Results in Chemistry 6 (2023) 101010

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Results in Chemistry
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Development of mucoadhesive in-situ nasal gel formulation for enhanced

bioavailability and efficacy of rizatriptan in migraine treatment
Kaushik Suhagiya a, Chetan H. Borkhataria a, *, Sumit Gohil a, Ravi A. Manek a, Kalpesh A. Patel b,
Nilesh K. Patel a, Dhaval V. Patel c
a
B K Mody Government Pharmacy College, Government Polytechnic Campus, Nr Aji Dam, Rajkot 360003, Gujarat, India
b
Government Pharmacy College, Dr SS Gandhi Government Engineering Campus, Majura Gate, Surat, Gujarat 395001, India
c
Sr. Specialist, QA, Moderna Therapeutics, USA

A R T I C L E I N F O A B S T R A C T

Keywords: This research article focuses on the development of an in-situ nasal gel formulation for the antimigraine agent,
Rizatriptan Rizatriptan (RZT), to enhance its bioavailability and efficacy in treating migraine. The aim of the study was to
Mucoadhesive overcome the challenges of rapid mucociliary clearance. Carbopol 934P (1–2 %w/v) was added to the primary
In-situ nasal gel
solution, followed by HPMC K4M (0.4–1.2 %w/v). Different copolymers were tested, and HPMC K4M was chosen
Carbopol 934P
due to its compatibility with the other ingredients. The pH of all formulations was in the nasal pH range, and the
HPMC K4M
drug content was above 97% in all batches. Gelling time varied from 1.3 to 8.6 sec, with low gelling time
observed when viscosity and mucoadhesive strength were high. The results were analyzed using a statistical
model with interactive and polynomial terms. The statistical analysis showed a good correlation between
dependent and independent variables, with R2 values of 0.97758 and 0.948931 for viscosity and mucoadhesive
strength, respectively. The Cmax of RZT in the brain was significantly higher (p < 0.05) via intra-nasal (340.27
ng) route as compared to the oral route and the same was the case for AUC. Finally, it was concluded that the RZT
nasal in-situ gel system improves bioavailability compared with oral route and could provide fast action in
migraine therapy.

Introduction hypersensitivity to odours (hypersomnia) or light (photophobia). The

Nasal drug delivery has been a successful route of administration
since ancient times. It offers better brain availability for many drugs and
is considered a valuable tool in treating various brain disorders. The
olfactory region of the nose is particularly important, because it contains
specialized ciliated nerve cells for smell perception, receives ophthalmic
and maxillary divisions of the trigeminal nerve, and has direct access to
cerebrospinal fluid [1,2].
Migraine is a syndrome characterized by intense headache and
nausea that occur at irregular intervals and last for several hours. Ac­
cording to the Global Burden of Disease (GBD) study, migraine is the
second most common cause of disability and the leading cause among
women under 50 [3]. Classic migraine is typically heralded by an “aura”
accompanied by spreading homonymous visual field defects with col­
oured sharp edges, known as “fortification” spectra [4].
Patients with migraine may also experience difficulty focussing on
certain objects, an insatiable appetite for particular foods, and
exact cause of these symptoms is unknown, but it is likely related to a
disturbance in cranial blood flow. In addition to a genetic predisposi­
tion, precipitating factors such as psychological stress, lack of sleep, and
certain foods are required to provoke an attack.
Migraine’s pharmacotherapy has two goals: stopping the acute
attack and preventing subsequent attacks. Rizatriptan (RZT) is an anti­
migraine agent belonging to BCS class III category. It is a small molecule
with a molecular weight of 269.34 g/mol and a short biological half-life
of 2–3 hrs, which results in low brain availability due to insufficient time
for penetration and low permeability. RZT is a selective agonist of se­
rotonin 1B and 1D-type receptors, causing vasoconstriction and activa­
tion of 5-HT1 receptors on peripheral terminals of the trigeminal nerve
innervating cranial blood vessels, thereby contributing to its anti­
migraine action in humans. It is used for migraine attacks with or
without aura. RZT undergoes significant first-pass metabolism, with
51% of the oral dose excreted as the indole acetic acid metabolite before
urinary excretion. To overcome RZT first-pass metabolism and

* Corresponding author.
E-mail address: chetanborkhataria@gmail.com (C.H. Borkhataria).

