Article Not peer-reviewed version

Nanostructured Emulsomes for the

Enhanced Delivery of Cabazitaxel: A
Novel Approach to Cancer Treatment
and Drug Release Profiling

Ramya Teja Medarametla * , J. N. Suresh Kumar , Venkata Gopaiah Kurra , D. Naveen Reddy , B. Venkamma ,
G. Anand Babu , G. Raghavendra , M. Shaggir

Posted Date: 3 June 2025

doi: 10.20944/preprints202505.2468.v1

Keywords: emulsomes; lipophilic; cabazitaxel; pegylation; drug release kinetics

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Article

Nanostructured Emulsomes for the Enhanced

Delivery of Cabazitaxel: A Novel Approach to Cancer
Treatment and Drug Release Profiling
Ramya Teja Medarametla*, J. N. Suresh Kumar, K. Venkata Gopaiah, D. Naveen Reddy,
B. Venkamma, G. Anand Babu, G. Raghavendra and M. Shaggir

Narasaraopeta Institute of Pharmaceutical Sciences, Narasaraopet, Palnadu

* Correspondence: ramyatejamedarametla@[Link]

Abstract: Objective: The present study aimed to develop and optimize a Cabazitaxel (CTX)-loaded
sterically stabilized emulsome (SS-ES) system to enhance drug stability and achieve sustained release,
using tristearin as the core lipid via the solvent evaporation method. Methods: Two emulsome
formulations were developed: a simple emulsome (P-ES) and a sterically stabilized emulsome (SS-
ES). Key formulation variables included the lipid-to-stabilizer ratio (DSPC, CHOL, and DSPE-PEG),
the molar ratio of phospholipids to PEG-lipid components, solid lipid content, phospholipid
concentration, and the ratio of organic to aqueous phase. The prepared formulations were
characterized for particle size, release kinetics, and pH sensitivity. Results: The mean particle size of
P-ES was 275 ± 5.52 nm, while SS-ES exhibited a smaller size of 195 ± 6.4 nm, indicating improved
nano-sizing with PEGylation and steric stabilization. Both formulations demonstrated pH-sensitive
and sustained drug release profiles, with SS-ES showing a slower and more controlled release
compared to P-ES. Release kinetics followed a Fickian diffusion mechanism, best fitting the Higuchi
model. The SS-ES formulation exhibited enhanced physical stability and potential for prolonged
systemic circulation. Conclusion: Sterically stabilized emulsomes represent a promising nanocarrier
platform for the delivery of hydrophobic drugs like Cabazitaxel, offering enhanced stability,
sustained release, and potential for prolonged bioavailability.

Keywords: emulsomes; lipophilic; cabazitaxel; pegylation; drug release kinetics

Introduction
The emergence of nanotechnology has revolutionized drug delivery strategies by providing
innovative carrier systems that can be engineered to align with the specific physicochemical
properties of therapeutic compounds. Nanocarriers, with their tunable surface characteristics and
internal structures, offer enhanced delivery options for both small-molecule drugs and biologics. In
recent years, research has emphasized the design of tumour-specific nanocarriers to minimize
systemic exposure and reduce unintended side effects1.
One of the critical limitations associated with many chemotherapeutic agents is their poor
aqueous solubility, which hampers formulation development and diminishes therapeutic
effectiveness. Traditional solubilization techniques, such as the use of organic solvents and
surfactants, often provoke adverse immune reactions or toxicity. To address these issues,
nanoparticulate systems—especially those based on lipids and polymers—have been explored to
enhance solubility, safeguard the drug molecule, and eliminate the need for harmful excipients 2.
Cabazitaxel (Ctx), a second-generation taxane used in cancer treatment, is an example of a
hydrophobic drug that presents formulation challenges. The commercial formulation, Taxol® ,
utilizes a combination of Cremophor EL and ethanol, which is linked to significant toxicity. As a safer
alternative, emulsome-based delivery systems have gained interest due to their ability to encapsulate
hydrophobic drugs without the use of aggressive solubilizers.

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Emulsomes are hybrid systems that integrate the properties of liposomes and solid lipid
nanoparticles. They feature a lipid-based solid core surrounded by a phospholipid bilayer, making
them suitable for high drug loading and improved physical stability. PEGylation, achieved by
incorporating polyethylene glycol (PEG)-linked lipids, further enhances circulation time by reducing
recognition and clearance by the immune system. Moreover, these systems can be tailored for
controlled and sustained drug release, which can potentially improve therapeutic outcomes and
patient adherence3,4.
The present study focuses on the development and optimization of PEGylated emulsomes, also
known as sterically stabilized emulsomes (SS-Es), encapsulating Cabazitaxel. The goal was to
evaluate their potential as a long-circulating and biocompatible nanocarrier for anticancer therapy.
Key physicochemical properties and drug release kinetics were investigated to determine their
suitability for clinical application.

