inhibition zone against

Candida albicans. By using the previously indicated disk
diffusion technique, the antifungal activity of the investigated
formulations (F1 to F6), as well as the aqueous dispersion
of ItZ, was assessed.

2.2.10.4. Ulcer index evaluation. Male albino rats weighing

150 to 250 g were used for this investigation. The in-vivo
study was performed according to the institutional guidelines
of the Animal Ethics Committee of Cairo Agriculture for
Experimental Animals, Cairo, Egypt, Approval No. (137-10-
22). The ulcer index was assessed for seven animal groups,
each consisting of three animals, using a previously described
methodology (Alkhalidi et al., 2020). Group 2 received the Figure 1. Solubilization properties of ItZ in various surfactants.
ItZ aqueous suspension and was the positive control. Group
3 received formulation F1; Group 4 received the optimized
nanoemulgel, or formula F2; Group 5 received formulation
F4; Group 6 received formulation F5; and Group 7 received
formulation F6. Group 1 received only normal saline and was
the negative control.
After 3 days of therapy with each formulation, the ulcer-
ative effect on the oral mucous membranes of every treated
group was graded as 1 (normal-colored epithelial lining), 2
(red-colored epithelial lining), 3 (ulcers < 1 mm), 4 (ulcers >
1 mm but with no bleeding areas), and 5 (ulcers > 1 mm and
with hemorrhagic streaks).

Figure 2. Solubilization properties of ItZ in various co-surfactants.

3. Results and discussion
3.1. Assessment of ItZ solubility
The ItZ had the best level of solubility when it was mixed
with the surfactant labrasol and co-surfactant transcutol,
according to the investigations into its solubility in various
surfactants and co-surfactants. Figures 1 and 2 show the
relationship between the types of surfactants and
co-surfactants utilized and the percentage of ItZ solubilized.
In order to determine the highest and lowest limits of each
component of the SNEDDS, labrasol and transcutol were
chosen for the pseudoternar y phase diagram
construction.

3.2. The ability of certain surfactants and

co-surfactants to emulsify ItZ-ThO-EOs with a high Figure 3. The pseudoternary phase plot of labrasol, transcutol, and thyme EO.
degree of tenacity
In order to stabilize the system created by combining water
seen in the pseudoternary phase diagram in Figure 3. These
and oil, a surfactant is frequently employed, with the goal
concentration ranges were used to create the statistical
of reducing the particle size. According to Tsai et al. (2014),
design that was later used for the creation and optimization
the oil phase typically consists of natural triglycerides, hydro-
of several ItZ-ThO-SNEDDS formulations.
carbons, and other nonpolar hydrophobic molecules, while
the aqueous phase typically contains solutes and electrolytes
that have been dissolved in water. To accurately pinpoint the
3.3. Visual examination of ItZ-ThO-SNEDDS
range of concentrations of the thyme EO, labrasol, and trans-
cutol that should produce regions of NE, a pseudoternary An examination of the ItZ-ThO-SNEDDS with the naked eye
phase diagram was built for the current study. revealed translucid dispersions that were clear and did not
The precise concentration levels of thyme EO, labrasol, exhibit any coalescence or breakage. This implied that the
and transcutol for making NEs were found to be 10% to concentration ranges that were specified were exact in the
25%, 40% to 70%, and 20% to 40%, respectively, as can be establishment of stable NEs.
6 A. M. SINDI ET AL.

Table 2. Extreme vertices mixture design and the associated responses.

Y1:Globule size Y2: ItZ loading Y3: Inhibition
Run A:ThO B:Labrasol C:Transcutol (nm) capacity (%) Zone (mm) PDI
1 0.179 0.621 0.200 103 ± 1.6 63 ± 2.0 18 ± 0.7 0.11
2 0.169 0.536 0.294 143 ± 2.2 45 ± 1.0 9 ± 0.3 0.15
3 0.169 0.536 0.294 144 ± 1.9 46 ± 0.9 10 ± 0.5 0.12
4 0.192 0.471 0.336 151 ± 2.7 52 ± 1.3 12 ± 0.6 0.09
5 0.151 0.449 0.400 172 ± 2.5 41 ± 1.1 8 ± 0.4 0.10
6 0.169 0.536 0.294 142 ± 1.1 45 ± 0.5 9.5 ± 0.8 0.18
7 0.112 0.640 0.247 98 ± 0.9 40 ± 1.8 7.5 ± 0.3 0.28
8 0.250 0.485 0.264 188 ± 2.9 59 ± 0.4 16 ± 0.7 0.21
9 0.100 0.593 0.306 104 ± 3.1 39 ± 1.0 7 ± 0.5 0.25
10 0.100 0.700 0.200 84 ± 2.1 34 ± 0.3 6 ± 0.9 0.30
11 0.250 0.421 0.329 229 ± 3.3 57 ± 0.8 15 ± 0.4 0.32
12 0.179 0.621 0.200 102 ± 1.2 64 ± 1.1 20 ± 1.0 0.29
13 0.250 0.550 0.200 177 ± 1.8 61 ± 1.0 17 ± 0.3 0.22
14 0.100 0.500 0.400 161 ± 2.8 30 ± 1.5 5 ± 0.5 0.26
15 0.250 0.421 0.329 230 ± 3.9 56 ± 0.9 15 ± 0.1 0.17
16 0.118 0.538 0.344 138 ± 1.1 34 ± 0.3 6 ± 0.2 0.11
17 0.198 0.402 0.400 240 ± 3.0 54 ± 0.6 13 ± 0.6 0.19