https://doi.org/10.1016/j.rechem.2023.101010
Received 8 May 2023; Accepted 13 June 2023
Available online 16 June 2023
2211-7156/© 2023 The Author(s). Published by Elsevier B.V. This is an open access article under the CC BY-NC-ND license (http://creativecommons.org/licenses/by-
nc-nd/4.0/).
K. Suhagiya et al.
permeability issues, nasal drug delivery may be a suitable alternative,
but further research is needed.
Animal studies have demonstrated that the concentration of cocaine
in the brain is higher after nasal administration compared to intravenous
injection at early time points, the existence of a pathway from the nose
to the brain [5,6]. For instance, in a mouse model, [3H]-dopamine
reached the olfactory lobe after nasal administration, and after 4 hrs,
the concentration in this tissue was 27 times higher than after intrave­
nous injection [7].
One of the significant challenges faced by formulators is to overcome
the protective barriers of the nasal cavity without causing permanent
tissue damage. The primary issue with nasal solutions is their rapid
clearance from the nasal cavity, with a half-life of clearance of 15–20
min for both liquid and powder formulations that lack mucoadhesive
properties [8,9]. Therefore, using mucoadhesive gel formulations could
be a possible strategy to extend the residence time at the nasal absorp­
tion site, thereby enhancing drug uptake. Approaches to improving
nasal bioavailability include using viscosity-enhancing or in-situ gelling
polymers to prolong the contact time with the nasal surface. An in-situ
gel is a drug delivery system that undergoes a sol-to-gel phase transi­
tion by changing specific physicochemical parameters such as ionic,
temperature, or pH [10,11]. This drug delivery system can release the
drug in a sustained manner; maintaining relatively constant plasma
profiles. As RZT is a BCS class III drug with high hydrophilicity, its ab­
sorption is rate-limiting by permeation. Therefore, using nasal in-situ gel
could enhance its bioavailability by delivering the drug to the brain
through the olfactory region while also eliminating the adverse effects of
first-pass metabolism and gastrointestinal disturbances [12]. Elissa et al.
(2022) developed glycerol monooleate, poloxamer 407 and tween 80
containing GS-loaded cubosomal in-situ gel utilizing a melt dispersion-
emulsification technique and were able to obtain improved bioavail­
ability and brain distribution [13].
The objective of the present study was to prepare an in-situ nasal gel
with improved residence time in the nasal cavity, achieving prolonged
drug release, quick onset of action by accessing the nose-to-brain route,
and assess its efficacy both in vitro and in vivo. The work also covers a
statistical approach in optimizing polymer concentration and study of
the same on viscosity and mucoadhesive strength which could in turn
affect the diffusion behaviour of the drug. These could benefit the pa­
tients suffering from migraine in real life without need of frequent and
high dosing and fast action of the drug via nose to brain. Challenges
would be the minimization of patient variability in human trials and
acceptance of this drug delivery system although there are nasal sprays
available in market, in-situ gels are yet to hit the market after due
approval from the regulatory authorities.
Material and methods
Materials
Carbopol 934P was supplied from Coral Pharma Chem (India).
Rizatriptan supplied was supplied from Sunrise remedies pvt ltd (India).
Various grades of HPMC polymer and Polyethylene glycol 400 were
procured via S D Fine Chem (India). All chemicals and solvents used
were of analytical grade.
Detachment weight(m)
Mucoadhesive strength (N) = Acceleration due to gravity (g)
Area exposed(A)
2
Results in Chemistry 6 (2023) 101010
Method for preparation of in-situ nasal gel
In-situ nasal gels prepared by cold technique [14,15]. Dissolve RZT
completely in distilled water to prepare 0.5 %w/v solution. Polymer
(Carbopol 934P in concentration range 1–2 %w/v) was added to the
above solution using a magnetic stirrer at 200 rpm to obtain the primary
solution. Copolymer (HPMC K4M in concentration range 0.4–1.2 %w/
v)) added into above primary solution and placed in cool conditions
overnight to obtain the final solution. The final solution was then sub­
jected to various evaluation tests.
Formulation optimization using design of experiment
It was decided from the preliminary trials to use a 32 full factorial
design in the present study to evaluate the effect of independent vari­
ables, i.e., concentration of carbopol 934P and HPMC K4M. In this
design, two factors were evaluated at three levels, and experimental
trials were performed for nine possible combinations. The concentration
of carbopol 934P (X1) and HPMCK4M (X2) were chosen as independent
variables in full factorial design, while viscosity (Y1) and mucoadhesive
strength (Y2) were taken as dependent variables. The coded and decoded
values of independent factors for all runs of 32 full factorial designs are
as in Table 1. The results were finally subjected to desirability function
and optimized batch selected based on that. The optimized batch was
developed and evaluated further. The concentration of drug in all nine
batches was 0.5 % w/v, volume of PEG kept constant at 1 mL, and final
volume up to 10 mL for each batch made up using distilled water.
Viscosity, pH and drug content of RZT in-situ gel
The viscosity, pH, and drug content of the formulated gels were
assessed in this study. To determine the pH of each formulation, a pH
meter was used, which was calibrated initially using buffer solutions of
pH 4 and pH 9. The viscosity of the developed formulations was
measured using a thermostatically controlled Brookfield viscometer.
The samples were sheared at 20-rpm with a suitable spindle no. 5 at a
temperature of 30 ◦ C; and the results were recorded. For drug content
determination, 0.5 g of formulation was taken in 50 mL volumetric flask
and diluted with 25 mL of SNF pH 6.4. The mixture was shaken for 10
min in an incubator to dissolve the drug in SNF pH 6.4. The solution was
then filtered and appropriately diluted to determine the drug content
spectrophotometrically, using a calibration curve plotted at 227 nm.
Mucoadhesive strength
In this study, we investigated the mucoadhesive properties of goat
nasal mucosa using a modified physical balance method. To simulate the
nasal environment, we wet the mucosa with simulated nasal fluid (SNF)
at a pH of 6.4. Next, we secured the mucosa to two glass slides and
applied in-situ gel between them by exerting a light fingertip force for
30 s. To adjust the physical balance, we placed a glass beaker on the
other side. We then added water from a burette and recorded the weight
of water needed to detach the two slides from each other. Mucoadhesive
strength was calculated using equation (1) [16,17].
(1)
K. Suhagiya et al. Results in Chemistry 6 (2023) 101010