Materials and Methods

Materials
Cabazitaxel was generously supplied by Neon Laboratories Ltd., Mumbai, India. Lipid
components including DSPC (1,2-distearoyl-sn-glycero-3-phosphocholine), cholesterol, and DSPE-
PEG (PEGylated phospholipid) were purchased from Lipoid GmbH, Germany. Tristearin was
sourced from Sigma-Aldrich, Germany. All other chemicals used were of analytical grade and
procured locally.

Preparation of Emulsomes

Both plain emulsomes (P-Es) and PEGylated emulsomes (SS-Es) were prepared using a modified
single-step emulsification technique followed by solvent evaporation. Tristearin was utilized as the
solid lipid forming the core, while DSPC and cholesterol comprised the lipid bilayer. For steric
stabilization, DSPE-PEG was included in selected formulations.
The process began by melting the lipid phase at 80 °C, followed by the addition of Cabazitaxel
pre-dissolved in aqueous ethanol. The mixture was stirred at 1000 rpm for 15 minutes to ensure
uniform distribution. In parallel, the phospholipids (with or without PEGylation) were dispersed in
pre-heated distilled water and mixed at 5000 rpm. The lipid solution was then added gradually (1
ml/min) to the aqueous phase under continuous stirring at 80 °C and homogenized at 10,000 rpm for
30 minutes6,7.
Rapid cooling using an ice bath facilitated the formation of a phospholipid bilayer around the
solid lipid core. Residual solvents and unencapsulated drug were removed via dialysis against
distilled water using a 10 kDa molecular weight cutoff membrane. The final product was sterile-
filtered (0.22 µ m), cryoprotected with 5% sucrose, and lyophilized for storage at 4 °C. The
composition of the optimised formulation is stated in Table 1.

Table 1. Composition of optimized formulations.

SL
Formulation PL: DSPE-PEG PL: SL Organic: aqueous Homogenization
concentration
code (w/w) (w/w) phase volume (v/v) time (min)
(%)
P-Es - 1:1 1:10 10 15
SS-Es 1:3 1:1 1:10 10 15
P-Es: Plain emulsomes, SS-Es: Sterically stabilized emulsomes, PL: phospholipid, DSPE-PEG: 1,2-Distearoyl-
sn-glycero-3-phosphoethanolamine- Poly(ethylene glycol), SL: Solid lipid.

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Optimization Parameters

To achieve optimal characteristics such as particle size and entrapment efficiency, the following
formulation parameters were systematically varied:
• Ratio of phospholipids to solid lipid (PL:SL)
• Molar ratio of lipid to PEG-lipid
• Concentration of both lipid and phospholipid components
• Volume ratio of organic to aqueous phase
• Homogenization time

Characterization of Emulsomes

Particle Size and Distribution9,10

Dynamic light scattering (DLS) was employed to measure particle size and polydispersity index
(PDI) using a Nanoplus 5.01 Zetasizer. Samples were diluted (1:9) in deionized water. Low PDI
values indicated homogeneity and colloidal stability.

Zeta Potential

Surface charge was analyzed to assess colloidal stability and predict in vivo interactions. Zeta
potential measurements were performed using the same instrument in flow-through mode.

Morphological Studies

Transmission electron microscopy (TEM) was used to assess the vesicle morphology, including
shape and surface texture. A droplet of the formulation was placed on a copper grid, air-dried, and
visualized under a Tecnai G2 TEM at 100 kV. Scanning electron microscopy (SEM, NOVA NanoSEM
450) provided additional 3D structural insights.

Entrapment Efficiency11

Drug entrapment was measured using a gel filtration technique involving Sephadex G-50
columns. Sephadex (1.2 g) was pre-swollen in saline for 5 hours and stored at 4 °C. Mini-columns
were prepared using disposable syringes and packed with the hydrated gel. A 0.2 ml sample of the
emulsomal formulation was applied and centrifuged at 2000 rpm for 3 minutes. The eluted fraction,
containing encapsulated drug, was collected. The emulsomes were lysed with 1 ml of 0.1% Triton X-
100, and Cabazitaxel concentration was measured at 229 nm using a UV-Vis spectrophotometer.
Entrapment efficiency was calculated using the following formula:
Entrapment Efficiency (%) = (Total drug – Free drug) / Total drug ×100

In Vitro Drug Release Study12,13

Drug release profiles were determined using dialysis in phosphate buffer (pH 4.0) at 37 ± 1 °C.
A 1 ml aliquot of the drug-loaded emulsomes was enclosed in a dialysis membrane and placed in 20
ml buffer. Samples were taken at defined intervals for 24 hours, with the buffer volume maintained
by replacing each withdrawn sample with fresh medium. Drug concentration was quantified
spectrophotometrically at 229 nm.

