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Research Article
Biocompatible Phospholipid-Based Mixed Micelles for Tamoxifen Delivery:
Promising Evidences from In - Vitro Anticancer Activity and Dermatokinetic
Studies
Pramod Kumar,
1
Rajendra Kumar,
2
Bhupinder Singh,
2,3
Ruchi Malik,
1
Gajanand Sharma,
3
Deepak Chitkara,
4
O. P. Katare,
3
and Kaisar Raza
1,5
Received 12 September 2016; accepted 22 November 2016
Abstract. Tamoxifen (TAM) is frequently prescribed for the management breast cancer,
but is associated with the challenges like compromised aqueous solubility and poor
bioavailability to the target site. It was envisioned to develop phospholipid-based mixed
micelles to explore the promises offered by the biocompatible carriers. Various compositions
were prepared, employing soya lecithin, polysorbate 80, sodium chloride/dextrose, and water,
by self-assembled technique. The formulations were characterized for micromeritics and
evaluated for in vitro drug release, hemolysis study, dermatokinetic studies on rodents, and
cytotoxicity on MCF-7 cell lines. Cellular uptake of the system was also studied using
confocal laser scanning microscopy. The selected composition was of sub-micron range
(28.81 ± 2.1 nm), with spherical morphology. During in-vitro studies, the mixed micelles
offered controlled drug release than that of conventional gel. Cytotoxicity was significantly
enhanced and IC
50
value was reduced that of the naïve drug. The bioavailability in epidermis
and dermis skin layers was enhanced approx. fivefold and threefold, respectively. The
developed nanosystem not only enhanced the efficacy of the drug but also maintained the
integrity of skin, as revealed by histological studies. The developed TAM-nanocarrier
possesses potential promises for safe and better delivery of TAM.
KEY WORDS: bioavailability; dermal kinetics; MCF-7 cell lines; MTT assay; percutaneous delivery; skin
compatibility; topical delivery.
INTRODUCTION
Various new drug molecules with potential biological
efficacy are being explored, but most of them pose delivery
challenges like compromised solubility, lower bioavailability,
and poor pharmacokinetic profile (1,2). Tamoxifen (TAM) is
one such drugs, which finds immense applications in the
management of various cancers, esp. breast cancer. Generally,
TAM is by oral and parenteral route for the management of
cancer. It is listed as a BCS class II drug and many-a-times its
citrate salt is used in the pharmaceutical products owing to
the poor solubility issues. To circumvent the delivery
challenges associated with TAM, variety of drug delivery
carriers including dendrimers nanosponges (3), graphene (4),
lipid nano capsules (5), nanostructured lipid carriers (6),
polymeric nanoparticles (7), hollow manganese ferrite nano-
carriers (8), chitosan/lecithin nanoparticles (9), self-
nanoemulsifying drug delivery systems (10), flexible mem-
brane vesicles, liposomes (11), lipoplex (12), vesicles (13),
lipid vesicles (14), lecithin organogels (15), nanoemulsions
(16), gold nanoparticles (17), and cyclodextrin nanoparticles
(18) have been developed. In our laboratory, recently, a few
more attempts have been made to deliver TAM employing
pluronic lecithin organogels (19), flexible membrane vesicles,
and polymeric micelles (20,21). Though all these attempts are
novel, but they either require tedious synthetic approaches or
employ non-economic and bio-incompatible excipients.
A close scrutiny by this research group revealed that
phospholipid-based mixed micelles have not yet been ex-
plored for topical delivery of TAM. The main ingredient,
phospholipid, is a well-established biomaterial with immense
biocompatibility and delivery promises. On the other hand,
Tween 80 is non-ionic, biocompatible, and non-immunogenic
1
Department of Pharmacy, School of Chemical Sciences and
Pharmacy, Central University of Rajasthan, Bandar Sindri, Distt.,
Ajmer, Rajasthan, India 305 817.
2
UGC-Centre of Excellence in Applications of Nanomaterials,
Nanoparticles & Nanocomposites, Panjab University, Chandigarh,
India 160 014.
3
Division of Pharmaceutics, University Institute of Pharmaceutical
Sciences, Panjab University, Chandigarh, India 160 014.