Table 1
Coded and Decoded Value for 32 Factorial Design.
Coded value Decoded value
Batchcode Concentration of Carbopol Concentration of HPMC K4M Concentration of Carbopol(%w/v) Concentration of HPMC K4M (%w/v)
X1 X2 X1 X2

F1 − 1 − 1 1 0.4
F2 0 − 1 1.5 0.4
F3 +1 − 1 2 0.4
F4 − 1 0 1 0.8
F5 0 0 1.5 0.8
F6 +1 0 2 0.8
F7 − 1 +1 1 1.2
F8 0 +1 1.5 1.2
F9 +1 +1 2 1.2

Determination of gel strength
Gel strength was determined following method reported by Yong
et al. (2001); a 50-gram sample was placed in a 100-milliliter graduated
measuring cylinder and gelled in a thermostatically controlled water
bath at 37±◦ C [18]. This gel was formed by adding SNF to the sample.
To measure the gel strength, a weight of 35 g was placed onto a disk with
a diameter of 2.3 cm, a clearance from the sidewall of the cylinder of 0.4
cm, and a thickness of 0.5 cm. The gel strength was determined by
measuring the time (in seconds) required to move the piston 5 cm down
through the gel. The gel strength is influenced by factors such as mo­
lecular weight and degree of cross-linking. In cases where the apparatus
took more than five min to drop into the gel, additional weights were
added on top of the apparatus; and the gel strength was described by the
minimal weights required to push the apparatus 5 cm down through the
gel.
Ex vivo permeation study on nasal mucosa
In this research study, an ex vivo diffusion experiment was con­
ducted using freshly excised goat nasal mucosa, excluding the septum
portion, obtained from a local slaughterhouse in SNF (pH 6.4). To allow
tissue equilibration, the nasal mucosa was kept in SNF for 1 h. The su­
perior nasal concha was identified and separated from the nasal mem­
brane; and the excised superior nasal membrane was mounted onto a
Franz diffusion cell. To stabilize the tissue, SNF was added to both
compartments of the cell and stirred for 15 min using a magnetic stirrer.
After 15 min, the solutions from both compartments were removed, and
25 mL of fresh SNF was added to the receptor compartment. A volume of
0.5 mL of in-situ gelling solution was placed in the donor compartment
of Franz diffusion cell. Samples were withdrawn from receptor
compartment at predetermined time intervals and analyzed at 227 nm.
Stability studies
analysis. In 200 µl of brain, add 5-µl sodium peroxide vortex mix for 30
sec in Eppendorf tube. Then, add 400 µl di-chloro-methane: acetyl ac­
etate (1:4) for the extraction of RZT from brain tissue. Vortex mix for 60
sec and centrifuge at 15,000 RPM, collect 300 µl organic layer. To this
solution, add 100 µl of HPLC water to reconstitute it and inject it into
HPLC column. Chromatographic separations were achieved on an ODS
C8 column and, C8 (5 μm) Guard column. The column temperature was
maintained at 40 ◦ C. The mobile phase consisted of an isocratic solvent
system of acetonitrile: (water–triethanolamine) (30: (70:0.25), v/v), pH:
2.75 adjusted by using 80% phosphoric acid) at a flow rate of 1 mL/min.
Results and discussion
Analysis of viscosity, mucoadhesive strength, gelling time, gel strength and
drug content of design batches
Preliminary batches prepared using different grades of HPMC as
copolymer with RZT and carbopol 934P and subjected to viscosity and
drug content analysis. First, mix RZT and carbopol 934P, primary so­
lution was prepared and then HPMC E5 LV solution was added to it, and
there was precipitation due to changes in pH. With HPMC E15 LV, there
was very high viscosity like gel and after 2 hrs viscosity decreased and it
was like solution. RZT also precipitated out meanwhile. Next, with
HPMC E50 LV, again the problem of precipitation was evident. HPMC
k4M gave solution without precipitation and preliminary batches with
its different concentrations (0.3–0.8 %w/v) resulted in solution with
viscosity in the range of 1283 ± 1.89 cp − 2300 ± 1.5 cp and
mucoadhesive strength in the range of 0.1968 ± 0.014 N − 0.3767 ±
Table 2
Viscosity, mucoadhesive strength, gelling time, gel strength and drug content of
design batches.
Batch Viscosity Mucoadhesive Gelling Gel Drug
(cps) strength (N) time strength content
(sec) (sec) (%)