Drug Release Kinetics14

Release data were analyzed using different kinetic models, including zero-order, first-order,
Higuchi, and Korsmeyer-Peppas equations. The best-fitting model was determined based on
correlation coefficients (R²), and the diffusion exponent (n) from the Korsmeyer-Peppas model
provided insight into the release mechanism, distinguishing between diffusion-controlled and
anomalous transport.

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Plasma Protein Binding Assay

To assess the stealth characteristics conferred by PEGylation, emulsomes were incubated in a

5% BSA solution. Changes in particle size and PDI were monitored using DLS. Minimal variation in
these parameters indicated effective PEG shielding and reduced protein adsorption.

Statistical Analysis

All data were expressed as mean ± standard deviation (n=3). Statistical analysis was conducted
using one-way ANOVA followed by Tukey–Kramer post hoc tests via GraphPad Prism (version
3.00). A p-value less than 0.05 indicated statistical significance. (Table 2)

Table 2. Physicochemical properties of optimized formulations.

Formulation code Size (nm) PI Zeta potential (mV) %EE

P-Es 275±5.52* 0.112±0.011** -30.5±2.0* 72.54±3.41*
SS-Es 195±6.4** 0.124±0.012** -35.8±2.1* 75.56±3.25**
Data represents mean ± SD (n=3), *Statistically significant difference at *p<0.05,** Statistically significant
difference at p<0.01.

Formulation Variables in Emulsomes Optimization15

To optimize the emulsomal formulation for effective Cabazitaxel delivery, several key
formulation parameters were systematically evaluated. These included the molar ratio of
phospholipids to DSPE-PEG, the total lipid-to-solid lipid mass ratio (DSPC, cholesterol, and DSPE-
PEG to tristearin), the concentrations of tristearin and phospholipids, and the volume ratio between
the organic and aqueous phases. Each of these factors was analysed for its influence on particle size
(PS) and drug entrapment efficiency (%EE), to develop a stable Nano system capable of efficiently
encapsulating Cabazitaxel in a size range suitable for intravenous delivery.

Impact of Phospholipid to DSPE-PEG Molar Ratio on Nanoparticle Characteristics16,17

DSPE-PEG is incorporated into emulsomes to enhance their stability by providing a steric shield
that limits particle aggregation and promotes long circulation. In formulations lacking DSPE-PEG
(referred to as plain emulsomes), the average particle size was relatively large, measured at
320.5 ± 5.8 nm (Figure 1). Introducing DSPE-PEG led to a progressive decrease in particle size, with
the most significant reduction observed at a phospholipid to DSPE-PEG molar ratio of 1:5. At this
ratio, the particle size was minimized to 190.5 ± 4.77 nm. A slightly higher size of 195.5 ± 5.6 nm was
noted at the 1:3 ratio, which still represented a statistically significant reduction compared to non-
PEGylated emulsomes (p < 0.01).
Beyond the 1:3 molar ratio, increasing the DSPE-PEG content further had minimal additional
effect on reducing particle size, indicating that 1:3 was the optimal ratio for this formulation. The size
reduction is attributed to the steric stabilization effect provided by the hydrophilic PEG chains, which
create a repulsive barrier between particles and prevent their agglomeration during formation.
This optimized formulation also achieved a high entrapment efficiency, reaching 78.20 ± 2.8%
(Figure 1A, 1B), suggesting that PEGylation not only stabilizes the particles but also promotes
effective drug incorporation. A low polydispersity index (PDI) was observed, indicating a narrow
size distribution and uniformity of the emulsomes. Zeta potential analysis revealed a surface charge
of around –32 mV, which is consistent with stable colloidal systems. The high negative charge
contributes to repulsive interactions between particles, preventing aggregation and thereby
enhancing the formulation’s shelf life.

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Figure 1. A. Graphs showing the effect of PL to DSPE-PEG ratio on size and entrapment efficiency of
emulsomes.

