Piroxicam Loaded Solid Lipid Nanoparticles (SLNs): Potential for Topical Delivery

Abstract

Aim: The objective of this study was to develop suitable lipid nanocarriers for topical delivery of Piroxicam, known for its anti-inflammatory and anti-arthritic properties and to increase its therapeutic potential and residence time at the site of inflammation and in systemic circulation. Materials and Methods: Piroxicam loaded Solid lipid nanoparticles (SLNs) were prepared by high-speed homogenization followed by ultrasonication technique. Physicochemical evaluation of SLNs involved particle size and zeta potential analysis, electron micrographic analysis, drug loading, entrapment and release studies. Drug excipient interactions were studied by Fourier transform infrared spectroscopy (FTIR) and Differential scanning calorimetry (DSC). Prepared nanoparticles were assimilated into a topical gel system and evaluated for viscosity, in vitro permeation properties and in vivo efficacy assessment by carrageenan induced rat paw edema study. Results: Evaluation of prepared SLNs revealed the particle size, drug loading, entrapment efficiency and in vitro release to significantly differ (p<0.05) with different lipid and surfactant concentrations present in formulations. SEM and TEM were useful aids in studying the morphology of the SLNs. FTIR and DSC revealed no likely interaction amongst the drug and/or any excipients. In vitro permeation and in vivo anti-inflammatory study of the SLN gel revealed it to be a sustained permeation system, maintaining drug concentration over an extended period of time and providing a better therapeutic potential on topical dermal application when compared with a free drug gel. Conclusion: Our results proved the suitability and capability of fabricated piroxicam loaded SLN gel system in treating arthritic pain and inflammation when delivered topically.

Full text

S82 Indian Journal of Pharmaceutical Education and Research | Vol 53 | Issue 2 (Suppl)| Apr-Jun, 2019
Original Arcle
www.ijper.org
DOI: 10.5530/ijper.53.2s.52
Correspondence:
Ms. Harshita Krishnatreyya
Department of Pharmaceuti-
cal Sciences Dibrugarh
University Dibrugarh, Assam-
786004, INDIA.
Phone: +91 8638095195
E-mail: harshita.
krishnatreyya@gmail.com
Submission Date: 13-09-2018;
Revision Date: 28-12-2018;
Accepted Date: 20-03-2019
Piroxicam Loaded Solid Lipid Nanoparticles (SLNs):
Potential for Topical Delivery
Harshita Krishnatreyya1,*, Sanjay Dey2, Paulami Pal1, Pranab Jyoti Das1, Vipin Kumar Sharma3,
Bhaskar Mazumder1
1Department of Pharmaceutical Sciences, Dibrugarh University, Dibrugarh, Assam, INDIA.
2Department of Pharmaceutics, School of Pharmacy, Techno India University, West Bengal, INDIA.
3Department of Pharmaceutical Sciences, Gurukul Kangri University, Haridwar, Uttarakhand, INDIA.
ABSTRACT
Aim: The objective of this study was to develop suitable lipid nanocarriers for topical
delivery of Piroxicam, known for its anti-inammatory and anti-arthritic properties and to
increase its therapeutic potential and residence time at the site of inammation and in
systemic circulation. Materials and Methods: Piroxicam loaded Solid lipid nanoparticles
(SLNs) were prepared by high-speed homogenization followed by ultrasonication
technique. Physicochemical evaluation of SLNs involved particle size and zeta potential
analysis, electron micrographic analysis, drug loading, entrapment and release studies.
Drug excipient interactions were studied by Fourier transform infrared spectroscopy (FTIR)
and Differential scanning calorimetry (DSC). Prepared nanoparticles were assimilated
into a topical gel system and evaluated for viscosity, in vitro permeation properties and
in vivo efcacy assessment by carrageenan induced rat paw edema study. Results:
Evaluation of prepared SLNs revealed the particle size, drug loading, entrapment efciency
and in vitro release to signicantly differ (p<0.05) with different lipid and surfactant
concentrations present in formulations. SEM and TEM were useful aids in studying the
morphology of the SLNs. FTIR and DSC revealed no likely interaction amongst the drug
and/or any excipients. In vitro permeation and in vivo anti-inammatory study of the
SLN gel revealed it to be a sustained permeation system, maintaining drug concentration
over an extended period of time and providing a better therapeutic potential on topical
dermal application when compared with a free drug gel. Conclusion: Our results proved
the suitability and capability of fabricated piroxicam loaded SLN gel system in treating
arthritic pain and inammation when delivered topically.
Key words: Solid lipid nanoparticles, Piroxicam, Topical, Inammation, Dermal.
INTRODUCTION
Piroxicam belongs to the oxicam class
of Non-steroidal anti-inammatory drug
(NSAID) used in the treatment of rheu-
matoid arthritis, osteoarthritis, alkalosing
spandylitis and acute gout.1,2 It is proposed
as a Biopharmaceutical Classication Sys-
tem (BCS) class II drug with low solubility
(pKa 6.3) and high permeability.3 Admin-
istration of piroxicam by traditional meth-
ods (orally or intramuscularly) resulted in
severe side effects and toxicity like nausea,
vomiting, dyspepsia, epigastric pain, gas-
trointestinal ulceration and higher risk of
renal failure or bleeding.4,5 As opposed to
the traditional methods, transdermal and/
or topical drug delivery system possesses
several advantages such as protection from
gastrointestinal enzymatic degradation,
bypass of hepatic rst-pass metabolism and
lower risk of systemic side effects.6 There-
fore, this study was designed so as to obtain
a safe and effective therapeutic outcome by
developing a suitable carrier for the topical
delivery of piroxicam. The main obstacle
of delivery of drug through topical route is
the permeation of drug through skin stra-
tum corneum. There are several strategies
such as the use of vesicular formulation as a

Krishnatreyya, et al.: Piroxicam Nanoparticles for Topical Delivery
Indian Journal of Pharmaceutical Education and Research | Vol 53 | Issue 2 (Suppl)| Apr-Jun, 2019 S83
delivery vehicle, use of penetration enhancers (Chemi-
cal approach) and physical methods (Like: iontopho-
resis, sonophoresis, etc.) have been reported earlier.7-13
Unfortunately, the physical and chemical methods are
expensive and may cause skin irritation.