Short term accelerated stability study performed for optimized in- F1 1033 ± 0.2105 ± 0.003 8.6 ± 11.00 ± 97.96 ±
situ gelling solution at 40 ± 2 ◦ C temperature and 75 ± 5 % relative 0.763 0.23 1.73 0.63
F2 2217 ± 0.3142 ± 0.001 4.3 ± 29.33 ± 98.10 ±
humidity for one month and evaluated for drug content, ex vivo drug
0.763 0.20 0.57 1.35
diffusion, pH and viscosity. F3 3250 ± 0.4318 ± 0.004 2.3 ± 32.66 ± 98.32 ±
1.500 0.07 1.52 0.08
F4 1933 ± 0.3477 ± 0.004 5.6 ± 30.00 ± 97.52 ±
Protocol for determining RZT brain concentration in wister rats using
0.288 0.35 2.00 0.43
HPLC analysis F5 2416 ± 0.3687 ± 0.001 3.6 ± 38.33 ± 97.83 ±
0.288 0.24 0.57 0.40
The protocol described and executed in the present study was with F6 3167 ± 0.4578 ± 0.004 1.6 ± 42.00 ± 98.55 ±
the permission of the Committee for the Purpose of Control and Super­ 0.763 0.15 1.00 0.13
F7 2883 ± 0.3293 ± 0.0004 4.3 ± 36.67 ± 98.59 ±
vision of Experiments on Animals (CPCSEA), Ministry of Social Justice 0.280 0.19 1.52 0.69
and Empowerment (Protocol No. BKMGPC/IAEC19/RP22/2017). Wis­ F8 3100 ± 0.3462 ± 0.0100 4.6 ± 39.33 ± 98.86 ±
ter rats (150–180 gm) were housed in groups of 30 animals in cages and 1.500 0.87 0.58 0.27
maintained under standard conditions (12-hour light/dark cycle, 24℃, F9 4067 ± 0.4796 ± 0.0014 1.3 ± 51.33 ± 98.99 ±
1.040 0.04 1.52 0.20
and 35–60% humidity), with free access to pelleted diet and purified
drinking water. RZT brain concentration was determined by HPLC *Mean ± SD(n = 3).