Figure 1. B. Plot showing the change in zeta potential of emulsomes on the addition of DSPE-PEG to PL. Data
represents mean ± SD (n=3).

Effect of Organic-to-Aqueous Phase Volume Ratio18,19

Another critical variable in emulsomes preparation is the ratio between the volumes of organic
and aqueous phases used during emulsification. In this study, the organic phase volume was held
constant while the volume of the aqueous phase was varied between 5 and 20 ml. Initial increases in
aqueous volume improved dispersion efficiency, leading to a notable reduction in particle size. This
effect can be attributed to the enhanced dilution environment, which minimizes droplet collisions
and supports better emulsification.
However, when the aqueous phase volume exceeded a certain threshold, an opposite trend was
observed—particle size began to increase. This is likely due to a dilution-related reduction in shear
energy during homogenization, which diminishes the capacity to break lipid droplets into finer
particles. Furthermore, excessive dilution can destabilize emulsification and promote the formation
of larger aggregates.
Entrapment efficiency also followed a similar trend. As particle size decreased with increasing
aqueous volume, %EE initially improved. However, beyond the optimal point, further increases in
aqueous phase volume led to a decline in drug loading. This drop could be due to the increased
diffusion of the drug into the surrounding aqueous medium during processing or loss during solvent
evaporation, which reduces the proportion of drug retained in the lipid matrix.

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When comparing plain emulsomes (P-Es) and sterically stabilized emulsomes (SS-Es), both
showed a decrease in particle size as the organic-to-aqueous phase ratio increased from 1:2 to 1:10.
For P-Es, the size decreased from 270.5 ± 2.5 nm to 230.2 ± 3.5 nm. The SS-Es exhibited an even greater
reduction, from 256.5 ± 2.6 nm to 195.5 ± 3.28 nm (Figure 2). However, beyond this 1:10 ratio, further
increases in aqueous phase led to a size increase, likely due to inefficient emulsification at lower
energy densities. Based on these observations, an organic-to-aqueous phase ratio of 1:10 was
considered optimal for achieving small, stable nanoparticles with desirable drug loading
characteristics. At this optimized ratio, the entrapment efficiency was observed to be 70.25 ± 4.4% for
P-Es and 75.50 ± 2.5% for SS-Es, indicating effective drug loading in both formulations, with slightly
superior performance in the PEGylated emulsomes.

Figure 2. Graph showing the effect of organic to aqueous phase volume on size and entrapment efficiency of
emulsomes. Data represents mean ± SD (n=3).

Effect of Phospholipid to Solid Lipid (PL:SL) Ratio on Particle Size and Entrapment Efficiency20,21
The phospholipid-to-solid lipid ratio (PL:SL) plays a pivotal role in determining the physical
characteristics and stability of emulsomal formulations. Varying this ratio from 0.25:1 to 1.5:1 led to
a significant decrease in particle size across both plain emulsomes (P-Es) and sterically stabilized
emulsomes (SS-Es). For P-Es, particle size was reduced from 650.5 ± 1.5 nm to 310.2 ± 2.5 nm, while
SS-Es showed a size reduction from 556.4 ± 3.5 nm to 235.5 ± 3.6 nm, indicating improved dispersion
and structural uniformity with increasing phospholipid content.
At the extreme of a 0:1 PL:SL ratio—meaning the absence of phospholipids—the lipid-based
particles lacked structural integrity, resulting in aggregation and significantly larger particle sizes
(approximately 840.5 ± 3.7 nm for P-Es and 650.8 ± 4.5 nm for SS-Es) as shown in Figure 3A. This
observation emphasizes the essential role of phospholipids in stabilizing the emulsomal shell and
maintaining nanoscale particle dimensions.
In formulations with low PL:SL ratios (0.25:1 to 0.5:1), the produced emulsomes were
comparatively larger and exhibited low zeta potential values (Figure 3B), which suggests partial or
insufficient encapsulation of the solid lipid core. This incomplete coating likely resulted in unstable
particles that were prone to aggregation due to limited steric or electrostatic repulsion and stronger
attractive forces between particles.
Improved formulation characteristics were observed near a 1:1 PL: SL ratio. At this ratio, particle
sizes were minimized (310.2 ± 2.5 nm for P-Es and 235.5 ± 3.6 nm for SS-Es), and the zeta potential
values reached –32 ± 0.12 mV and –35 ± 0.15 mV, respectively. These surface charge values are
generally considered ideal for ensuring colloidal stability, as particles with zeta potentials exceeding
±30 mV are less likely to aggregate due to increased electrostatic repulsion.
The enhanced stability at this ratio is likely due to the effective encapsulation of the lipid core
by the phospholipid shell, which reduces interfacial tension and supports the formation of uniformly