4
Department of Pharmacy, Birla Institute of Technology and Science,
Pilani Campus, Vidya Vihar, Pilani, Rajasthan, India 333 031.
5
To whom correspondence should be addressed. (e-mail:
drkaisar@curaj.ac.in; razakaisar_pharma@yahoo.co.in)
AAPS PharmSciTech ( #2016)
DOI: 10.1208/s12249-016-0681-1
1530-9932/16/0000-0001/0 #2016 American Association of Pharmaceutical Scientists
surfactant, approved by various federal agencies for i.v., oral,
and topical routes. Henceforth, it was envisioned to develop
phospholipid-based mixed micelles loaded with TAM and
explore the same for the delivery of TAM by topical route
that too employing simple dispersion technique.
MATERIAL AND METHODS
Materials
Phospholipid 90G was provided ex gratis by M/s Phos-
pholipid GmBH, Nattermannalle, Germany. Sodium chloride,
potassium dihydrogen phosphate, and dipotassium hydrogen
phosphate were purchased from M/s Central Drug House (P)
Ltd., New Delhi, India. Dextrose and ethanol were procured
from M/s RFCL Limited, New Delhi, India and M/s Jai
Chemical Pharma Works, Jaipur, resp. HPLC column and
membrane filters were provided by M/s Merck Specialties
Pvt. Ltd., Mumbai, India. HPLC grade water, methanol,
acetonitrile (ACN), and sodium lauryl sulphate were bought
from M/s Spectrochem Pvt. Ltd., Mumbai, India. Dialysis
membrane (average flat width 22.54 mm, average diameter
14.3 mm, capacity 1.61 mL/cm, molecular cut off 12,000 to
14,000 Da, and pore size of 2.4 nm) was procured from the M/
s Himedia Laboratories Private Limited, New Delhi, India.
Tamoxifen (TAM) and dimethyl sulphoxide (DMSO) were
procured from M/s Sigma-Aldrich, New Delhi, India. MCF-7
cancer cell lines were acquired from European Collection of
Cell Cultures (ECACC), a Culture Collection of Public
Health, England. All other chemicals were of analytical grade
and used without additional purification. Distilled water was
employed throughout the study.
Animals
Wistar rats (6–8 weeks old, 200 ± 20 g) and female Laca
mice (4–6 weeks, 15 ± 5 g) were obtained from Central
Animal House, Panjab University, Chandigarh, India. All
the pathogen-free animals were kept at a temperature of 25 ±
2°C and a relative humidity of 70 ± 5% under natural light/
dark conditions for at least 48 h before dosing. The experi-
ments were performed according to the animal ethical guide-
lines. All the animal protocols were duly approved by
Institutional Animal Ethics Committee (IAEC/S/14/79-2014-
15), Panjab University, Chandigarh, India.
Methods
Preparation of TAM-Loaded Phospholipid-Based Mixed
Micelles
Mixed micelles of TAM were prepared by spontaneous
micelles formation, as described by Song et al., 2011 (22).
Briefly, a total of 27 batches were prepared for the final
selection of the formulation (n= 3), as per the compositions
shown in Table I. The 09 formulations were designed in a
strategic manner and coded from F
1
–F
9
. Initial three for-
mulations (F
1
–F
3
) were prepared without NaCl/dextrose, with
NaCl (0.9% w/v) and dextrose (5% w/v), respectively. In next
three formulations, i.e., F
4
–F
6
, the amount of phospholipid
was doubled and the formulations without NaCl/dextrose,
with NaCl (0.9% w/v) and dextrose (5% w/v) were prepared.
In the last set of F
7
–F
9
, the level of Tween 80 was increased
from532gto1064mgandtheformulationswithout
NaCl/dextrose, with NaCl (0.9% w/v) and dextrose (5% w/
v) were prepared. For selection of ethanol amount, prelimi-
nary studies ranging from 1 to 10% w/wof ethanol were
performed and 6% w/wwas selected on the basis of drug and
lipid solubility. In brief, for any coded formulation, TAM,
Tween 80, and PL were dissolved in dehydrated ethanol with
continuous stirring. Subsequent to that, this homogeneous
phase was added in a streamlined manner in 60 s to 5% (w/v)
dextrose solution or 0.9% (w/v) NaCl solution or plain water
to give clear micellar dispersion. The resulting mixed micellar
dispersions were filtered from the membrane filter (0.22 μm)
and stored in refrigerator till use.