Nanoparticles composed of lipid materials such as wax
or fats seem attractive carrier system for the transder-
mal delivery of drug.14,15 They have several advantages
over traditional method of transdermal drug delivery.
Nanoparticles made of solid lipid display unique ben-
ets in drug delivery and produce various desirable
effects. They moisten the skin through occlusion by
providing skin hydration.16 Solid lipid nanoparticles
(SLNs) are suitable for use as damaged or inamed skin
as they are made up of physiological and biodegradable
lipids that are non-irritant and non-toxic. Several stud-
ies reported the favourable incorporation of drugs into
SLN and its advantages as a colloidal carrier system.17-19
In the present study, SLN was selected as delivery car-
rier for delivering piroxicam through topical route.
The present work was designed mainly with the aim
of formulating, evaluating and consequently screening
piroxicam incorporated SLNs for the possibility of any
anti-inammatory efcacy when applied by the topical
route of drug delivery.
MATERIALS AND METHODS
Piroxicam was procured as a gift sample from Alkem
Laboratories, Sikkim, India.
Phospholipon 80 was obtained as gift sample from
Lipoid, Ludwigshafen, Germany. Stearic acid and Plu-
ronic F68 were bought from Himedia Chemicals, Mum-
bai, India. All reagents and chemicals used in this study
were of analytical grade and used as received without
additional purication.
Fabrication of Piroxicam loaded SLNs
Piroxicam loaded SLNs were developed by high-speed
homogenization followed by ultrasonication technique.
The composition of piroxicam loaded SLNs is pre-
sented in Table 1. In brief, accurate weighed amounts
of piroxicam and surfactants were dissolved in 10 mL
of methanol. On the other hand, exact amount of
lipid was taken in 10 ml of acetone and stirred for 5
min. Both the mixtures were maintained at the same
temperature (60ºC). After thermal treatment, both of
these were admixed. The mixture was then sonicated
for 15 min using a probe sonicator (Ultrasonic Proces-
sor, Hielscher, Germany). This solution constituted the
organic phase. In another beaker, weighed quantity of
surfactant was dissolved in 25 mL of distilled water and
heated at 60ºC. This solution constituted the aqueous
phase. Both organic and aqueous phases were mixed
together and homogenized by high-speed homogenizer
(Ultra Turrax® T 25, Janke and Kunkle GmbH, Ger-
many) at 15000 rpm for 8 min. The homogenized solu-
tion was sonicated for 20 min to prevent agglomeration
of smaller-size particles. The obtained formulation was
stirred using magnetic stirrer (Remi, Mumbai, India) for
30 min, ltered through 0.2 µm Whatman lter paper
and centrifuged at 20,000 rpm for 20 min (Remi Instru-
ments, India). The residue was collected while discard-
ing the supernatant and collected sediment was further
lyophilized to obtain its free-owing powder form.
Characterization of SLNs
Particle size and zeta potential analysis
Particle-size analysis of piroxicam SLNs was performed
by dynamic light scattering (DLS) method using a Mal-
vern Zetasizer Nano ZS (Malvern Instruments, UK).
DLS yielded mean particle size and polydispersity index
(PDI), which measures the width of size dispersion.
The particle size and PDI of the samples were investi-
gated by calculating average of 10 measurements at an
angle of 90ºC in 10 mm diameter disposable plastic cell
at 25ºC. Before measurement, the samples were diluted
in distilled water to yield a suitable scattering intensity.
Zeta potential determination of piroxicam SLNs was
done by the same Malvern Zetasizer Nano ZS (Malvern
Instruments, UK). The measurements were carried
out in distilled water adapted to 50 mS/cm conductiv-
ity with sodium chloride solution (0.9% w/v) and eld
strength was applied 20 V/cm. During measurement
the pH range was 5.5 – 6.0. Instrumental zeta potential
was determined using the Helmholtz-Smoulchowsky
equation.
Electron microscopic analysis
Both Scanning electron microscopy (SEM) and Trans-
mission electron microscopy (TEM) were done to
directly observe the piroxicam loaded SLNs. These
techniques provide particle specics such as surface
topography and external morphology of the samples.
TEM has a small size limit of detection as compared to
SEM. Samples were sent to the North Eastern Hill Uni-
versity, Shillong, India for TEM while SEM analysis of
the samples were done in Oil India Limited, Duliajan,
Assam, India.