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dispersed nanoparticles. As a result, the emulsomes exhibit improved physical properties, including
smaller size, better dispersion, and higher entrapment efficiency.
However, exceeding the optimal phospholipid concentration resulted in diminishing returns.
With a total lipid content of 30%, excessive phospholipids that do not integrate into the emulsomal
matrix may form separate vesicular structures such as liposomes, particularly when the phospholipid
concentration surpasses its critical micelle concentration (CMC ~0.4 mg/ml). This leads to an increase
in particle size and a decline in zeta potential, signaling reduced formulation stability22.
Conversely, insufficient phospholipid content results in inadequate surface coverage of the lipid
core, compromising particle integrity despite potentially high surface charge values. This trade-off
highlights the importance of balance in formulation design was shown in graphical data in Figure-
3A, 3B & also in Figure- 4.

Figure 3. A. Graphs showing the effect of PL to SL ratio on size and entrapment efficiency of emulsomes. B. Plot
showing the change in the zeta potential of emulsomes with PL: SL ratio. Data represents mean ± SD (n=3).

Figure 4. Graphs showing the effect of SL concentration on particle size and entrapment efficiency of emulsomes.
Data represents mean ± SD (n=3).

Impact of Solid Lipid Concentration on Particle Size and Entrapment Efficiency23,24

The concentration of solid lipid was found to significantly influence both particle size and drug
entrapment within the emulsomal formulations. As the solid lipid content was increased from 5% to
20% w/w, a notable rise in particle size was observed. In the case of plain emulsomes (P-Es), particle
diameter increased from 198.5 ± 4.5 nm to 380.4 ± 2.4 nm, while for sterically stabilized emulsomes
(SS-Es), size grew from 182.3 ± 3.5 nm to 358.5 ± 1.5 nm (Figure 4). This size enlargement can be
attributed to the elevated viscosity of the organic phase at higher lipid concentrations, which slows
down the diffusion of lipids into the aqueous phase during formulation.

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When solid lipid concentration exceeded 10% w/w, an imbalance between the lipid core and the
phospholipid shell was observed. The available phospholipids were insufficient to fully encapsulate
the expanding lipid content, leading to particle coalescence, structural instability, and an associated
increase in size. Additionally, these higher lipid levels reduced the system’s ability to efficiently trap
the drug, resulting in a drop in entrapment efficiency of 35.8±2.1, respectively (Figure 7C and D).
Conversely, increasing solid lipid content initially led to improved drug entrapment. The greater
viscosity of the medium at higher lipid levels slowed the diffusion of Cabazitaxel (Ctx) into the
surrounding aqueous phase, facilitating greater retention within the hydrophobic lipid core. These
results point to a strong affinity between Ctx and the solid lipid matrix, particularly tristearin, which
contributed to enhanced drug loading.

Effect of Homogenization Time on Particle Size and Entrapment Efficiency

Homogenization time is a key processing parameter that directly affects the particle size
distribution and entrapment characteristics of emulsomes. This study examined the effect of varying
homogenization times between 5 and 15 minutes. It was observed that increasing homogenization
time led to a gradual reduction in average particle size and improved uniformity of particle
distribution (Figure 5). This can be explained by the higher energy input over time, which facilitates
the formation of smaller and more evenly dispersed lipid droplets.
However, extending homogenization beyond the optimal point caused a reversal in this trend.
Longer durations introduced excessive turbulence into the system, resulting in increased particle
collisions and aggregation. This effect compromised the structural integrity of the particles and
reduced the drug’s entrapment efficiency.
Optimal conditions were achieved at a homogenization time that produced particle sizes of
250.5 ± 2.5 nm for P-Es and 215.6 ± 2.3 nm for SS-Es. At this point, entrapment efficiencies reached
76.4 ± 3.2% and 80.2 ± 2.8% for P-Es and SS-Es, respectively, indicating enhanced formulation
performance at these conditions.