Particle Size, Zeta Potential, and PDI
Particle size, PDI, and zeta potential were determined by
means of Malvern Zetasizer (S. No. MAL1040152, Software
version v6.01, M/s Malvern Instruments Limited, Worcester-
shire, UK) installed at the Department of Pharmacy, Birla
Institute of Technology, Pilani, Rajasthan, India. The average
value of three measurements for each sample was reported as
the final result.
Transmission Electron Microscopy
Transmission electron microscopy (TEM) analysis was
performed using Hitachi H-7500 (M/s Hitachi High-
Technologies Europe GmbH, Krefeld, Germany) installed at
the Central Instrumentation Laboratory, Panjab University,
Chandigarh, India. For analysis, 20 μL of the sample were
deposited on carbon/membrane-coated copper grids and
stained with 1% phosphotungstic acid. Finally, the grid was
air dried and observed under electron microscope and micro
photographed.
Entrapment Efficiency
Entrapment efficiency (EE) of the drug in the mixed
micelles was determined by dialysis bag method. Mixed
micelles equivalent to 1 mg of TAM were packed in dialysis
bag and dipped in 30 mL of ethanol in a beaker for 2 h with
continuous stirring. After complete diffusion of un-entrapped
drug, the contents of the dialysis bag were analyzed for
entrapped drug.
In Vitro Drug Release Studies
Phospholipid-based mixed micelles equivalent to 1 mg of
TAM were placed in a pouch made up of dialysis membrane
and the sealed pouch was dispersed in 30.0 mL diffusion
medium, comprised of a solution of ethanol and phosphate
buffer saline, pH 5.6 in 1:9 v/vratio. The system was
maintained at 37 ± 1°C with gentle magnetic stirring at
50 rpm. Samples of 0.5 mL were withdrawn at the time
intervals of 5, 10, 15, 30, 45, 60, 90, 120, 180, 240, 360, 720, and
1440 min, and the receptor compartment was replaced by
equal volume of fresh diffusion medium, every time. The
amount of TAM released was determined by double-beam
Kumar et al.
UV-visible spectrophotometer at the λ
max
of 301 nm. Blank
micelles (without drug) were subjected to the same protocol
and the respective samples from these blank formulations
served as the blank for the UV-visible spectrophotometry to
nullify the interference, resulting from the possible leaching
of PL and Tween 80 (Cary 100 UV–vis, M/s Agilent
Technologies, Manesar, Haryana, India).
Ex Vivo Hemolysis Studies. Healthy Wistar rats were
employed for collection of the blood samples. Sampling of
1 mL of blood was done from retro-orbital plexus of the
animal and immediately collected in 124 mM sodium citrate
(1:9 v/vratio of sodium citrate solution and blood). Eryth-
rocytes were immediately harvested by centrifuging the blood
and washed thrice with normal saline. The collected RBCs
were re-suspended in normal saline. To this, RBC dispersion
of the test sample (TAM and TAM-loaded phospholipid-
based mixed micelles) was added. The resulting two test tubes
with the test samples and the third one with RBCs dispersed
in double-distilled water were incubated for 1 h at 37°C with
the aid of gentle shaking. Centrifugation at 2000×gfor 5 min.
was performed, after incubation and the supernatant was
analyzed spectrophotometrically at the λ
max
of 415 nm.
Hemolysis induced by double-distilled water was treated as
reference, i.e., 100% hemolysis (22).
MTT Assay. Human breast cancer cells, i.e., MCF-7 cells,
were grown in 96-well tissue-culture plates. Two different
concentrations (equivalent to 1 and 10 μg/mL of TAM) of test
samples (TAM and TAM micelles) were added to cells. Plates
were incubated at 37°C for 48 h using CO
2
incubator. To the
incubated plates, MTT solution of 20 μL (2.5 mg/mL) was
added and the set-up was gently stirred. Plates were again
incubated and centrifuged for 15 min at 400×g. The resulting
MTT-formazan crystals were dissolved in 150 μL of DMSO,
which were previously collected by discarding the superna-
tant. Plates were again stirred and measured for optical
density at 570 nm, and 620 nm as reference wavelength with
the help of microplate reader. Treated cells were washed and
pictured under inverted microscope for morphological
changes. The microphotographs were taken under resolution
of ×30 (n= 3).