Table 3. Regression analysis of the Y1, Y2, and Y3 responses.

Dependent variables R2 Adjusted R2 Predicted R2 F-value p-value Adequate precision
Y1 0.9995 0.9991 0.9960 2203.64 0.0001 148.1595
Y2 0.9975 0.9950 0.9799 118.06 0.0001 58.9733
Y3 0.9871 0.9741 0.8931 26.68 0.0002 25.2220

3.4. Assessment of ItZ-ThO-SNEDDS’ droplet size oily droplets of the NE and, hence, for bigger droplets to be
attained.
The NEs had droplet sizes of 84 ± 2.1 to 240 ± 3.0 nm (Table 2)
and PDI values between 0.09 and 0.32. This demonstrated
the formulations’ uniformity, stability, and size distribution.
3.5. Assessment of ItZ incorporation efficiency within
The collected globule size data were subjected to a special
ItZ-ThO-SNEDDS
quadratic model of polynomial analysis. The researched
model was effective at determining the substantial impact As demonstrated in Table 2, the loading of ItZ in the NEs
of the thyme EO (A), labrasol (B), and transcutol (C) on the ranged between 30 ± 1.5% and 64 ± 1.1%.
ItZ-ThO-SNEDDS’ droplet sizes, according to the chosen math- The resulting drug-loading data were subjected to a spe-
ematical design. As shown in Table 3, the model had a mod- cial quartic model of polynomial analysis. The ItZ-ThO-
ulated R2 value of 0.9991 and a predicted R2 value of 0.9960, SNEDDS’ drug-loading capacity was evaluated using the
which were closely correlated. An ANOVA data analysis pro- model to assess the major impacts of ThO (A), labrasol (B),
duced the equation shown below: and transcutol (C). As shown in Table 3, the chosen model
Droplet size = +527.85 A + 83.81 B + 262.16 C − 510.83 AB − achieved an adjusted R2 value of 0.9950 and a forecasted R2
479.36 AC − 185.83 BC − 2689.11 A2BC + 3526.21 AB2C − 1622.86 value of 0.9799. An ANOVA analysis of the data used the
ABC2 Eqn. (3) equation below:
The perturbation, 3D surface, and contour plots in Figure 4 ItZ loading capacity = − 36.98 A + 34.34 B + 10.14 C + 248.22
show how the parameters affected the size of the ItZ-ThO- AB + 267.58 AC + 55.50 BC + 792.65 A2BC − 1183.21 AB2C −
SNEDDS droplets. The graphs demonstrate how the amount 635.31 ABC2 Eqn. (4)
of different formulation variables in the formulations affected Figure 5 shows the main effect, contour, and 3D surface
the globule size of the generated NEs. plots that illustrate how the investigated parameters affected
According to the graph, labrasol (i.e. factor B) had the big- the drug loading in the ItZ-ThO-SNEDDS.
gest impact on the droplets’ size. The increase in surfactant As might be noticed from the preceding plots, the ThO
limits reduced the size of the globules, as shown in Figure 4(A), level (i.e. factor A) significantly increased the incorporation of
since this caused a lowering of the interfacial tension that ItZ into the NE’s droplets. This finding might be due to the
existed between the aqueous and nonaqueous phases (Hosny great capacity of the oil to absorb ItZ because of the lipophilic
et al., 2020). The fact that increasing the percentage of oil nature of ItZ. Further, improving the percentage of oil in the
reduced the surfactant and co-surfactant levels and, therefore, emulsion would lower the accompanying levels of labrasol and
limited their capacity to minimize the droplet size may help transcutol and, hence, lessen their ability to force the drug
to explain the increase in droplet size in relation to the into the surrounding aqueous milieu (Hosny et al., 2021).
increased amount of ThO. Larger oil globules were produced Additionally, as can be seen from the graph, large amounts of
as a result, and this was consistent with the outcomes men- labrasol or transcutol were associated with a lower loading
tioned in the literature (Ziani et al., 2011; Pavoni et al., 2020). capacity of ItZ in the NE. These outcomes could be ascribed
Moreover, boosting the ThO level might have allowed for more to the comparable decrease in oil levels and, hence, the
ItZ, which is a lipophilic agent, to be incorporated into the decreased space into which ItZ could be incorporated in the
 Drug Delivery 7