Physicochemical Characterization of Optimized Emulsomes25,26

Following optimization of formulation and process parameters, the emulsomes were subjected
to further physicochemical analysis. Key attributes assessed included particle size, morphology, and
surface charge (zeta potential), all of which are critical for evaluating colloidal stability and drug
delivery potential.
Imaging studies using Transmission Electron Microscopy (TEM) and Scanning Electron
Microscopy (SEM) confirmed the spherical nature of the emulsomes (Figures 6A and 6B). The
uniformity in particle size was further supported by low polydispersity index values, reflecting a
narrow size distribution. Mean particle sizes were recorded as 275 ± 5.52 nm for P-Es and 195 ± 6.4 nm
for SS-Es (Figures 7A and 7B), consistent with earlier observations.
Surface charge analysis revealed that both P-Es and SS-Es carried a zeta potential of
approximately –30.5 ± 2.0 mV. These values indicate strong repulsive forces between particles,
minimizing aggregation and supporting the stability of the formulations during storage.

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Figure 5. Graphs showing the effect of homogenization time on the particle size of emulsomes, data represents
mean ± SD (n=3).

Figure 6. A. TEM micrograph of SS-Es. The outer light periphery indicated by the arrow shows the presence of
PEG on the surface of the emulsomes. B. SEM micrograph of SS-Es.

Figure 7. Plots showing the size distribution of A. P-Es; B. SS-Es and zeta potential value of C. P-Es; D. SS-Es.

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In Vitro Drug Release27,28

One of the essential attributes of a successful anticancer drug delivery system is its ability to
remain stable in physiological conditions while preferentially releasing the drug in the more acidic
tumour microenvironment. To examine this, in vitro release studies were performed at two pH levels:
pH 7.4 to simulate normal physiological conditions, and pH 4.0 to mimic the acidic conditions
typically found in tumour tissues.
Both plain emulsomes (P-Es) and sterically stabilized emulsomes (SS-Es) demonstrated similar
release trends under both pH environments, suggesting that the incorporation of lipids and
PEGylated components did not drastically alter the drug's release mechanism. At pH 7.4, drug release
over the first 24 hours was measured at approximately 40.5 ± 2.7% for P-Es and 32.5 ± 1.5% for SS-Es
(Figure 7A). This initial phase of faster release is believed to result from the liberation of the drug
associated with the phospholipid layer. Subsequently, a more sustained release phase was observed,
extending over the following six days. During this extended period, cumulative release reached
75.50 ± 4.5% for P-Es and 66.80 ± 3.5% for SS-Es..
The slower release rate from SS-Es compared to P-Es is attributed to the presence of the PEG
coating, which provides structural reinforcement and acts as a barrier, limiting the diffusion of water
and reducing solvent access to the lipid matrix. This barrier effect contributes to a more controlled
and prolonged release profile.
When tested at acidic pH (4.0), both formulations exhibited a noticeably faster drug release.
Within the first 24 hours, P-Es released 52.5 ± 1.5% of the encapsulated drug, while SS-Es released
48.6 ± 2.5%. This accelerated release behavior in an acidic environment indicates a pH-sensitive
release mechanism. The reduced release rate at physiological pH helps maintain the drug within the
emulsomal carrier, which could reduce off-target toxicity and enhance safety.
The enhanced drug release under acidic conditions can be explained by the partial disruption of
the phospholipid bilayer and increased permeability of the emulsome membrane at lower pH. Acidic
conditions may also promote hydrolytic degradation of the lipid matrix, especially tristearin,
resulting in faster drug diffusion. This behavior supports the system’s potential for site-specific
release in the tumour environment29,30.

Figure 8(a). Plot showing the in vitro drug release kinetics as per the various mathematical models. A. Zero-order
drug release model. B. First-order drug release model.

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Figure 8 (b). Plot showing the in vitro drug release kinetics as per the various mathematical models. C. Higuchi
model D. Korsmeyer-peppas model. Data represents mean ± SD (n=3).

Drug Release Kinetics Study31,32

To elucidate the release behaviour of Cabazitaxel (Ctx) from emulsomal formulations, in vitro
cumulative release data were analysed using multiple kinetic models. This analysis aimed to
determine the underlying mechanism controlling drug release. The models employed included zero-
order, first-order, Higuchi, and Korsmeyer-Peppas, each represented by specific plots: cumulative
percentage of drug released (Q) versus time (t), log cumulative drug remaining versus time, Q versus
the square root of time (√t), and log Q versus log time (log t), respectively (Figure 8A & 8B).
The correlation coefficients (R²) derived from the linear regression of each model were compared
to identify the best-fitting kinetic profile (Table 3). Among these, the Higuchi model showed the
highest correlation, indicating that the drug release from emulsomes is primarily governed by a
diffusion-controlled mechanism.
To further characterize the nature of diffusion, the Korsmeyer-Peppas model was applied. The
diffusional exponent ‘n’, determined from the slope of the log-log plot, was found to be less than 0.5.
This low ‘n’ value confirms that the drug release conforms to Fickian diffusion, suggesting that the
release process is dominated by the passive diffusion of the drug through the emulsomal matrix.
These findings highlight that the emulsomes provide a sustained release platform where the
drug diffuses gradually over time, supporting their potential as a controlled drug delivery system for
Cabazitaxel with parameters for kinetic models observed given in table-3.