Confocal Laser Scanning Microscopy. MCF-7 cells were
grown in culture medium and transferred into 96 plates at the
density of 15,000 cells/cm
2
. Coumarin 6 dye was loaded into
the mixed micelles in the concentration of 50 μg/mL.
Tamoxifen-loaded mixed micelles were transferred into the
96-well plates containing cultured cells and incubated at 37°C
for 24 h. Cells were washed thrice employing PBS 7.4. Ice
cold methanol was added to fix the cells and again it was
washed with PBS 7.4. To stain cell nuclei, 300 nM DAPI
solution was added into cell culture containing mixed micelles
for 05 min. Cells were again washed employing PBS 7.4 to
remove excess amount of DAPI. The cells were scanned and
observed under confocal laser scanning microscope (Nikon
C2 Plus, with NIS Elements Version 4.3 Software, M/s
NIKON Instruments INC., Melville, NY, USA), installed at
UGC-Centre of Excellence in Applications of Nanomaterials,
Nanoparticles & Nanocomposites, Panjab University, Chan-
digarh, India. Coumarin-6 dye was excited at 488 nm and
emitted at 500–550 nm. DAPI was excited and emitted at
405 nm and 417–477 nm, respectively. The photographs of
fluorescent nuclei and the cells were clicked at ×60 magnifi-
cation. All the experiments were performed in the triplicate
(n= 3).
Ex Vivo Dermatokinetic Studies. Wistar rat skin was used
for the studies on Franz diffusion cells (M/s Permegear, Inc.,
PA, USA), as per the previous reported method (23). After
sacrificing the animals, the hair on the dorsal side of animals
were removed. The skin was harvested, freed of adhering fat
layers, and mounted on Franz diffusion cells having a cross-
sectional area of 3.142 cm
2
and receptor volume of 30.0 mL.
Methylene blue dye test was performed to test the integrity of
the skin (24). The diffusion medium in the receptor compart-
ment was composed of ethanol–phosphate-buffered saline
mixture, pH 5.6 in 1:9 v/vratio. The assembly was maintained
at 37 ± 1°C with the help of thermo-regulated outer water
jacket, while the diffusion medium was stirred continuously at
50 rpm using a magnetic stirrer. Diffusion cells were covered
with aluminum foil to avoid contact of the donor as well as
receptor components with the light. Formulation (conven-
tional gel and TAM-loaded mixed micelles; equivalent to
1 mg of TAM) were applied on the skin with the help of
micro-spatula and the drug amount was determined by mass
difference. However, the mixed micellar dispersion was
spread on rat merely by means of a pre-calibrated dropper.
The formulations stayed fix throughout the study, as the
donor was sealed with parafilm, and the study was continued.
Permeation study was performed for duration of 6 h. The
whole skin was removed from the Franz cell at the respective
Table I. Composition of Various Mixed Micelle Formulations
S. No. Formula code TAM (mg) PL (mg) T 80 (mg) EtOH (g) NaCl (mg) Dextrose (mg) Water (g)
1. F1 20 60 532 1.2 –– 18.2
2. F2 20 60 532 1.2 180 –18.0
3. F3 20 60 532 1.2 –1000 17.2
4. F4 20 120 532 1.2 –– 18.1
5. F5 20 120 532 1.2 180 –17.9
6. F6 20 120 532 1.2 –1000 17.1
7. F7 20 120 1064 1.2 –– 17.6
8. F8 20 120 1064 1.2 180 –17.4
9. F9 20 120 1064 1.2 –1000 16.6
Biocompatible Phospholipid-Based Mixed Micelles for Tamoxifen
sampling time and washed thrice to remove any adhering
formulation. The clean skin sample was soaked in hot water
(60°C) for 30 s to facilitate the detachment of epidermis from
dermis. The resulting wedge of the rat skin, exposing the
cleavage of epidermis and dermis, was pulled out with the
help of a forceps. Both of these separated sections were cut
into pieces of small size in separate containers, and kept in
methanol (5 mL) for 1 day for complete drug extraction, in
refrigerator. After filtering the solution through a membrane
(0.45 μm), the filtrate was analyzed using the validated RP-
HPLC technique (LC-2010C HT, M/s Schimadzu Co., Ltd.,
Chiyoda-ku, Tokyo, Japan) with the conditions as follows:
Merck HPLC Column: Oyster BDS C18 (250 × 4.6 mm,
5μm); mobile phase acetonitrile: 50 mM potassium phos-
phate pH 3.0 (45:55%, v/v); column temperature: 30°C;
detection wavelength 254 nm; flow rate: 1.0 mL/min; injection
volume: 5 μL; run time: 30 min.; and detector used: PDA
(SPD-M20A) (15). The various dermatokinetic parameters
were calculated on MS-office Excel using of one compartment
open body model (21).