3
K. Suhagiya et al.
0.01 N. HPMC K100M when used resulted in gel and not used further in
the study. Finally, HPMC K4M was selected as the copolymer with
carbopol 934P and different trial batches were prepared and evaluated.
Changes in polymer and copolymer concentrations appreciably affected
the viscosity of the formulation and mucoadhesive strength in pre­
liminary studies; therefore, the concentration of carbopol 934P and
HPMC K4M were selected as independent parameters to check the net
effect on viscosity and mucoadhesive strength of the prepared
formulations.
As per the design of experiment (32 factorial design), nine different
batches were prepared, evaluated and results recorded as in Table 2. The
pH of all formulations was in the range of 5.2 to 6.5, which is in the nasal
pH range. Gelling time varied from 1.3 to 8.6 sec and the changes are in
continuation of changes in mucoadhesive strength and gel strength. Low
gelling time was observed when viscosity was high and so was the
mucoadhesive strength and gel strength. F1 batch showed higher gelling
time as compared to other batches that was due to low concentrations of
polymer and copolymer. Drug content was above 97% in all batches so
independent variables have no effect on it and results were in an
acceptable range of 95–100%. Gel strength on the other hand was varied
from 11.00 ± 1.73 to 51.33 ± 1.52 sec but was in accordance with the
change in viscosity. The highest viscosity was of F9 i.e. 4067 ± 1.040 cps
while lowest was of F1 i.e. 0.2105 ± 0.003 cps and it is the case with gel
strength, highest for F9 and lowest for F1. Therefore, for statistical
optimization using factorial design, viscosity (Y1) and mucoadhesive
strength (Y2) were chosen as independent variables.
A statistical model incorporating interactive and polynomial terms
was used to evaluate the responses.
Y=b0+b1X1+b2X2+b11X1X1+b22X2X2+b12X1X2
The polynomial equations used to draw conclusions after considering
the magnitude of the coefficients and the mathematical sign (positive or
negative). R2 values for viscosity (cps), mucoadhesive strength (N) were
0.97758 and 0.948931, respectively, indicating a good correlation be­
tween dependent and independent variables. The results of ANOVA; F
calculated values for viscosity and mucoadhesive strength were 26.161
and 11.148, respectively (Table 3). The tabulated F value was found to
be 9.01. Calculated F values were greater than tabulated for both
dependent variables; therefore, selected factors have a significant effect
on dependent variables. From the results of regression analysis, both
independent variables had a statistically significant influence on
dependent variables as significance F value < 0.05.
For viscosity, the response surface plot (Fig. 1) revealed that a cor­
responding decrease in viscosity was observed with a decrease in the
concentration of Carbopol 934P and HPMC K4M and vice-versa. The
coefficient of X1 was higher than X2, which indicated that the effect of
concentration of Carbopol 934P was more significant than the effect of
concentration of HPMC. Only X1 and X2 were significant model terms
that affect the viscosity since their P value was < 0.05. The interaction
term, X1X2, and quadric terms, X1X1 and X2X2, were not significant. It
means that both polymer and copolymer affect viscosity individually
only and there was no synergistic effect.
Table 3
Summary of results of ANOVA.
Viscosity (cps)
Source of df SS MS F Significance
variation F Short-term accelerated stability study:
Regression 5 6,117,505 1,223,501 26.161 0.011
Residual 3 140301.1 46767.03
Total 8 6,257,806
Mucoadhesive strength (N)
Source of df SS MS F Significance
variation F (f2) for the percentage of drug diffusion. The results showed that f1 was
Regression 5 0.051 0.01 11.148 0.037
Residual 3 0.002 0.0009
Total 8 0.054
4
Results in Chemistry 6 (2023) 101010
The ideal combination of ingredients was chosen by meeting the
required standards while staying within the minimum and maximum
limits of formulation variables. To determine where the perfect blend
may be found, a comprehensive desirability function that considers all
the variables studied was employed. The desirable ranges in this func­
tion ranged from zero to one, indicating the least to the most desirable
variables. The optimized batch desirability was calculated according to
the process shown in Fig. 2. The target value for viscosity (Y1) was
determined from the preliminary study to be between 2400 and 2600
cps, which was sufficient to minimize burst release and maintain good
pourability for administration.
The target value for mucoadhesive strength (Y2) was also deter­
mined from the preliminary study as 0.3100–0.3800 N detachment
force, which was sufficient to stick to nasal mucosa. As a result, the
targeted values for viscosity and mucoadhesive strength were set at
2600cps and 0.3500 N, respectively. The optimized batch (OB) was
determined to have a composite desirability of 1, was made up of X1 =
1.27% w/v (Carbopol 934P concentration) and X2 = 1.03% w/v (HPMC
K4M concentration), as shown in Fig. 2.
Ex vivo permeation studies on nasal mucosa
The diffusion profile of all batches was compared; and the results are
presented in Fig. 3. These results indicate that polymer concentration
has significant impact on diffusion. For instance, F1 batch, which had
the lowest amounts of carbopol 934P and HPMC K4M; exhibited a
higher drug release of 94% within 4 hrs. Conversely, the F9 batch, which
had the highest amounts of these polymers, had a lower drug release of
55% within 8 hrs. As the concentration of polymer increased, the initial
burst release decreased due to the corresponding increase in viscosity.
The F1 batch; for example, had a 48.62% drug release in the first 30 min,
while the F9 batch had only 10.14%. In summary, these findings prove
that increasing the concentration of polymer and copolymer leads to
decreased, drug release and initial burst release. On kinetic modelling,
percentage cumulative drug permeated from In-situ nasal gel, r2 value
was found highest for Korsmeyer-Peppas model. N-value for Korsmeyer-
Peppas model was 0.420, which indicates fickian diffusion (Table 4).
The release mechanism from the polymers would be diffusion, after
swelling of polymers to develop gel. Swelling of the polymer structure
will facilitate the diffusion of drug, which ultimately will lead to the
diffusion of drug from the nasal membrane to the brain and exert a
therapeutic effect. Here, high molecular weight of the polymer is the
rate limiting step for their passage through the nasal membrane and
entry into the brain. It is important to note that higher polymer viscosity
can result in a denser and more compact matrix, reducing the diffusion
rate of drug molecules. This slower diffusion can lead to decreased burst
release and more controlled release profile over time. Hydrogels, can
absorb water and swell. The drug release from these systems can be
influenced by the polymer swelling capacity and its ability to retain the
drug within its structure. It was reported that the swelling capacity of
hydrogel increases with increase in carbopol 934P concentration [19]
and so is the viscosity. Higher polymer viscosity may enhance the
polymer’s ability to swell, forming a more retentive network that hin­
ders the burst release of the drug. HPMC K4M is frequently used to trap
drug particle and retard the release making it suitable for controlled
drug release [20].
Table 5 presents the results of a one-month stability study of the RZT
in-situ gel formulation, showing no significant changes in any of its
properties. To assess the stability of the formulation, the dissolution data
was used to calculate the dissimilarity factor (f1) and similarity factor
2.62 and f2 was 83.86, indicating that the drug diffusion data before and
after stability were equivalent. These findings confirm that the prepared
RZT in-situ gel was stable after the one-month period of stability.
K. Suhagiya et al. Results in Chemistry 6 (2023) 101010