Figure 4. The impacts of various independent factors on the droplet size of various ItZ-ThO-SNEDDS are depicted in (A) a perturbation plot, (B) a contour
plot, and (C) a 3D surface plot.

Figure 5. The impacts of various factors on the drug loading of various ItZ-ThO-SNEDDS are depicted in the (A) perturbation, (B) contour, and (C) 3D surface
plots.

NE droplets. It could also be related to the interfacial tension– Figure 6 shows the main effect, contour, and 3D surface
lowering properties of labrasol and transcutol. graphs that illustrate how the investigated parameters
impacted the fungal growth inhibition zones of ItZ-ThO-
SNEDDS against C. albicans.
3.6. Evaluation of fungal growth inhibition zones
As might be seen from the above graphs, ThO has a great
of ItZ-ThO-SNEDDS against Candida albicans
capacity for expanding the growth inhibition zones of C.
As is shown in Table 2, the inhibition zones against C. albicans albicans. Ergosterol is a special sterol that can only exist in
created by the NE formulations ranged from 5 ± 0.5 to 20 ± 1.0 cm. the outer membrane of a fungus and is crucial to its healthy
The zone inhibition data were fitted to a special quartic development and operation. As a result, substances affecting
model of polynomial analysis. The ItZ-ThO-SNEDDS’ growth the level of ergosterol might have antifungal properties
zone inhibition capabilities were examined using the selected (Kowalczyk et al., 2020). Thymol, the major component of
statistical model to assess the major impacts of ThO (A), ThO, has potential antifungal action that is based on how
labrasol (B), and transcutol (C). As shown in Table 3, the it affects the metabolism of fatty acids, especially ergosterol,
chosen model had an adjusted R2 value of 0.9741 and a in fungal cells (De Lira Mota et al., 2012). Among other
predicted R2 value of 0.8931. An ANOVA analysis of the data things, it produces oxidative stress and an increase in reac-
used the equation below: tive oxygen species, and this lowers the levels of capsular
Inhibition Zone = − 23.62 A + 6.05 B − 2.54 C + 105.23 AB + polysaccharide and the extracellular polymer matrix (EPS)
103.74 AC + 19.93 BC + 401.09 A2BC − 562.56 AB2C − 295.23 (Al-Shahrani et al., 2017). In cell membranes of Candida and
ABC2 Eqn. (5) Cryptococcus that had been exposed to thymol, ergosterol
8 A. M. SINDI ET AL.

Figure 6. The impacts of various factors on the fungal growth inhibition zones of various ItZ-ThO-SNEDDS are depicted in the (A) perturbation, (B) contour,
and (C) 3D surface plots.

Figure 7. Bar chart and desirability ramp for process optimization. The levels of the parameters that were studied and the predicted estimations for the
dependent variables of the optimal formulation.

levels decreased, as previously stated in the literature. This 0.62 g/ml of labrasol, and 0.2 g/ml of transcutol, according to
resulted in disruptions of the membrane integrity and Statgraphics software, which also suggested many other
membrane-associated enzymes, significant damage, and, options that represented different combinations of the vari-
ultimately, cell death (Poonam et al., 2019). ables under study. The optimal formulation had a drug-loading
The decrease in the size of the inhibition zones associated capability of 67%, a droplet size of 105 nm, and an inhibition
with the increase in labrasol (B) and transcutol (C) levels zone of 18.177 mm, and it acquired a desirability of 0.931. The
might be correlated with the comparable decrease in the desirability ramp is shown in Figure 7, which also shows the
level of ThO and, hence, its lowered ability to destroy the levels of the independent variables and the anticipated values
fungal cell membrane. The decrease in the area of the growth of the measured responses for the best formulation. Table 4
zone area observed when the surfactants were increased shows that the parameters of the optimum formulation’s ver-
might be explained by the expected decrease in ItZ loaded itable and anticipated values were very nearly identical, with
into the NE and, hence, the decreased overall antifungal no appreciable discrepancies (p > .05), and this supported the
activities of the formulations. accuracy, validity, and precision of the equations.