Skin Compliance Studies. Pathogen-free female Laca
mice were employed for the skin compliance studies and
were allocated into three groups of four animals each. Hair
was removed by use of depilatory cream (Veet, M/s Reckitt
Benckiser, Gurgaon, India). Hair-removed skin was wiped
three to four times with saline-presoaked cotton to remove
the adhere materials. Each group from 1 to 3 was treated with
conventional hydrogel, mixed micelles formulation, and saline
solution (treated as control), respectively (equivalent to 2 mg
of TAM/dose), once a day. Each animal was kept in separate
cage under dark/light standard laboratory conditions (tem-
perature 25 ± 2°C and relative humidity 70 ± 5%). The
respective application was continued for 2 weeks. The
animals were sacrificed by cervical dislocation, and skin was
harvested in 10% formalin solution. Skin histopathology was
performed, after staining the skin with hematoxylin and eosin
(24).
RESULTS AND DISCUSSION
Particle Size, PDI, and Zeta Potential
Figure 1shows the micromeritic as well as surface charge
profiles of the developed nanocarriers. The size range of the
micelles varied from 10.08 to 222.60 nm, whereas the PDI
values were consistently below 0.280. Lower values of PDI
assured homogeneity in the particle size of the dispersed
system. However, slightly negative values of zeta potentials
may be ascribed to the presence of alcohol, whereas
phospholipid and Tween 80 tend to bring the zeta potential
values close to neutral. As a general observation, the particle
size was slightly enhanced when phospholipid levels were
increased, whereas increased Tween 80 levels resulted in
smaller-sized micelles systems. On the other hand, the
dextrose seems to increase the size of the micelles; however,
in conjugation with Tween 80, the size enhancement was
negligible. To proceed further, the formulation-coded F
1
was
selected, as it offered maximum sustenance (>90 days).
Transmission Electron Microscopy
The transmission electron microscopy (TEM) micro-
photograph of selected mixed micelles is portrayed in
Fig. 2. The microphotograph has been captured at
×400,000 magnification. The photograph depicts the pres-
ence of turgid micellar vesicles with near spherical
geometry. The micellar dispersion was composed of non-
aggregated spherical nanoconstructs without significant
aggromelation.
Entrapment Efficiency
EE of the mixed micelles was found to be 86.56 ± 2.60%.
Content of TAM in 1 mL was 0.86 mg. This higher drug
entrapment assured adequate loading of the substantial
amount of the chemotherapeutic agent for sustained release.
In Vitro Drug Release
The drug release profile from the studied system has
been shown in Fig. 3. The amount of naïve drug diffused
3919-1
F1
F2
F3
F4
F5
F6
F7
F8
F9
Values for Particle size, PDI and zeta potential
Formulation Type
Zeta potential PDI Particle Size
Fig. 1. Average size, PDI, and zeta potential values of various
developed systems (n=3)
Fig. 2. TEM microphotograph of mixed micelles at ×400,000
Kumar et al.
across the semipermeable membrane was about 75% to
that of the amount of drug diffused from the mixed
micelles in the studied period of 24 h. The average drug
diffusion flux for the naïve drug was 7.35 μgcm
−2
h
−1
,
whereas for the micelle-loaded TAM, the obtained value
was 9.94 μgcm
−2
h
−1
. The enhanced diffusion of a poorly
soluble drug can be ascribed to the solubility enhanced by
the strategically formulated micellar composition.