Fig. 1. Response surface plot for dependent variables viscosity and mucoadhesive strength.

Isolated tissue sample (Brain)

The administration of RZT by various route, the Cmax of RZT in brain

is significantly higher (p < 0.05) in intra-nasal (340.27 ng) route as
compare to the oral, the maximum concentration of drug was released
by in-situ nasal gel within the 60 min due to this the onset of action
becomes fast. Direct access of brain by trigeminal nerve and olfactory

Table 4
Comparison of kinetic models.
Kinetic models R2
Zero order 0.675
First order 0.849
Higuchi 0.958
Fig. 2. Optimization by desirability function using Design expert10 (Desir­ Korsmeyer-Peppas 0.991 n = 0.420
ability = 1). Hixson-Crowell 0.818

Fig. 3. Ex vivo permeation studies on nasal mucosa (Batch F1-F9).

5
K. Suhagiya et al. Results in Chemistry 6 (2023) 101010

Table 5 0.97758 and 0.948931, respectively. Ex-vivo release study results sug­
Effect of stability study on various parameters of optimized batch. gested that as the concentration of polymer and copolymer increases,
Response After stability Before stability drug release and initial burst release (effect) decreases. The higher Cmax
and AUC values for the in-situ nasal gel suggest that this route of
Physical appearance White milky solution White milky solution
pH 6.1 6.1 administration could be more effective in delivering RZT to the brain.
Drug content (%) 99.26 ± 0.154 98.99 ± 0.204
Gelling time (sec) 5.66 ± 0.577 6.33 ± 0.771 CRediT authorship contribution statement
Gel strength (sec) 36.33 ± 0.356 37.33 ± 0.451
Viscosity (cps) 2633 ± 0.763 2516 ± 0.942
Mucoadhesive strength (N) 0.3546 ± 0.004 0.3438 ± 0.006 Kaushik Suhagiya: Conceptualization, Methodology, Data curation,
Diffusion at 5 min (%) 18.573 ± 0.173 23.791 ± 0.201 Writing – original draft. Chetan H. Borkhataria: Conceptualization,
Diffusion at 8 hr (%) 94.469 ± 0.168 93.373 ± 0.194 Methodology, Data curation, Writing – original draft. Sumit Gohil:
Methodology, Investigation, Writing – review & editing. Ravi A.
Manek: Investigation, Writing – review & editing. Kalpesh A. Patel:
Table 6 Resources, Writing – review & editing. Nilesh K. Patel: Resources,
Estimation of RZT concentration in brain (4 mg/Kg) at various time interval Writing – review & editing. Dhaval V. Patel: Supervision, Validation,
after I.V., oral & in-situ nasal gel administration. Writing – review & editing.
Sr. No Time (min) Concentration (ng/ml)
I.V. Oral In-situ nasal gel