3.7. Itz-ThO-SNEDDS optimization 3.8. In-vitro dissolution behavior of the ItZ-ThO-

SNEDDS–loaded hydrogel formulations
The optimized NE formulation was created using the most
appropriate qualities revealed by the aforementioned data. The evaluated formulations’ drug release profiles are shown
The best combination was composed of 0.18 g/ml of ThO, in Figure 8. The drug suspension had a very low cumulative
 Drug Delivery 9

Table 4. Values obtained experimentally and realistically for the ideal NE formulation.
Droplet size ItZ loading Inhibition zone
Solution ThO Labrasol Transcutol (nm) capacity(%) (mm) desirability
Predicated value 0.1810 0.6190 0.2000 102.989 64 18.77 0.931
Experimental value 0.1810 0.6190 0.2000 105 67 18.00 0.931

Figure 8. ItZ in-vitro release characteristics of the formulations.

Table 5. Ex-vivo permeation parameters of ItZ through sheep buccal mucosa.

Parameters of permeation F2 F4 F3 F5 ItZ Aqueous suspension
Cumulative amount permeated, Q24 (μg/cm2)a 3125 2614 1882 800 750
Steady state flux, Jss, (μg/cm2.min)b 52.4 44.9 31.4 16.2 18.8
Permeability coefficient, P, (cm/min)c 4.1 × 10-3 3.5 × 10-3 2.6 × 10-3 1.1 × 10-3 1.2 × 10-3
Diffusion coefficient, D, (cm2/min)d 2.03 × 10-4 1.72 × 10-4 1.21 × 10-4 0.58 × 10-4 0.51 × 10-4
Enhancement factor (Fen)e 4.16 3.48 2.5 1.06 –
N.B. a: Q = the cumulative ItZ amount permeated thru membrane unit area; b: Jss = obtained from slop of curve plotted between Q and time; c: P = ItZ
Jss/original concentration; d: D = Calculated from the slop of curve plotted between Q and square root of time; e: EF: Q of studied formulations/Q of
ItZ suspension.

percentage of the drug released (22 + 2%) after 60 minutes; contained in the NE of same components (i.e. F4 and F2).
this might be due to the low water solubility of ItZ. Formulation F5, which contained ItZ powder in a gel base
Formulation F4, which was a nanoemulgel without honey, but no ThO, had a percentage of release of 19 ± 1% of ItZ.
had a drug release percentage of 85 ± 3%. Such a high per- This low result might have been due to the presence of ItZ
centage of release of the optimized formulation compared in a powder form but no surfactants that could have solu-
with the other examined formulations might be attributable bilized it, as well as to its viscous gel base, which further
to the NE content of amphiphilic molecules, which may hindered its diffusion and release.
improve drug release by enhancing ItZ dissolution (Hosny
et al., 2021). Additionally, the small globule size of the NEs 3.9. Ex-vivo transmucosal penetration test of the
provided a broader surface area for drug release and aided hydrogel formulations
in increasing the amount of drug released, enabling a higher
amount of absorption and a quicker initiation of the drug’s As might be observed from Table 5, formulation F2, which
effect (Hosny et al., 2021). In addition, the lack of honey in was composed of the optimal formulation–loaded hydrogel
the formulation might have contributed to a decrease in the that contained honey, achieved a Q24 value of 3125 μg/cm2
formulation’s viscosity, which could be expected to enhance with a 4.16-fold increase in permeation compared with the
the ItZ release. Meanwhile, formulation F2, which contained ItZ aqueous suspension, which had a Q24 of 750 μg/cm2. Such
the optimal nanoemulgel and honey, had a drug release a result demonstrates that the NEs were significantly involved
percentage of 72 ± 4%. The decrease in ItZ release from for- in enhancing the ItZ penetration through the mucosa. The
mulation F2 compared with formulation F4 could be due to NEs’ vital role in increasing drug permeation can be ascribed
the higher viscosity of the F2 with its honey content, which to its content of surfactant, which might contribute to drug
might have restrained the drug release (Salem et al., 2022). diffusion from the formulation and exert a considerable effect
Formulation F3, which contained ItZ powder and ThO dis- on fluidizing the mucosal cell membrane, leading to enhanced
persed in a gel base, attained a percentage of release of ItZ permeation (Salem et al., 2020). In addition, the small size
39 ± 4% of ItZ. Such a low release percentage may have of the NE globules provided a larger surface area for drug
resulted from poor ItZ solubility due to the lack of amphi- permeation. Simultaneously, formulation F4 had a Q24 of
philic molecules in this formulation compared with the gel 2614 μg/cm2. Such a decrease in the Q24 compared with F4,
10 A. M. SINDI ET AL.