Drug loading and entrapment efciency
Drug loading and entrapment efciency of piroxicam
SLNs were adjudged by the centrifugation technique.20
1 ml of centrifuged SLN sediment (As mentioned in
the SLN preparation method) was taken and diluted
with acetonitrile. The diluted sample was analysed by
UV-Visible spectrophotometer (UV-1700, Shimadzu,
Tokyo, Japan) at λmax of 230 nm. The drug loading and

Krishnatreyya, et al.: Piroxicam Nanoparticles for Topical Delivery
S84 Indian Journal of Pharmaceutical Education and Research | Vol 53 | Issue 2 (Suppl)| Apr-Jun, 2019
entrapment efciency of piroxicam in SLNs were calcu-
lated according to following equations:
determined
In vitro
Drug Release Study
In vitro release study of piroxicam SLNs was studied
using the Franz diffusion apparatus method. Typically, 1
mL of the centrifuged SLN sediment (As mentioned in
the SLN preparation method) was placed on the dialy-
sis membrane having a molecular weight cutoff: 10,000
-12,000 (HiMedia, Mumbai, India). The membrane
was adjusted between the donor and receptor sections
of the diffusion apparatus which was then lled with
200 mL phosphate buffer, pH 5.8, which served as the
release medium. The release medium was stirred at 175
rpm in a magnetic stirrer (Remi, Mumbai, India) and the
temperature was kept at 37 ± 0.5ºC during the experi-
mental procedure. At specic time intervals, 1 mL of
aliquot was withdrawn and reinstated with equal volume
of fresh buffer to preserve the sink condition. Aliquots
were diluted suitably with acetonitrile and the quantity
of piroxicam contained in the samples was evaluated
using a UV-Visible spectrophotometer (UV-1700, Shi-
madzu, Tokyo, Japan) at? max of 230 nm. The release
of individual SLN formulations was done in triplicate.
Drug Release Kinetics
The in vitro drug release pattern from piroxicam SLNs
was predicted by applying the drug release data to dif-
ferent release kinetic models like: zero-order, rst-order,
Higuchi, Korsmeyer-Peppas and Hixson-Crowell. For
each release kinetics model, the values of R2 (Correla-
tion coefcient) and n (Release exponent of Korsmeyer-
Peppas kinetics), K (Rate constant) and SSR (Sum of
squared residual) were determined for each formulation
and all the models and the possible methods of drug
release were identied.
Optimization of Suitable SLN Formulation
Suitable formulation from amongst the prepared piroxi-
cam SLN formulations was selected, based on their
optimum particle size and zeta potential, high drug
loading, entrapment efciency and better in vitro drug
release properties. The optimized formulation was fur-
ther scrutinized and used for additional studies.
Incorporation of Piroxicam-loaded SLN into Gel
System
Gel containing appropriate formulation of piroxicam
SLN and pure piroxicam were made by using 0.5% w/v
of carbopol 934P as gel forming polymer. Firstly, gel
was prepared by diffusing carbopol 934P in distilled
water containing glycerol (10%) and kept for saturation
for 3 h. Lyophilized piroxicam-loaded SLN was admixed
with aqueous carbopl 934P dispersion under controlled
stirring using a magnetic stirrer (Remi, Mumbai, India)
to obtain uniform and smooth dispersion containing
a nal concentration of 20% w/w piroxicam-loaded
SLN. The pH of the piroxicam-loaded SLN enriched
gel was adjusted to 6.0 using sufcient quantity of tri-
ethanolamine. Piroxicam conventional gel was prepared
in similar process and was used as reference during ef-
cacy evaluation.
Evaluation of Piroxicam SLN gel system
Appearance
Organoleptic properties like color, odor, phase sepa-
ration and gel capacity to be washed out were visually
observed in the SLN gel formulation.
Physicochemical properties
pH of piroxicam SLN gel was evaluated using a digital
pH meter (Model-355, Systronic, India). The viscosity
and rheological properties of the water based piroxicam
SLN gels as compared to prepared conventional gel was
determined using a Brookeeld viscometer (Brookeld
Engineering Laboratories, USA) with spindle no. 6 at 10
rpm. All measurements were performed at a tempera-
ture of 37 ± 0.5ºC.
Skin Irritation
A small quantity of the prepared SLN gel was applied
on the skin and evaluated for its easy removal after 1
min. This was done to observe the presence of rough or
granular surface particles and examine the skin for any
sign/symptoms of irritation.
Drug-excipient interaction study
Fourier transform infrared spectra (FTIR) spectroscopy
FTIR spectra was evaluated to check the compatibility
of the drug with formulation excipients and to ensure
proper entrapment of drug in formulation. The FTIR
spectra was recorded using an FTIR spectrophotometer
(Shimadzu, Tokyo, Japan; Model: 8400) in between the
range of 600-4000 cm-1 using KBr discs to recognize
the variations in the characteristic peaks of individual
drug piroxicam and piroxicam loaded SLN formula-
tions. Pure drug was pulverized with dried potassium
bromide. This mixture was then pressed in a hydrau-
lic press to form a transparent pellet and the pellet
FTIR spectrum was measured. To observe the spectra
of the piroxicam SLN gel, a drop of sample (Around
5–50µL) was spread on the plate and the IR spectrum
was recorded. Distinctive functional groups present in

Krishnatreyya, et al.: Piroxicam Nanoparticles for Topical Delivery
Indian Journal of Pharmaceutical Education and Research | Vol 53 | Issue 2 (Suppl)| Apr-Jun, 2019 S85
each sample was identied and compared to check for
specic drug excipient interactions.
Differential Scanning Calorimetry (DSC)
Thermal behaviour of pure piroxicam drug and piroxi-
cam-loaded SLNs was analysed using a differential scan-
ning calorimeter (Perkin Elmer, USA). Approximately
10 mg of sample was placed in aluminium crimp cells
and heated at the scanning rate of 10ºC/min from 30 to
400º C in a nitrogen atmosphere. Aluminium oxide was
used as the reference material to calibrate the tempera-
ture and energy scale of the DSC instrument.