1 5 106.83 ± 21.42 0 ± 0.0 165.56 ± 22.79 Declaration of Competing Interest

2 60 75.11 ± 18.84 41.55 ± 10.52 340.27 ± 31.23
3 120 51.68 ± 12.30 31.40 ± 12.79 250.00 ± 23.87 The authors declare that they have no known competing financial
4 180 43.78 ± 11.75 29.82 ± 8.57 238.66 ± 24.25 interests or personal relationships that could have appeared to influence
5 240 35.88 ± 9.63 28.25 ± 6.48 201.23 ± 29.13
6 300 34.35 ± 11.87 21.38 ± 6.27 197.90 ± 22.43
the work reported in this paper.
7 360 32.83 ± 10.86 14.51 ± 2.94 209.27 ± 19.68
8 420 23.31 ± 7.59 8.66 ± 3.42 164.46 ± 21.43 Data availability
9 480 15.69 ± 3.71 3.97 ± 1.64 128.62 ± 24.95
C max (ng) 106.832 41.401 340.27 ± 31.23
Data will be made available on request.
T max (min) 5 60 60
Total AUC(ng/ml × min) 338.73 171.26 1602.36
T1/2 (min) 211.56 225.01 440.61 References
Vd(L) 9.36 19.81 3.05
% Drug in brain 26 10.3 85 [1] B. Vigani, S. Rossi, G. Sandri, M.C. Bonferoni, C.M. Caramella, F. Ferrari, Recent
advances in the development of in situ gelling drug delivery systems for non-
Data are represented as (Mean ± SEM n = 3).
parenteral administration routes, Pharmaceutics 12 (9) (2020), https://doi.org/
= p < 0.0001, significant difference as compare to oral route. 10.3390/pharmaceutics12090859.
[2] N.G.N. Swamy, Z. Abbas, Mucoadhesive in situ gels as nasal drug delivery systems:
An overview, Asian J. Pharm. Sci. 7 (3) (2012).
bulb by nasal route of administration gives significantly higher (p < [3] T.J. Steiner, L.J. Stovner, R. Jensen, D. Uluduz, Z. Katsarava, Migraine remains
0.05) drug enrichment in to the brain as compare to the Oral route of second among the world’s causes of disability, and first among young women:
administration. From this study percentage drug in brain was signifi­ findings from GBD2019, J. Headache Pain 21 (1) (2020), https://doi.org/10.1186/
s10194-020-01208-0.
cantly higher (p < 0.05) in intra-nasal (85%) route as compare to the
[4] H. Lüllmann, K. Mohr, L. Hein (Eds.), Color Atlas of Pharmacology, Georg Thieme
oral route (10.3%). Verlag, 2017.
The Table 6 shows the estimation of RZT concentration in the brain [5] K. Selvaraj, K. Gowthamarajan, V.V.S.R. Karri, Nose to brain transport pathways an
at various time intervals after IV, oral, and in-situ nasal gel adminis­ overview: potential of nanostructured lipid carriers in nose to brain targeting, Artif
Cells Nanomed. Biotechnol. 46 (8) (2018), https://doi.org/10.1080/
tration in rats. The higher Cmax and AUC values for the in-situ nasal gel 21691401.2017.1420073.
suggest that this route of administration could be more effective in [6] F. Erdő, L.A. Bors, D. Farkas, Á. Bajza, S. Gizurarson, Evaluation of intranasal
delivering RZT to the brain. Mathure et al (2023) demonstrated the delivery route of drug administration for brain targeting, Brain Res. Bull. 143
(2018) 155–170.
similar findings with sumatriptan succinate by formulating liposomal [7] M. Dahlin, U. Bergman, B. Jansson, E. Björk, E. Brittebo, Transfer of dopamine in
thermosensitive in-situ nasal gel [12]. In another study enhanced brain the olfactory pathway following nasal administration in mice, Pharm. Res. 17 (6)
bioavailability of 131.17% was achieved for rasagiline mesylate using (2000), https://doi.org/10.1023/A:1007542618378.
[8] A. Kant, S. Reddy, M. M. S, J.S. V, C. N., In situ Gelling System - An overview,
transferosomal in-situ gel [21]. The in-situ nasal gel has the advantage of Pharmacologyonline 2 (1) (2011).
bypassing the blood–brain barrier, and the drug can directly enter the [9] A.K. Agrawal, M. Das, S. Jain, In situ gel systems as “smart” carriers for sustained
brain through the olfactory and trigeminal nerve pathways, which could ocular drug delivery, Expert. Opin. Drug Deliv. 9 (4) (2012) 383–402.
[10] Devi R, Chaudhary A, Pandit V. Mucoadhesive insitu nasal gel-A novel approach.
explain the higher brain concentrations seen with this route of admin­ J. Adv. Drug Deliv. Published online 2014.
istration [22]. [11] M. Singh, D. Dev, D.N. Prasad, A recent overview: in situ gel smart carriers for
ocular drug delivery, J. Drug Deliv. Ther. 11 (6-S) (2021) 195–205.
[12] D. Mathure, A.D. Sutar, H. Ranpise, A. Pawar, R. Awasthi, Preparation and
Conclusion
optimization of liposome containing thermosensitive in situ nasal hydrogel system
for brain delivery of sumatriptan succinate, Assay Drug Dev. Technol. 21 (1)
In this study, a design of experiments (32 factorial design) was used (2023) 3–16, https://doi.org/10.1089/adt.2022.088.
to prepare and evaluate nine different batches of the formulation, [13] E.M. Eissa, M.H. Elkomy, H.M. Eid, A.A. Ali, M.A.S. Abourehab, A.M. Alsubaiyel, I.
A. Naguib, I. Alsalahat, A.H. Hassan, Intranasal delivery of granisetron to the brain
varying the concentrations of carbopol 934P and HPMC K4M as inde­ via nanostructured cubosomes-based in situ gel for improved management of
pendent parameters to check their net effect on the viscosity and chemotherapy-induced emesis, Pharmaceutics 14 (7) (2022) 1374.
mucoadhesive strength of the prepared formulations. The data sug­ [14] U.C. Galgatte, A.B. Kumbhar, P.D. Chaudhari, Development of in situ gel for nasal
delivery: Design, optimization, in vitro and in vivo evaluation, Drug Deliv. 21 (1)
gested low gelling time when viscosity was high, and so was the (2014) 62–73.
mucoadhesive strength and gel strength. The statistical model used to [15] M.M. Omar, N.E. Eleraky, A.M. El Sisi, O.A. Hasan, Development and evaluation of
evaluate the responses incorporated interactive and polynomial terms in-situ nasal gel formulations of nanosized transferosomal sumatriptan: Design,
optimization, in vitro and in vivo evaluation, Drug Des. Devel Ther. (2019) 13,
and showed good correlation between dependent and independent https://doi.org/10.2147/DDDT.S235004.
variables. The R2 values for viscosity and mucoadhesive strength were [16] S. Shelke, S. Shahi, K. Jadhav, D. Dhamecha, R. Tiwari, H. Patil, Thermoreversible
nanoethosomal gel for the intranasal delivery of Eletriptan hydrobromide,