Table 6. Ulcer index values of different animal groups treated with the tested received F4) or the lack of ThO in Group 6 (which received
formulations (means ± SD, n = 6).
the gel containing ItZ powder only). Such outcomes empha-
Group treated Tested Formulation Ulcer Index size the ulcer-reducing properties of both honey and ThO.
Group 1 Normal saline 1 The ulcer-healing activity of ThO was further proved by the
Group 2 aqueous dispersion of ItZ 5
Group 3 formulation F1 1 results for Group 7, which received F6, which was composed
Group 4 formulation F2 1 of the gel loaded with NE but without ItZ; this group had
Group 5 formulation F4 3 an ulcer index of 2. This low ulcer index may have been due
Group 6 formulation F5 3
Group 7 formulation F6 2 to the ThO content. ThO is well known for its anti-inflammatory
N.B. F1: plain hydrogel; F2: optimum NE-loaded gel; F4: optimum NE-loaded and antioxidant properties, which contribute to its ability to
gel but without honey; F5: gel loaded with ItZ powder and ThO; F6: gel heal lesions and ulcers; this is due to its main phenolic com-
containing only ThO. pounds, thymol and carvacrol (Nagoor Meeran et al., 2017;
Kohlert et al., 2002).
even though both contained the ItZ NE, might be due to the
presence of honey in F2. This could provide greater viscosity,
which might have allowed for more intimate contact and a 4. Conclusion
prolonged residence time of the formulation within the mucosa ItZ was effectively developed as a functional NE in combi-
and, thus, a greater chance for ItZ to permeate the mucosal nation with ThO. A pseudoternary phase diagram was used
membrane (Laffleur, 2014). Moreover, F3, which contained ItZ to estimate the ideal concentrations of ThO, labrasol, and
powder and ThO dispersed in a gel base, had a Q24 of 1882 μg/ transcutol in combination to create the necessary drug deliv-
cm2, while F5, which contained ItZ powder in a gel base but ery system. The created systems appeared to be reasonably
no ThO, had a Q24 of 800 μg/cm2. Such a large difference in stable, as evidenced by the NEs, which had globule sizes
the Q24 values of these two formulations may be attributable between 84 ± 2.1 and 240 ± 3.0 nm with acceptable homoge-
to the oil’s potential to increase permeability by altering and neous distribution. Additionally, the created formulations
modifying cell membrane characteristics, such as by boosting had a drug-loading efficiency of between 30 ± 1.5% and
ItZ solubility and penetration through the mucosa or by 64 ± 1.1% and a fungal growth inhibition zone of up to
improving the membrane fluidity, as was previously described 20 mm. The extreme vertices mixture design was used to
in the literature (Rizg et al., 2021; Jiang et al., 2017). obtain the best formulation, which consisted of the variables
ThO, labrasol, and transcutol at levels of 0.18 g/ml, 0.62 g/
3.10. Ulcer index determination ml, and 0.2 g/ml respectively. The optimal formulation was
loaded into a honey-based oral gel and showed enhanced
The data in Table 6 show that F2 (i.e. the optimum release and transmucosal permeation. When tested in rats,
ItZ-ThO-SNEDDS-loaded gel) had the lowest ulcer index. It the optimized ItZ-ThO-SNEDDS–loaded gel displayed the
was comparable to that of the negative control group, which lowest ulcer index value when compared with other formu-
received normal saline, and Group 3, which received plain lations. Overall, this study showed that ItZ-ThO–based NEs
gel. This finding revealed the substantially low ulcer-forming in combination with a honey-based gel could offer an effec-
capacity of F2. Such noticeable activity might be ascribed to tive defense against oral diseases brought on by microbial
the nature of the gel base carrying F2, which had honey infections.
among its components. Honey has certain physical traits that
encourage its application as a medicinal drug to fight certain
infections (Hixon et al., 2019). These characteristics of honey Acknowledgments
are also connected to its ability to reduce inflammation and The Deanship of Scientific Research (DSR) at King Abdulaziz University
promote wound healing, antioxidant and the capacity to (KAU), Jeddah, Saudi Arabia, has funded this project, under grant no.
scavenge free radicals. It is an immune stimulator that can RG-5-166-43
improve the immunological response (Almasaudi et al., 2016).
It can be used to treat a variety of ulcers; it can also be
utilized for wound healing and skin sanitation. In addition Disclosure statement
to its anti-inflammatory and bacteria-fighting capabilities, it The authors declare no conflicts of interest.
accelerates the healing of wounds and reduces the ulcer
index (Almasaudi et al., 2017).
These results were further confirmed when they were Ethical Approval statement
compared with the results for Group 2, which received the The in-vivo study was performed according to the institutional guidelines
aqueous dispersion containing the drug and had an ulcer of the Animal Ethics Committee of Cairo Agriculture for Experimental
index of 5. Group 2 received ItZ twice daily for 14 days, and Animals, Cairo, Egypt, Approval No. (137-10-22).
although ItZ has antifungal properties, because it is a syn-
thetic drug it contributed to the injury of the buccal mucosal
membrane. Groups 5 and 6, which received F4 and F5, ORCID
respectively, both had ulcer indexes of 3. The change in the Amal M. Sindi http://orcid.org/0000-0002-5286-1756
ulcer index for Groups 5 and 6 from that of Groups 3 and Waleed Y. Rizg http://orcid.org/0000-0002-3088-4084
4 might be due to the lack of honey in Group 5 (which Khaled M. Hosny http://orcid.org/0000-0002-4904-1378
 Drug Delivery 11