Biological Evaluation of Phospholipid-Based Mixed Micelles
Ex Vivo Hemolysis Studies
Percentage hemolysis (2.1%) was decreased to its half
value (1.12%) to that of TAM-encapsulated phospholipid-
based micelles. On the other hand, the blank micelles
were well-tolerated by the erythrocytes (0.11% hemolysis),
owing to biocompatible and immunoneutral components.
The obtained results assured that encasement in mixed
micelles can significantly enhance the blood-compatibility
of the anticancer agents, indicating a potential surfactant-
free anticancer product. Though the developed system is
for topical application, but hemolysis study is important as
there are reports that drug-loaded nanocarriers can reach
the systemic circulation. Henceforth, the findings ensured
substantial hemo-compatibility, if the drug-loaded nano-
carriers reach the systemic circulation.
MTT Assay
The results obtained from MTT assay in MCF-7
breast cancer cell lines are portrayed in Fig. 4a, b.
Cytotoxicity of blank phospholipid-based mixed micelles
was found negligible at the studied concentration of 1 and
10 μg/mL. However, pure TAM offered cytotoxicity in the
range of 16 to 35% at these concentrations. On the other
hand, incorporation in mixed micelles substantially en-
hanced the cytotoxicity by approx. two times. This can be
Fig. 3. In vitro release pattern of TAM from naïve drug and mixed micelles (n=3)
0
30
60
TAM Blank Micelles TAM-Mixed Micelles
% Growth Inhibition
Human Breast Cancer Cell Lines
MCF-7 (1 µg/mL) MCF-7 (10 µg/mL)
bcdea
a
b
Fig. 4. a Graphical presentation of the cancer cell cytotoxicity on MCF-7 cells offered by
various treatments. bMicrophotographs of MCF-7 cell lines: (a) control; (b) TAM, 1 μg/
mL; (c) TAM, 10 μg/mL; (d) TAM-loaded mixed micelles, 1 μg/mL; (e) TAM-loaded
mixed micelles, 10 μg/mL
Biocompatible Phospholipid-Based Mixed Micelles for Tamoxifen
ascribed to the better adhesion and penetration of drug
encased in phospholipid-based carriers. The resemblance
in the biological membranes of the cells and the mixed
micelles, i.e., phospholipid bilayer might have significantly
contributed to the better penetration of the drug-loaded
carriers. On the other hand, enhanced solubilization of
this highly permeable, but solubility compromised drug
can also be one of the major contributing factors.
Confocal Laser Scanning Microscopy
It was observed in Fig. 5that coumarin-6 tagged TAM-
loaded mixed micelles got an easy access to the cytoplasm as
well as cell nuclei of the cancer cells. The blue stain
represents the DAPI-stained nuclei, whereas the green color
represented the invasion by coumarin-6 tagged drug-loaded
nanocarriers. Interestingly, it was also seen that plain dye was
unable to penetrate the cells. Significant portion of nucleo-
plasm was also observed to be penetrated by the nano-
carriers, indicating enhanced cellular uptake. Data derived
from the confocal laser scanning microscopy correlates the
enhanced cytotoxic efficacy of the mixed micelles, and
justifies the result of MTT assay as well as dermatokinetic
studies.
Ex Vivo Dermatokinetic Studies
Dermatokinetic studies confirmed substantial deposition
of TAM in epidermis and dermis of Wistar rat skin, as shown
in Fig. 6a, b. It was observed that the mixed micelles
significantly enhanced the epidermal and dermal bioavailabil-
ity of TAM in substantial amounts vis-à-vis the conventional
gel, as shown in Table II. The bioavailability enhancement in
epidermis was about five times, whereas in dermis, it was
around three times to that of the conventional gel. Mixed
micelles offered enhanced permeation to epidermis (approx.
three times) and dermis (about two times) in comparison to
the conventional gel. The results are in consonance with the
drug diffusion studies employing semipermeable membrane.