Table 3. Parameters for kinetic models applied to in vitro drug release data from emulsomes formulations.

Zero
Formulation First order Higuchi Korsmeyer-peppas
order
R2 K R2 K R2 K R2 n
P-Es (pH 7.4) 0.8875 0.3011 0.9512 0.0025 0.9947 4.4170 0.9766 0.3252
SS-Es (pH 7.4) 0.9470 0.3166 0.9755 0.0024 0.9912 4.5573 0.9925 0.3942
P-Es (pH 4.0) 0.9128 0.3255 0.9766 0.0033 0.9958 5.5072 0.9968 0.2659
SS-Es (pH 4.0) 0.9383 0.2860 0.9755 0.0022 0.9997 4.3476 0.9858 0.2730
R: regression coefficient. K: release constant, n: diffusion coefficient.

Plasma Protein Absorption Study33,34

Evaluation of Stealth Properties Against Protein Adsorption

One of the primary obstacles in designing long-circulating nanoparticle systems is their rapid
clearance by the Reticuloendothelial System (RES), triggered by the adsorption of opsonin proteins
onto the nanoparticle surface. This opsonization process flags the particles for uptake by

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macrophages through phagocytosis. To mitigate this, DSPE-PEG was incorporated into the
phospholipid shell of the emulsomes to provide a stealth characteristic, which helps evade immune
detection and enhances systemic persistence.
To examine this stealth effect, the emulsomal formulations were incubated in a 5% bovine serum
albumin (BSA) solution, with distilled water serving as the control medium. Changes in particle size
and distribution were monitored over time to evaluate protein interaction and aggregation
behaviour.
Following 24 hours in distilled water, the particle sizes of both plain emulsomes (P-Es) and
sterically stabilized emulsomes (SS-Es) remained consistent, indicating good baseline stability.
However, when exposed to BSA, a difference in protein interaction became evident. After 12 hours
of incubation, both formulations maintained stable sizes, suggesting minimal initial protein
adsorption. By 24 hours, P-Es exhibited a significant size increase, reaching approximately
575 ± 2.5 nm, indicative of substantial protein binding and possible aggregation.
In contrast, SS-Es displayed only a slight increase in size, confirming that the PEGylated surface
effectively minimized protein adhesion. This resistance to protein adsorption supports the role of the
PEG layer as a steric shield that reduces opsonin binding and protects the emulsomes from RES-
mediated clearance. Consequently, this stealth behavior contributes to extended circulation time and
may enhance the therapeutic potential of the drug delivery system was also shown in Figure 9..

Figure 9. Change in size of emulsomes on incubation with 5%BSA, data represents mean ± SD (n=3).

Discussion
The formulation of an effective delivery system for Cabazitaxel (CTX) remains a crucial
pharmaceutical challenge due to its low aqueous solubility, high systemic toxicity, and poor oral
bioavailability. Traditional formulations, such as those using Cremophor EL, have been associated
with significant hypersensitivity reactions, prompting the search for safer and more efficient
alternatives. In this context, Compritol-based emulsomes offer a novel and biocompatible approach
that addresses many of these limitations.
Rationale for Using Emulsomes
Emulsomes combine the structural advantages of liposomes and solid lipid nanoparticles
(SLNs). By encapsulating CTX in a solid lipid core (Compritol or tristearin) and surrounding it with
a phospholipid bilayer, the system not only increases the drug’s solubility but also improves its
stability and controlled release properties. Unlike conventional systems, these emulsomes were
designed without additional surfactants or co-solvents, reducing the risk of toxicity and enhancing
patient safety.