References Hixon KR, Klein RC, Eberlin CT, et al. (2019). A critical review and per-
spective of honey in tissue engineering and clinical wound healing.
Alhakamy NA, Hosny KM, Aldryhim AY, et al. (2022). Development and Adv Wound Care 8:403–15.
optimization of ofloxacin as solid lipid nanoparticles for enhancement Hosny K, Asfour H, Rizg W, et al. (2021). Formulation, optimization, and
of its ocular activity. J Drug Deliv Sci Technol 72:1. evaluation of oregano oil nanoemulsions for the treatment of infec-
Ali SA, Sindi AM, Mair YH, et al. (2021). Oral gel loaded by ethotrans- tions due to oral microbiota. Int J Nanomed 16:5465–78.
fersomes of antifungal drug for oral thrush: preparation, characteri- Hosny KM, Alhakamy NA, Sindi AM, et al. (2020). Coconut oil nanoemul-
zation, and assessment of antifungal activity. J Drug Deliv Sci Technol sion loaded with a statin hypolipidemic drug for management of
66:102841. burns: formulation and in vivo evaluation. Pharmaceutics 12:1061.
Ali SA, Sindi AM, Mair YH, Khallaf RA. (2021). Oral gel loaded by etho- Hosny KM, Khallaf RA, Asfour HZ, et al. (2021). Development and opti-
transfersomes of antifungal drug for oral thrush: preparation, char- mization of cinnamon oil nanoemulgel for enhancement of solubili-
acterization, and assessment of antifungal activity. J Drug Delivery ty and evaluation of antibacterial, antifungal and analgesic effects
Sci Technol 66:102841. against oral microbiota. Pharmaceutics 13:1008.
Alkhalidi HM, Hosny KM, Rizg WY. (2020). Oral gel loaded by fluconazole– Hosny KM, Rizg WY, Khallaf RA. (2020). Preparation and optimization of
sesame oil nanotransfersomes: development, optimization, and as- in situ gel loaded with rosuvastatin-ellagic acid nanotransfersomes
sessment of antifungal activity. Pharmaceutics 13:27. to enhance the anti-proliferative activity. Pharmaceutics 12:263.
Alkhalidi HM, Naguib GH, Kurakula M, et al. (2018). In vitro and preclin- Hosny KM, Sindi AM, Alkhalidi HM, et al. (2021). Development of omega-3
ical assessment of factorial design based nanoethosomal transdermal loxoprofen-loaded nanoemulsion to limit the side effect associated
film formulation of mefenamic acid to overcome barriers to its use with NSAIDs in treatment of tooth pain. Drug Deliv 28:741–51.
in relieving pain and inflammation. J Drug Del Sci Technol 48:450–13. Hussein RM, Kandeil MA, Mohammed NA, Khallaf RA. (2022). Evaluation
Almasaudi SB, Abbas AT, Al-Hindi RR, et al. (2017). Manuka honey exerts of the hepatoprotective effect of curcumin-loaded solid lipid nanopar-
antioxidant and anti-inflammatory activities that promote healing of ticles against paracetamol overdose toxicity: Role of inducible nitric
acetic acid-induced gastric ulcer in rats. Evid Based Complement oxide synthase. J Liposome Res 8:1–11.
Alternat Med 2017:5413917. Jiang Q, Wu Y, Zhang H, et al. (2017). Development of essential oils as
Almasaudi SB, El-Shitany NA, Abbas AT, et al. (2016). Antioxidant, skin permeation enhancers: penetration enhancement effect and
anti-inflammatory, and antiulcer potential of manuka honey against mechanism of action. Pharm Biol 55:1592–600.
gastric ulcer in rats. Oxid Med Cell Longev 2016:3643824. Katz J, Wallet S, Cha S. (2010). Periodontal disease and the oral-systemic
Al-Shahrani MH, Mahfoud M, Anvarbatcha R, et al. (2017). Evaluation of connection: “is it all the RAGE?”. Quintessence Int 41:229–37.
antifungal activity and cytotoxicity of Thymus vulgaris essential oil. Kohlert C, Schindler G, März RW, et al. (2002). Systemic availability and
PC 7:34–40. pharmacokinetics of thymol in humans. J Clin Pharmacol 42:731–7.
Bendif H, Peron G, Miara MD, et al. (2020). Total phytochemical analysis Kosakowska O, Bączek K, Przybył JL, et al. (2020). Morphological and
of Thymus munbyanus subsp. coloratus from Algeria by HS-SPME-GC-MS, chemical traits as quality determinants of common thyme (Thymus