In vitro
Permeation Study
The in vitro permeation study of piroxicam SLN gels
was done using pig ear epidermal skin obtained from
the local slaughter house. The subcutaneous fat on the
skin was removed and soaked in 2M sodium bromide
solution for 36 h. The pig ear epidermis was peeled
from the dermis, washed with double distilled water
and used for further study. The isolated epidermis was
mounted carefully across the donor and receptor sec-
tions in Franz Diffusion cell with the epidermal side up
and an effective diffusion area of 3.8 cm.2 The receptor
section remained lled with 50 mL of phosphate buffer
(pH 5.8) and was maintained at 37 ± 0.5ºC with stirring
at 600 rpm. The skin membranes were calibrated for 30
min before placing gel into donor compartment. Then
1 g of piroxicam SLN gel and conventional gel were
placed individually in the donor section of the diffusion
cell. Sampling was done at predetermined intervals. At
specic points, 2.5 ml aliquots were withdrawn from the
receptor section and immediately an equal volume of
receptor uid was replaced in it. Piroxicam concentra-
tion in withdrawn sample was analysed spectrophoto-
metrically using a UV-Visible spectrophotometer at max
260 nm. The permeation of both gels were carried out
in triplicate.
Permeation Data Analysis
The permeation prole of piroxicam from piroxicam
loaded SLN gel and conventional gel was made by plot-
ting the cumulative amount of piroxicam permeated
per unit pig skin epidermis area (µg/cm2) versus time.
The steady state ux (Jss, µg/cm2/h) of piroxicam was
determined from the slope of graph by applying linear
regression analysis. The permeability coefcient (Kp)
of piroxicam was calculated using following equation:
Where, Jss is the steady state ux (µg/cm2/h) of piroxi-
cam from the gel and C is the initial piroxicam concen-
tration available in donor section. The enhancement
ratio (Er) of piroxicam SLN gel was determined by fol-
lowing expression:
In vivo Anti-Inammatory Efcacy of Piroxicam SLN Gel
Ethical statement for the use of experimental animals
Wistar rats (Rattus norvegicus) were used as experi-
mental animals. All animal experiments complied with
the ARRIVE guidelines12 and were carried out in accor-
dance with the U.K. Animals (Scientic Procedures) Act
1986 and associated guidelines. They were used as per
the guidelines of the Institutional Animal Ethics Com-
mittee of the Department of Pharmaceutical Sciences,
Dibrugarh University, Assam, India (Regd No. 1576/
GO/a/11/CPCSEA, Dated: 17/02/2012) under the
approval number: IAEC/DU/95. All applicable inter-
national, national and/or institutional guidelines for
the care and use of animals were followed. The in vivo
anti-inammatory potential of formulated piroxicam
SLN gel was evaluated by the carrageenen induced paw
edema method using Wistar albino rats as the experi-
mental animals. Animal care and handling throughout
the experimental procedure were carried out in accor-
dance with the CPSEA guidelines.
The rats were acclimatized at the commencement of the
study and were divided into four groups (n=6) as fol-
lows: Group 1: Normal saline (control) Group 2: Carra-
geenan and normal saline (toxicant) Group 3: Piroxicam
conventional gel (standard) Group 4: Piroxicam loaded
SLN gel (test) For the paw edema study, a digital ple-
thysmometer was used to measure the volume of the
rat paw (in ml). The plethysmometer was designed so
as to measure small changes in volume. The rats were
marked on the left hind paw at a specic position. Every
now and then, the paw was dipped to that xed mark in
an electrolyte uid so that every time the paw is dipped
in the electrolyte uid, it would be dipped to that xed
mark which would assure constant paw volume. The
original paw volume of the rats were measured and
noted. Paw edema was induced in the rats using freshly
prepared 1% carrageenan saline solution. 0.1ml of pre-
pared carrageenen solution was injected into the plantar
region of the left hind paw of the rats. After the induc-
tion of edema, the enlarged paw volumes were noted.
Prepared free drug gel as well as SLN gel formulation
were applied to the third and fourth groups.
Readings of the paw volume were then taken after 1,
2, 4, 8, 10 and 24 h of time intervals using the plethys-
mometer. The percentage edema rate and percentage
edema inhibition rate were calculated using following
equations:
% Edema rate = Vt V0/ V0 ×100 %

Krishnatreyya, et al.: Piroxicam Nanoparticles for Topical Delivery
S86 Indian Journal of Pharmaceutical Education and Research | Vol 53 | Issue 2 (Suppl)| Apr-Jun, 2019
%Edema inhibition rate= ES E
t/ES ×100. Here, V0
and Vt indicate mean paw volume before and after car-
rageenan injection (mL), respectively, ES and Et indi-
cate the edema rate of standard group and test group,
respectively.
Statistical Analysis
All data were presented as Mean ± Standard deviation
(S.D). Statistical analysis was carried out by one way
analysis of variance (ANOVA) by utilizing the two-tailed
paired ‘t’ test (GraphPad InStat Demo Version). P val-
ues <0.05 were considered to be statistically signicant.
RESULTS AND DISCUSSION
Fabrication of Piroxicam SLNs
Formulation of piroxicam SLNs was done by high-
speed homogenization and ultrasonication technique.
Formulation compositions are presented in Table 1.
In particular, application of homogenization method
resulted in the formation of unstable dispersion with
broad dimensional distribution. Conversely, application
of homogenization followed by sonication resulted in
a stable and homogenous formulation which was free
from aggregation. Stearic acid was used as lipid matrix
for the fabrication of piroxicam SLNs. Pluronic F68
and phospholipon 80 were used as surfactant and co-
surfactant respectively. Varied proportions of the lipid
matrix and pluronic F68 were utilized to constitute the
SLNs. The fabricated piroxicam loaded SLNs were sub-
jected to different characterization.