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Formulation and Physicochemical Characteristics

The use of a modified solvent evaporation technique proved efficient in producing reproducible
formulations with uniform particle size, high drug entrapment efficiency, and stable zeta potential.
The mean diameters of the simple and stabilized emulsomes-275 ± 5.52 nm (P-ES) and 195 ± 6.4 nm
(SS-ES), respectively—fell within the optimal nanoscale range for passive tumor targeting via the
enhanced permeability and retention (EPR) effect.
The smaller particle size and tighter distribution of SS-ES suggest that PEGylation and steric
stabilization not only enhance physical stability but also contribute to more predictable
pharmacokinetics. The phospholipid to PEG-lipid molar ratio and lipid/stabilizer ratios were critical
in modulating particle size and stability, underscoring the importance of careful optimization in
formulation development.
Drug Release and Mechanism
Both formulations exhibited pH-sensitive, sustained release profiles, which are ideal for cancer
therapy. The acidic tumor microenvironment facilitated drug release, while physiological pH
conditions maintained drug encapsulation, minimizing off-target toxicity. The release kinetics
followed the Higuchi model, indicating a Fickian diffusion-controlled mechanism—suggesting that
the drug was primarily released from the matrix through diffusion, a desirable trait for sustained
therapeutic action.
The slower release from SS-ES compared to P-ES can be attributed to the stealth characteristics
imparted by PEGylation, which may also result in reduced opsonization and extended systemic
circulation. This is essential for allowing the drug carrier to accumulate at the tumor site before
releasing its payload.
Therapeutic Implications and Biocompatibility
By eliminating harmful excipients like Cremophor EL, the developed emulsome system
enhances biocompatibility and reduces the risk of immune-related adverse effects. The PEG layer,
aside from extending circulation time, opens up possibilities for active targeting via ligand
conjugation (e.g., folate, transferrin, or antibodies), potentially enabling receptor-mediated
endocytosis in specific cancer cells.
Additionally, the modular nature of the emulsome design supports the co-encapsulation of
multiple drugs, gene delivery components, or imaging agents, making it a multifunctional
nanoplatform for combination therapy or theranostic applications.
Comparative Advantage and Future Directions
Compared to other nanoparticle systems like micelles or polymeric nanoparticles, emulsomes
offer the unique advantage of a solid lipid matrix with tunable surface characteristics. Their ease of
scale-up and reproducibility further support their industrial feasibility. However, the current study
is limited to physicochemical characterization and in vitro release. Future investigations should focus
on:
• In vivo pharmacokinetic and biodistribution studies
• Toxicological evaluation in animal models
• Comparative studies with conventional CTX formulations
• Ligand-targeted formulations for site-specific delivery
• Evaluation of therapeutic efficacy in cancer-bearing models
The developed sterically stabilized emulsomes present a safe, efficient, and versatile nanocarrier
platform for Cabazitaxel delivery. Their physicochemical stability, controlled release, and
biocompatible composition mark a significant advancement over traditional formulations. These
findings pave the way for further preclinical and clinical exploration, with the potential to transform
the delivery of Cabazitaxel and other hydrophobic anticancer agents.

Conclusions

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Formulating an effective delivery system for Cabazitaxel (Ctx) remains a significant challenge
due to its limited water solubility and poor bioavailability. In response, this study investigated the
use of Compritol-based emulsomes as a safer, biocompatible alternative to traditional Cremophor-
containing formulations. These emulsomes successfully encapsulated the drug within their solid
lipid core, eliminating the need for harmful solubilizers, surfactants, or co-solvents.
A major advantage of this approach lies in the natural stabilization provided by the
phospholipid layer surrounding the lipid core, which allowed the development of stable nanoscale
formulations without external surfactants. The emulsomes were prepared using a modified solvent
evaporation technique, resulting in a reproducible system capable of efficiently encapsulating
lipophilic compounds.
The optimized Cabazitaxel-loaded emulsomes demonstrated favourable physicochemical
attributes such as appropriate particle size, uniform morphology, stable surface charge, and high
entrapment efficiency. Moreover, they exhibited a pH-responsive release profile—remaining stable
under physiological conditions with limited drug leakage, while enabling enhanced release in acidic
environments mimicking tumour sites. This targeted release behaviour highlights their potential for
minimizing systemic toxicity while maximizing therapeutic efficacy at the disease site.
Surface modification with PEG further enhanced the stealth characteristics of the emulsomes,
reducing recognition and clearance by the reticuloendothelial system (RES) and extending circulation
time. Additionally, the phospholipid shell offers a versatile platform for ligand attachment, paving
the way for targeted delivery to specific receptors, thereby broadening their utility for treating a wide
range of diseases beyond cancer.
In conclusion, sterically stabilized emulsomes present a promising, non-toxic nanocarrier
platform for the delivery of Cabazitaxel and potentially other poorly soluble therapeutic agents. This
preliminary work underscores their potential as a next-generation drug delivery system for improved
treatment outcomes.

Conflict of Interests: ll authors have contributed equally. The authors have no conflict of interest.

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