6
K. Suhagiya et al.
J. Mater. Sci. Mater. Med. 27 (6) (2016), https://doi.org/10.1007/s10856-016-
5713-6.
[17] A. Kempwade, A. Taranalli, Formulation and evaluation of thermoreversible,
mucoadhesive in situ intranasal gel of rizatriptan benzoate, J. Sol-Gel Sci. Technol.
72 (1) (2014) 43–48.
[18] C.S. Yong, J.S. Choi, Q.-Z. Quan, J.-D. Rhee, C.-K. Kim, S.-J. Lim, K.-M. Kim, P.-
S. Oh, H.-G. Choi, Effect of sodium chloride on the gelation temperature, gel
strength and bioadhesive force of poloxamer gels containing diclofenac sodium,
Int. J. Pharm. 226 (1-2) (2001) 195–205.
[19] A. Mahmood, A. Mahmood, M.A. Ibrahim, Z. Hussain, M.U. Ashraf, M.M. Salem-
Bekhit, I. Elbagory, Development and Evaluation of Sodium Alginate/Carbopol
934P-Co-Poly (Methacrylate) Hydrogels for Localized Drug Delivery, Polymers
(Basel) 15 (2) (2023) 311.
Results in Chemistry 6 (2023) 101010
[20] W. Hadinugroho, S. Martodihardjo, A. Fudholi, S. Riyanto, J. Prasetyo,
Hydroxypropyl Methylcellulose as Hydrogel Matrix and Citric Acid-Locust Bean
Gum as Negative Matrix for Controlled Release Tablet, ACS Omega 8 (8) (2023)
7767–7778, https://doi.org/10.1021/acsomega.2c07432.
[21] ElShagea HN, Makar RR, Salama AH, Elkasabgy NA, Basalious EB. Investigating the
Targeting Power to Brain Tissues of Intranasal Rasagiline Mesylate-Loaded
Transferosomal In Situ Gel for Efficient Treatment of Parkinson’s Disease.
Pharmaceutics. 2023;15(2). doi:10.3390/pharmaceutics15020533.
[22] S.V. Dhuria, L.R. Hanson, W.H. Frey, Intranasal delivery to the central nervous
system: mechanisms and experimental considerations, J. Pharm. Sci. 99 (4) (2010)
1654–1673, https://doi.org/10.1002/JPS.21924.