Characterization of Piroxicam Loaded SLN
Particle size and zeta potential
Particle size and zeta potential data of piroxicam SLNs
are depicted in Table 2. The particle size of piroxicam
SLNs was found in the range of 136.0 ± 15.6 nm to
279.1 ± 21.3 nm. It was clear from Table 2 that the par-
ticle size decreased with increasing concentration of
Pluronic F68 in piroxicam loaded SLN formulations.
The increasing of pluronic F68 content in SLN formu-
lations could decrease the interfacial tension between
lipid matrix and the dispersion medium (Aqueous
phase), consequently favour the formation of SLNs with
smaller particle size.21 The particle size was increased
upon increasing the amount of stearic acid in the SLN
formulations. The polydispersity index (PDI) of piroxi-
cam SLNs was found to be within an acceptable range
of 0.206 ± 0.091 to 0.479 ± 0.045. Lower PDI values
of formulations are considered better as they indicate
the formation of uniform particles. Optimum concen-
trations of Pluronic F68 and stearic acid are relevant as
the PDI of SLNs were found to increase with increas-
ing lipid and surfactant concentrations. Zeta potential
is a potential parameter to access the stability of SLN
dispersion.22 The zeta potential of piroxicam SLNs was
found in the range of -14.5 ± 2.7 mV to -27.5 ± 1.9
mV. Generally, particles are considered stable when the
zeta potential of colloidal dispersed particles above 30
mV and this is due to electrostatic repulsion between
particles.23 The zeta potential values of SLNs revealed
the stability of the formulated piroxicam SLNs. Values
of zeta potential were found to increase with increased
Pluronic F68 content in SLNs whereas increased stearic
acid content reduced the zeta potential of formulated
SLNs.
Electron Microscopy Analysis
TEM photomicrographs of piroxicam SLNs is pre-
sented in Figures 1. The TEM photomicrograph
revealed the ideal spherical shape of piroxicam SLNs
which were distinct and free of aggregates (Figures 1A,
1B). Magnication of single SLN shows smooth surface
and uniform dispersion of piroxicam throughout the
lipid matrix (Figures 1A). Figures 2 shows SEM photo-
micrographs of piroxicam SLNs. Smooth and uniform
surface of a nanoparticle (Crushed) can be seen in Fig-
ures 2A while Figures 2B shows the overall formulation
with the presence of distinct SLN particles.
Drug Loading and Entrapment Efficiency
The drug loading and entrapment efciency data of
piroxicam SLNs are shown in Table 2. The drug load-
ing and entrapment efciency of piroxicam SLN was
found in the range of 0.437 ± 0.08% to 0.649 ± 0.09%
and 50.9 ± 1.77% to 74.0 ± 1.35%, respectively. The
concentration of Pluronic F68 and stearic acid might
have played crucial roles in improving piroxicam incor-
poration in the SLNs. The entrapment of piroxicam
in SLNs was increased when both Pluronic F68 and
stearic acid are increased. Pluronic F68 also contributes
towards increasing the solubility of Piroxicam within
the lipid matrix. The enhanced entrapment efciency of
piroxicam in SLN upon addition of stearic acid could
be due increasing the amount of available lipid matrix
for incorporation of piroxicam.
In vitro
Drug Release and Release Kinetic Study
The in vitro release prole of piroxicam from differ-
ent SLNs are shown in Figures 3. The percentage of
piroxicam release from formulations varied from 44.6 ±
1.286% to 81.70 ± 1.02% within 24 h of release study.
The in vitro release prole revealed a faster release at the
initial stage followed by prolonged release. Such a release
can be advantageous as it provides the initial therapeutic
concentration of the drug which provides faster relief
after dermal application. However, the sustained release

Krishnatreyya, et al.: Piroxicam Nanoparticles for Topical Delivery
Indian Journal of Pharmaceutical Education and Research | Vol 53 | Issue 2 (Suppl)| Apr-Jun, 2019 S87
effect as shown in release pattern may provide the
drug over a prolonged period of time. The initial faster
release can be explained by the enrichment of piroxi-
cam at outer surface of SLN and the prolonged release
can be due to slower diffusion of piroxicam from the
solid matrix of SLN.
Drug Release Kinetics
The obtained SLN release data were tted with the Zero
order, First order, Higuchi, Hixson Crowell and Kors-
meyer Peppas kinetic drug release models (Table 3). The
best t with the highest regression coefcient was found
in case of the Korsmeyer Peppas model while the drug
was also found to show zero order model release. Zero
order release indicates the release from systems which is
not dependent of the amount of substances present. In
case of Korsmeyer Peppas models, the diffusion expo-
nent “n” in all the cases has been found to be as follows:
0.5< n < 1.0 where n is the diffusion exponent; which
indicates drug release from the prepared SLN disper-
sion is found to follow Anomalous (Non Fickian) diffu-
sion mechanism.
Optimization of Piroxicam Loaded SLN Gel System
On the basis of optimized values obtained from particle
size, drug entrapment and release studies, the piroxicam
SLN formulation F2 (Containing 1.00% Stearic acid and
1.50 % Pluronic F-68) was chosen for further study. For
topical delivery, the SLNs were lyophilized and incorpo-
rated into carbopol 934P gel system.
Evaluation of Piroxicam SLN Gel System
Appearance:
Piroxicam loaded SLN incorporated gel
was found to be clear and smooth in texture, translucent
and showed homogenous consistency.
Physicochemical parameters
Optimum pH and viscosity values of the SLN gel
was obtained which was adequate for topical cutane-
ous administration as shown in Table 4. The viscosity
measurement studies of the prepared SLN gel was per-
formed by Brookeld rheometer as shown in Figures 4.