The elimination (K
e
) rate constant for both the layers were
found to be significantly lower for mixed micellar system
(p< 0.01). The drug clearance from epidermis was retarded
by approx. eight times and from dermis, this retardation was
of the order of 21 times to that of the conventional hydrogel.
As per the applied dose per 3.14 cm
2
,themaximum
concentration achieved could have been 318.47 μg/cm
2
; this
amount was leached to various layer in various time points,
depending on the characteristics of carriers, drug, skin, and
sink on the contrary; no peak of 4-hydroxy tamoxifen or n-
Fig. 5. Confocal laser scanning microphotographs of (×60): aControl cells with stained
nuclei. bCoumarin-6 tagged TAM-loaded mixed micelles
ab
0
80
160
036
Concentration (µg/cm2)
Time (h)
TAM-Gel TAM-Mixed micelles
0
100
200
036
Concentration ( µg/cm2)
Time (h)
TAM-gel TAM-mixed micelles
Fig. 6. a Graphical representation of the amount of drug present in the epidermis of Wistar rats at various time intervals. b
Graphical representation of the amount of drug present in the dermis of Wistar rats at various time intervals
Kumar et al.
desmethyl tamoxifen was observed in whole study, indicating
no degradation of drug. This indicates that the developed
system possesses better skin delivery potential as well as
forms a drug depot in skin layers. This depot formation will
help to sustain the drug release to the target site for longer
periods. This desired pharmacokinetic change in skin offers
huge potential in cancer chemotherapy, especially in breast
cancers.
Skin Compliance Studies
Histopathological microphotographs of animal skin
treated with normal saline, conventional formulation, and
mixed micelles have been depicted in Fig. 7. The studies
confirmed the biocompatibility of the developed system on
the skin, as no marked changes in skin histopathology
were observed, after the treatment. However, signs of skin
inflammation and acute damage were observed in the skin
sections of the animals receiving once-a-day application of
convention gel of TAM.
CONCLUSIONS
The developed TAM-nanocarrier possesses potential prom-
ises for safe and better delivery of TAM to the cancer cells with
substantial blood and skin compatibility. The enhancement in the
skin bioavailability and sustained release of drug from skin layers
by means of biocompatible nanocarriers are the unique desired
outcomes. Such biocompatible, simple, and scalable nanocarriers
provide a hope to enhance the outcomes from chemotherapy in
early stages of cancer. The same can be further explored to in-
vivo studies and such simple nanocarrier systems can emerge as
better alternative carriers for the delivery of various anticancer
agents.
Table II. Ex Vivo Dermatokinetic Modeling of Conventional Hydrogel and Mixed Micelles in Dermis and Epidermis
Dermatokinetic parameters Conventional hydrogel Mixed micelles
Epidermis Dermis Epidermis Dermis
AUC
0-∞
(μgcm
−2
h) 163.30 ± 9.8 127.70 ± 8.91 895.85 ± 71.67 434.96 ± 30.44
C
maxSkin
(μgcm
−2
) 37.92 ± 2.41 26.97 ± 2.07 261.14 ± 17.76 123.33 ± 8.01
T
maxSkin
(h) 0.91 ± 0.06 1.20 ± 0.08 0.47 ± 0.03 0.68 ± 0.05
K
p
(h
−1
) 2.70 ± 0.20 1.58 ± 0.09 8.00 ± 0.054 2.72 ± 0.22
K
e
(h
−1
) 0.30 ± 0.01 0.37 ± 0.02 0.21 ± 0.01 0.68 ± 0.04
Fig. 7. Photomicrograph of skin sections of mice treated with: asaline (control), b
conventional hydrogel, and cmixed micelles
Biocompatible Phospholipid-Based Mixed Micelles for Tamoxifen
ACKNOWLEDGEMENTS
The financial support from the University Grant Com-
mission (UGC), New Delhi in the form of Start-Up-Grant
(No. F.30-18/2014(BSR)) to Dr. Kaisar Raza is duly acknowl-
edged. The support in the form of UGC-National Fellowship
to the Mr. Pramod Kumar (F./2014-15/NFO-2014-15-OBC-
RAJ-8108/(SA-III/Website)) is also acknowledged.
COMPLIANCE WITH ETHICAL STANDARDS
Conflict of Interest The authors declare that they have no conflict
of interest.
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