NMR and HPLC-MSn studies. J Pharmaceut Biomed 186:113330. vulgaris L.), on the example of ‘StandardWinter’ cultivar. Agronomy
Buset SL, Walter C, Friedmann A, et al. (2016). Are periodontal diseases 10:909.
really silent? A systematic review of their effect on quality of life. J Kowalczyk A, Przychodna M, Sopata S, et al. (2020). Thymol and thyme
Clin Periodontol 43:333–44. essential oil-new insights into selected therapeutic applications.
Chang YL, Yu SJ, Heitman J, et al. (2017). New facets of antifungal ther- Molecules 25:4125.
apy. Virulence 8:222–36. Laffleur F, Keckeis V. (2020). Advances in drug delivery systems: work in
Chong CR, Sullivan DJ.Jr. (2007). New uses for old drugs. Nature 448: progress still needed? Int J Pharm X 2:100050.
645–6. Laffleur F. (2014). Mucoadhesive polymers for buccal drug delivery. Drug
Dangi YS, Soni MS, Namdeo KP. (2010). Oral candidiasis: a review. Int J Dev Ind Pharm 40:591–8.
Pharm Pharm Sci 2:36–41. Langemo DK, Hanson D, Anderson J, et al. (2009). Use of honey for
De Lira Mota KS, de Oliveira Pereira F, de Oliveira WA, et al. (2012). wound healing. Adv Skin Wound Care 22:113–8.
Antifungal activity of Thymus vulgaris L. essential oil and its constit- Molan P, Rhodes T. (2015). Honey: a biologic wound dressing. Wounds
uent phytochemicals against Rhizopus oryzae: interaction with ergos- 27:141–51.
terol. Molecules 17:14418–33. Molan PC. (2001). Potential of honey in the treatment of wounds and
DiMasi JA, Hansen RW, Grabowski HG. (2003). The price of innovation: burns. Am J Clin Dermatol 2:13–9.
new estimates of drug development costs. J Health Econ 22:151–85. Molan PC. (2002). Re-introducing honey in the management of wounds
Eva L, Christin M, Ahmed M, et al. (2018). Authorised medicinal product and ulcers – theory and practice. Ostomy Wound Manage 48:28–40.
Aspecton® Oral Drops containing thyme extract KMTv24497 shows Nagoor Meeran MF, Javed H, Taee HA, et al. (2017). Pharmacological
antiviral activity against viruses which cause respiratory infections. J properties and molecular mechanisms of thymol: Prospects for its
Herb Med 13:26–33. therapeutic potential and pharmaceutical development. Front
FDI World Dental Federation. (2015). The challenge of oral diseases. 2nd Pharmacol 8:380.
ed. Brighton, UK: Myraid Editions. Nilima T, Silpi B, Rakesh NB, et al. (2013). Antimicrobial efficacy of five
Ghasemi G, Alirezalu A, Ghosta Y, et al. (2020). Composition, antifungal, essential oils against oral pathogens: an in vitro study. Eur J Dent
phytotoxic, and insecticidal activities of Thymus kotschyanus essential 7:71–7.
oil. Molecules 25:1152. Okabayashi K, Imaji M, Osumi T, et al. (2009). Antifungal activity of
Girois SB, Chapuis F, Decullier E, et al. (2006). Adverse effects of anti- itraconazole and voriconazole against clinical isolates obtained from
fungal therapies in invasive fungal infections: review and meta-analysis. animals with mycoses. Nihon Ishinkin Gakkai Zasshi 50:91–4.
Eur J Clin Microbiol Infect Dis 25:138–49. Parihar S. (2011). Oral candidiasis – a review. Webmedcentral Dent 2:1–
Gleiter R, Horn H, Isengard H. (2006). Influence of type and state of 18.
crystallisation on the water activity of honey. Food Chem 96:441–5. Pavoni L, Perinelli DR, Bonacucina G, et al. (2020). An overview of micro-
Gregg Stetsko Manager (1986). Statistical experimental design and its and nanoemulsions as vehicles for essential oils: formulation, prepa-
application to pharmaceutical development problems. Drug Develop ration and stability. Nanomaterials (Basel 10:135.
Indust Pharm 12:1109–23. Poonam K, Neha A, Apurva C, et al. (2019). Delineating the biofilm in-
Guo S, Dipietro LA. (2010). Factors affecting wound healing. J Dent Res hibition mechanisms of phenolic and aldehydic terpenes against
89:219–29. Cryptococcus neoformans. ACS Omega 4:17634–48.
12 A. M. SINDI ET AL.