The SLN dispersions generally possessed a lipid matrix
of 10-20% and 90-80% water because of which they
were less visco-elastic by nature. The rheological prop-
erties of topical formulations inuences its spreadability
and contact time on the skin surface. The thixotropic
nature of the gel (Figures 4) was conrmed from the
curve obtained from rheological studies. The obtained
curves afrm that the gel is effectively viscous in nature
and the effect of shear rate on its viscosity can be
observed.
Skin irritation
The optimized SLN gel formulation produced no signs
of skin irritation (Table 4) or toxicity in experimental
animals and was hence, considered to be acceptable for
topical application.
Drug-Excipient Interaction Study
FTIR
The FTIR spectra of pure piroxicam and piroxicam
SLN gel is shown in Figures 4. The FTIR analysis exhib-
ited no distinct physical or chemical interaction of the
drug (Figures 5A) with either the lipids (Figures 5B, 5D)
or surfactant (Figures 5C). The FTIR spectrum of the
pure piroxicam (Figures 5A) showed a small peak at
3333.14 cm-1 corresponding to N-H and O-H stretching
vibrations. Major peaks were observed at 1526cm-1 and
1431cm-1 due to N-O asymmetric stretching. The pres-
ence and position of peaks in FTIR spectrum of piroxi-
cam SLN gel (Figures 5D) showed spectral similarity
to an extent with peak positions of FTIR spectrum of
pure piroxicam. The characteristic peaks of Piroxicam
were observed in the SLN gel indicating little interac-
tion while a few peaks (at near about 1500 cm-1) were
found to be missing which might be an indication of
proper entrapment of the drug within the lipid matrix.
Table 1: Composition of Solid-Lipid Nanoparticles Containing Piroxicam.
Formulation Drug (%w/v) Stearic acid (%w/v) Pluronic F68 (%w/v) Phospholipon 80 (%w/v) Water (%v/v)
F1 0.01 1.00 1.00 0.02 100
F2 0.01 1.00 1.50 0.02 100
F3 0.01 1.00 2.00 0.02 100
F4 0.01 0.75 1.00 0.02 100
F5 0.01 1.25 1.00 0.02 100
F6 0.01 1.25 2.00 0.02 100
F7 0.01 1.50 2.00 0.02 100

Krishnatreyya, et al.: Piroxicam Nanoparticles for Topical Delivery
S88 Indian Journal of Pharmaceutical Education and Research | Vol 53 | Issue 2 (Suppl)| Apr-Jun, 2019
Table 2: Characteristics of Fabricated Piroxicam Loaded SLNs.
Formulation
code
Particle size (nm)* PDI* Zeta potential
(mV)*
Drug loading
(%)*
Entrapment
efciency (%)*
F1 268.4 ± 16.8 0.325 ± 0.102 -25.2 ± 2.5 0.481 + 0.03 60.2 + 2.46
F2 237.0 ± 21.3 0.285 ± 0.015 -20.6 ± 1.0 0.547+ 0.07 64.8 + 1.32
F3 136.0 ± 15.6 0.206 ± 0.091 -19.2 ± 2.3 0.589 + 0.05 69.0 + 2.01
F4 242.4 ± 24.9 0.312 ± 0.023 -14.5 ± 2.7 0.437 + 0.08 50.9 + 1.77
F5 279.1 ± 28.7 0.479 ± 0.045 -27.5 ± 1.9 0.508 + 0.04 63.4 + 1.69
F6 190.0 ± 30.1 0.292 ± 0.082 -24.2 ± 1.6 0.649 + 0.09 74.0 + 1.35
F7 302.1 ± 22.4 0.465 ± 0.276 -22.1 ± 1.2 0.542 ± 0.21 62.3 ± 2.21
Table 3: The release data of the SLNs were fitted into different kinetic models to determine the mechanism of
drug release
Formulation code Kinetic model Equation R2
F1 Zero y = 0.144x+0.976 0.9021
First y =0.0133x+1.292 0.7545
Higuchi y = 0.116x+ 0.3814 0.8138
Hixson Crowell y = 0.234x+2.232 0.1848
Korsmeyer Peppas y =0 .6014x+0.572,n = 0.6014 0.9462
F2 Zero y = 0.6171x+0.572 0.9123
First y =0.0173x+1.533 0.2822
Higuchi y =0.1294x+0.396 0.8469
Hixson Crowell y =0 .989x+0.3324 0.4678
Korsmeyer Peppas y = 0.4830x-0.135, n = 0.4830 0.9594
F3 Zero y = 0.1710x+0.5311 0.7598
First y = 0.0182x+0.1603 0.3808
Higuchi y = 0.1306x+ 0.358 0.7508
Hixson Crowell y =0.6125x+ 0.221 0.5314
Korsmeyer Peppas y = 0.074x-0.1786, n = 0.074 0.8854
F4 Zero y = 0.1775x+0.5002 0.9621
First y = 0.0186x+0.1674 0.4498
Higuchi y = 0.1322x+0.3311 0.8737
Hixson Crowell y = 0.4036x+0.1232 0.4324
Korsmeyer Peppas y = 0.9261x-0.2123, n =0.9261 0.9036
F5 Zero y = 0.085x+0.3160 0.9868
First y = 0.0051x+0.1822 0.2076
Higuchi y = 0.6839x+ 0.2227 0.8504
Hixson Crowell y = 0.2711x+0.4118 0.5466
Korsmeyer Peppas y = 0.4832x-0.1015, n = 0.4832 0.9524
F6 Zero y = 0.7974x+0.1873 0.8127
First y = 0.0072x+0.184 0.4036
Higuchi y = 0.7324x+0.123 0.6188
Hixson Crowell y = 0.7345x+1.253 0.6127
Korsmeyer Peppas y = 0.8761x-0.044, n = 0.8761 0.9907
F7 Zero y = 0.119x+0.3053 0.9303
First y = 0.0213x+0.184 0.3781
Higuchi y = 0.7933x+0.2062 0.8014
Hixson Crowell y = 0.1098x+ 0.1876 0.5632
Korsmeyer Peppas y = 0.5212x- 0.4023, n = 0.5212 0.9234

Krishnatreyya, et al.: Piroxicam Nanoparticles for Topical Delivery
Indian Journal of Pharmaceutical Education and Research | Vol 53 | Issue 2 (Suppl)| Apr-Jun, 2019 S89
Table 4: Physicochemical Properties and Permeability Parameters of Piroxicam SLN Gel and Piroxicam Con-
ventional Gel (Mean ± SD;
n
= 3).
Formulation pH Viscosity(CP) Skin
Irritation
Steady state ux
(µg/cm2/h; JSS)
Permeability
coefcient
(cm/h; KP)
Enhancement
ratio (Er)
Piroxicam conventional
gel
7.28± 0.12 423.9± 0.58 Absent 8.006± 1.102 2.704 ± 0.892 -
Piroxicam loaded SLN
incorporated gel
7.25± 0.43 511.21± 0.90 Absent 8.832± 1.045 2.983 ± 0.782 1.10
Figure 1: TEM Images of Piroxicam Loaded SLNs. (A) Ideal
Spherical Shape of Piroxicam SLN (B) SLNs were Smooth,
Distinct and Free of Aggregates.
Figure 2: SEM Images of Piroxicam Loaded SLNs. (A) Surface
of a smooth, crushed nanoparticle. (B) Presence of distinct
SLN particles within the formulation.
DSC
The DSC thermogram of pure piroxicam and piroxicam
loaded SLN are depicted in Figure 6. The DSC ther-
mogram of pure piroxicam (Figures 6A) showed endo-
thermic peak corresponding to 203.63°C . This peak is
characteristic of melting point of piroxicam. However,
the DSC thermogram of piroxicam SLN (Figures 6B)
showed an undistinct peak at 53.21°C (onset time) and
it might be due to the melting of stearic acid which has
a melting point of around 54-58°C.24 Also, the formula-
tion was in a gel, i.e, liquid form already, which might
have been a reason for its low melting point. In the DSC
thermogram of the SLN formulation, the drug peak
was not clearly observed thereby indicating its entrap-
ment within the lipid matrix of the formulation.
Permeation of Piroxicam Loaded SLN Gel and
Permeation Data Analysis
The permeation prole of piroxicam from SLN gel
and prepared free drug gel are presented in Figures 7.
It depicted the steady increase of piroxicam permeation
across the pig ear epidermis with time. As expected,
piroxicam SLN gel was found to show better perme-
ation as compared to the conventional gel formulation.
Piroxicam SLN gel showed an initial lag phase during
permeation. The release of piroxicam from SLN gel
occurred in two-phases: initially, piroxicam entrapped
within the lipid matrix is unleashed into gel formula-
tions and secondly, piroxicam is released from the gel
to the dermal surface. The SLN gel possessed sustained
drug release over a period of 24 h. The slower release of
the drug from the gel formulation maintained the drug
concentration over a extended period of time. The per-
meability parameters of piroxicam SLN gel and conven-
tional piroxicam gel are given in Table 4. Parameters such
as steady-state ux (Jss) and permeability coefcient
(Kp) were signicantly (p<0.05) increased in the SLN
gel compared to the conventional gel formulation. The
permeability of piroxicam from SLNs was enhanced 1.1
times compared to conventional free drug gel. This indi-
cates that the SLN gel can not only improve the perme-
ation of piroxicam but can also improve its release rate.
This indicates the suitability of SLN gel for the topical
and/or dermal delivery of piroxicam. The enhancement
of permeation and release rate of piroxicam might be
the occlusive property of the lipid lm formed on the
skin surface. They decrease the trans epidermal water
loss thereby enhancing piroxicam penetration across
the membrane with increased hydration. The effect of
surfactant might be another reason for enhancing per-
meation and release rate of piroxicam through skin. It is

Krishnatreyya, et al.: Piroxicam Nanoparticles for Topical Delivery
S90 Indian Journal of Pharmaceutical Education and Research | Vol 53 | Issue 2 (Suppl)| Apr-Jun, 2019
well known that surfactants have predominant effect on
the permeability of drug through biological membranes.
The surfactant may permeate into the intracellular
regions of stratum corneum and increase their uidity,
eventually solubilizing and extracting the lipid compo-
nents.
In vivo
Efficacy of Piroxicam Loaded SLN
Incorporated Gel
The percentage edema inhibition by piroxicam SLN gel
and piroxicam conventional gel with respect to time are
presented in Figures 8. The percentage edema inhibition
Figure 8: Percentage Edema Inhibition by Piroxicam SLN Gel
(Test) Versus Prepared Piroxicam Conventional Gel (Stand-
ard) After Topical Application on Wister Albino Rats (Mean ±
SD;
n
= 6).
Figure 3: Release Pattern of Piroxicam from Different SLN
Formulations (Mean ± SD;
n
=3).
Figure 4: Viscosity and Thixotropy Curve of the F2 SLN Gel.
Both these Two Figures Determine the Influence of Time and
Temperature on Viscosity of F2 SLN Gel.
Figure 5: Comparative FTIR Spectra of (A) Piroxicam, (B)
Stearic acid, (C) Pluronic F68 (D) Phospholipon 90 G, (E)
Piroxicam SLN gel.
Figure 6: Comparative DSC Thermogram of (A) Piroxicam and
(B) Piroxicam SLN Gel.