Prasanna KR. (2012). Oral candidiasis – a review. Scholarly J Med 2:6–30. Sundoro A, Nadia K, Nur A, et al. (2012). Comparison of physical–chem-
Rajan B, Ahmed J, Shenoy N, et al. (2014). Assessment of quality of life ical characteristic and antibacterial effect between Manuka honey
in patients with chronic oral mucosal diseases: a questionnaire-based and local honey. J Plastik Rekonstruksi 1:3.
study. Perm J 18:e123-7. Talat M, Zaman M, Khan R, et al. (2021). Emulgel: an effective drug
Rizg WY, Hosny KM, Elgebaly SS, et al. (2021). Preparation and optimi- delivery system. Drug Dev Ind Pharm 47:1193–9.
zation of garlic oil/apple cider vinegar nanoemulsion loaded with Terezhalmy GT, Huber MA. (2011). Oropharyngeal candidiasis: etiology,
minoxidil to treat alopecia. Pharmaceutics 13:2150. epidemiology, clinical manifestations, diagnosis, and treatment. Crest
Salehi B, Mishra AP, Shukla I, et al. (2018). Thymol, thyme, and Oral‑B at Dentalcare.com Contin Educ Course 1–16.
o ther plant sources: health and potential uses. Phytother Res
­ Thimmaraju MK, Pamulaparthy V, Raghunandan N. (2012). Development
32:1688–706. and validation of RP-HPLC method for the determination of itraconazole
Salem HF, El-Menshawe SF, Khallaf RA, et al. (2020). A novel transdermal in bulk and capsule dosage form. J Anal Chem 2:10.
nanoethosomal gel of lercanidipine HCl for treatment of hypertension: Tsai M-J, Fu Y-S, Lin Y-H, Huang Y-B, Wu P-C. (2014). The effect of
optimization using Box-Benkhen design, in vitro and in vivo charac- Nanoemulsion as a carrier of Hydrophilic compound for transdermal
terization. Drug Deliv Transl Res 10:227–40. delivery. PLoS ONE 9:e102850.
Salem HF, Nafady MM, Ewees MGE, et al. (2022). Rosuvastatin Walther C, Döring K, Schmidtke M. (2020). Comparative in vitro analysis of
calcium-based novel nanocubic vesicles capped with silver inhibition of rhinovirus and influenza virus replication by mucoactive
nanoparticles-loaded hydrogel for wound healing management: op- secretolytic agents and plant extracts. BMC Complement Med Ther 20:380.
timization employing Box-Behnken design: in vitro and in vivo as- Wong RSH, Dodou K. (2017). Effect of drug loading method and drug
sessment. J Liposome Res 32:45–61. physicochemical properties on the material and drug release prop-
Sell SA, Wolfe PS, Spence AJ, et al. (2012). A preliminary study on the erties of poly (ethylene oxide) hydrogels for transdermal delivery.
potential of manuka honey and platelet-rich plasma in wound heal- Polymers (Basel) 9:286.
ing. Int J Biomater 2012:1–14. World Health Organization. Oral health fact sheet. (2012). Available at:
Sharifi-Rad M, Varoni EM, Iriti M, et al. (2018). Carvacrol and human http://www.who.int/ mediacentre/factsheets/fs318/en/
health: A comprehensive review. Phytother Res 32:1675–87. Ziani K, Chang Y, McLandsborough L, et al. (2011). Influence of surfactant
Silva AE, Demarco FF, Feldens CA. (2015). Oral healthrelated quality of life and charge on antimicrobial efficacy of surfactant-stabilized thyme oil
associated factors in Southern Brazilian elderly. Gerodontology 32:35–45. nanoemulsions. J Agric Food Chem 59:6247–55.