Figure 7: Permeation Profile of Piroxicam from SLN Gel and
Conventional Free Drug Gel Across Pig Epidermal Membrane.

Krishnatreyya, et al.: Piroxicam Nanoparticles for Topical Delivery
Indian Journal of Pharmaceutical Education and Research | Vol 53 | Issue 2 (Suppl)| Apr-Jun, 2019 S91
by piroxicam conventional gel was 5.49 ± 0.282% after
1 hr of topical application. However, percentage edema
inhibition after 1 hr on topical application of piroxicam
SLN gel was found to be 20.49 ± 0.543% After 24 h of
application, the percentage edema inhibition by piroxi-
cam conventional gel and piroxicam SLN gel were found
to be 20.08 ± 0.234% and 30.82 ± 0.237% respectively.
This indicates that edema inhibition by piroxicam SLN
gel was comparatively more and differed signicantly
(p < 0.05) compared to edema inhibition by piroxicam
conventional gel. This might be due to the impressive
release pattern of piroxicam from SLN. The fact that
piroxicam SLN gel was successful in inhibition of paw
edema to an extent greater than that of the piroxicam
conventional gel study revealed the applicability and/or
effectiveness of the piroxicam SLN gel system for the
treatment of arthritic pain and inammation.
CONCLUSION
Our study demonstrated the successful preparation of
piroxicam incorporated SLNs by high speed homogeni-
zation and ultrasonication. The formulated SLNs were
subjected to several characteristic evaluations. Evalu-
ation parameters revealed that the percentage of lipid
and surfactant have signicant (p<0.05) effects on the
particle size, drug loading, entrapment efciency and in
vitro release of piroxicam from the SLN formulation.
SLN formulation F2 was the most effective formulation
with optimum particle size, high entrapment efciency
and improved release prole. The in vitro permeation
study indicated revealed sustained permeation of piroxi-
cam from the SLN gel and maintained drug concentra-
tion over a prolonged period of time. The permeation
parameters indicated the enhancement of piroxicam
permeation from SLN gel as compared to conventional
gel. Piroxicam loaded SLN formulation was revealed
to have local as well as systemic effects as studied by
the in vivo anti-inammatory study. Additionally, topical
application of piroxicam SLN gel will be an advantage
thereby reducing the gastrointestinal side effects associ-
ated with its oral administration. The results depict that
the topical application of piroxicam SLN gel system is
an effective and safe alternative to the conventional gel
and oral formulation for the possible management of
inammatory pain and irritation associated with osteo-
arthritis, rheumatoid arthritis, edema etc.
ACKNOWLEDGEMENT
We acknowledge the administrative support obtained
from the Department of Pharmaceutical Sciences,
Dibrugarh University, Assam, India in carrying out this
research work.
FUNDING
The researchers are grateful to the All India Council for
Technical Education for nancial support (20/AICTE/
RIFD/RPS(POLICY-1)24/2013-2014).
CONFLICT OF INTEREST
The authors declare no conict of interest.
ABBREVIATIONS
BCS: Biopharmaceutical classication system; DSC:
Differential scanning calorimetry; Er: Enhancement
ratio; DLS: Dynamic light scattering; FTIR: Fourier
transform infrared spectra; Jss: Steady state ux; Kp:
Permeability coefcient; PDI: Polydispersity index;
SEM: Scanning electron microscopy; SLN: Solid lipid
nanoparticles; TEM: Transmission electron micros-
copy.
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Krishnatreyya, et al.: Piroxicam Nanoparticles for Topical Delivery
S92 Indian Journal of Pharmaceutical Education and Research | Vol 53 | Issue 2 (Suppl)| Apr-Jun, 2019
• Solid lipid nanoparticles are an emerging drug delivery
trend in the recent times. Our experiments have proved
the potentiality and appropriateness of piroxicam
loaded SLN gel system in treating inammatory pain
and irritation symptoms when delivered topically.
SUMMARY
PICTORIAL ABSTRACT
Harshita Krishnatreyya pursued her M. Pharm research work at the Department of Pharmaceutical
Sciences, Dibrugarh University in Assam, India. She is currently a registered Ph.D student at the
Department of Chemical Technology, University of Calcutta, Kolkata, India.
Pranab Jyoti Das, Ph.D from the Department of Pharmaceutical Sciences, Dibrugarh University in
Assam, currently works as a Drugs Inspector with the Central Drugs Standard Control, New Delhi,
India.
Sanjay Dey currently works as an Assistant Professor at the Department of Pharmacy in Techno
India University, Kolkata, India. He obtained his Ph.D degree from the Department of Pharmaceutical
Sciences, Dibrugarh University in Assam, India.
Paulami Pal is a Ph.D research scholar (WOS-A Project, DST) at the Department of Pharmaceutical
Sciences, Dibrugarh University in Assam, India.
Vipin Kumar Sharma works as an Assistant Professor (Pharmaceutics) in the Department of
Pharmaceutical Sciences, Gurukul Kangri Vishwavidyalaya, Haridwar. He was awarded Ph.D degree
from the Department of Pharmaceutical Sciences, Dibrugarh University in Assam, India.
Bhaskar Mazumder is a Professor in
the Department of Pharmaceutical
Sciences, Dibrugarh University, Assam.
His areas of specialization are novel drug
delivery system, dosage form design and
development etc.
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Cite this article: Krishnatreyya H, Dey S, Pal P, Das PJ, Sharma VK, Mazumder B. Piroxicam Loaded Solid
Lipid Nanoparticles (SLNs): Potential for Topical Delivery. Indian J of Pharmaceutical Education and Research.
2019;53(2S):s82-s92.