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EXPERIMENTAL INVESTIGATIONS IN ENCAPSULATION OF PHARMACEUTICALLY ACTIVE INGREDIENTS
_____________________________________________________________
A Synopsis of the Ph.D. Thesis
Submitted to
Gujarat Technological University, Ahmedabad
By
Yashawant Pralhad Bhalerao Enrollment No.: 129990905005 Branch: Chemical Engineering
Under the Supervision of
Dr.Shrikant J. Wagh Principal
Gujarat Power Engineering & Research Institute, Mehsana, Gujarat, India
Under Doctoral Progress Committee
Dr. S. A. Puranik Professor, Atmiya Institute of Technology,
Rajkot.
Dr. Sachin Parikh HOD, Chemical Engineering Department,
LDCOE, Ahmedabad.
GUJARAT TECHNOLOGICAL UNIVERSITY
JULY, 2019
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1. Abstract Microencapsulation comes as an important protection, storage technique, and controlled release
tool for several Pharmaceutically Active Ingredients (PAI), food, cosmetic and other medical
products. Purposes for encapsulation of PAI like thymol may be numerous, such as controlled
release, targeted controlled release, protection/preservation, economic utilization, formulations,
and modification/hiding undesirable property such as taste, odour and touch. Controlled release
helps to maintain an adequate drug concentration in the blood or in target tissues at a desired
value as long as possible and, for this, they are able to control drug release rate, hence control
release of PAI (thymol) is important. Controlled release formulation can be obtained by various
methods such as spray drying, solvent diffusion, nanoprecipitation etc.
The main objective of this study is to encapsulate thymol and determine its controlled or
sustained release by solvent diffusion and nanoprecipitation methods. In this work, the effect of
concentration of thymol (the core material), sodium alginate and ethyl cellulose (the shell
material), stirring speed for synthesis on sustained release and encapsulation efficiency of thymol
loaded beads and microparticles has been studied as discuss in table 1. For method optimization
the statistical design approach using 3 level 2 factors design and Plackett–Burman factorial
Design (PBD) was applied using the Design-Expert® (DoE) software(Version- 9.0.3.1, Stat-Ease
Inc., Minneapolis, MN). Obtained formulation of thymol loaded ethyl cellulose microparticles
shows maximum 98% drug release in 10 h. on the other hand, Formulation of thymol loaded
calcium chloride– sodium alginate beads showed that maximum 95.18±0.43 % drug release in 12
h. No chemical interaction between drug and polymer was found in FTIR study and
beads/particles obtained were spherical and distinct in nature.
Table 1. Overview of experimental methodology.
Core material
Shell material
Method Parameter studied Evaluation
Thymol Ethyl cellulose
Emulsification Solvent Diffusion
Process time, stirring speed for synthesis (RPM), Polymer concentration on encapsulation efficiency and control release
Optimization of the important process variables on Encapsulation Efficiency (EE) and Control Release (CR) by Emulsification solvent diffusion is achieved
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Thymol Ethyl cellulose
Nano precipitation
Process time, stirring speed for synthesis (RPM), Polymer concentration on encapsulation efficiency and control release
Optimization of the important process variables on EE and CR by Nano-precipitation techniques has been achieved
Thymol Sodium Alginate
Emulsification Solvent Diffusion
effect of concentration of thymol, sodium alginate, stirring speed on synthesis, and sustained release properties of thymol loaded sodium alginate beads using emulsion microencapsulation technique
Sustained release of thymol from Sodium Alginate Beads synthesized by Emulsification solvent diffusion Microencapsulation has been studied
2. Brief description on the state of the art of the research topic: The synopsis of the research work on “EXPERIMENTAL INVESTIGATIONS IN
ENCAPSULATION OF PHARMACEUTICALLY ACTIVE INGREDIENTS” gives a brief
description on i) Introduction ii) Literature review iii) Experiment and Analysis iv) Results and
Discussion v) Overall Conclusion and vi) Bibliography.
The introduction and literature review discuss merits, demerits, challenges, and factors affecting
microencapsulation, scope, aims &objectives of PAI microencapsulation, organization of thesis,
information about Emulsion Technique, Microencapsulation, Solvent diffusion,
Nanoprecipitation, Beads, Biodegradable polymers, and Design Expert.
The experimental methodology is to prepare thymol loaded ethyl cellulose micro particles using
solvent diffusion and nanoprecipitation techniques and methods of analysis is to study the effect
of concentration of thymol, sodium alginate, stirring speed on synthesis, and sustained release
properties of thymol loaded sodium alginate beads using emulsion microencapsulation
technique.
Main results are described in experiment and analysis and conclusions are described in results
are discussion and conclusion.
The following is the background of the overall research work undertaken:
Biodegradability of shell material along with its health compatibility and the half-life of the core
material and knowledge of microstructure for Control Release/Sustained Release (CR/SR) are
some of the main issues that must be addressed while studying encapsulation of PAI. The
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encapsulation of essential ingredients in core–shell or matrix particles has been investigated by
many researchers for various reasons, e.g. protection from oxidative decomposition and
evaporation, odour masking or merely to act as support to ensure controlled release. In order to
adapt to different types of active agents and shell materials different microencapsulation methods
have been developed generating particles with a variable shell thickness, range of sizes and
permeability, providing a tool to modify the release rate of the active principle.
Preparative conditions such as concentration ratio, temperature, stirring speed, and nature of
solvent used have deterministic effect on the polymer shell formed around the core material and
hence the release rate. Thus the final objective of encapsulation is controlled release of PAI. But
it (encapsulation) can be engineered according to the need. Microencapsulation can promote
pharmaceutical base products by introducing innovation, added functional properties and thus
added value.
Chemically thymol is 2-isopropyl-5-methylphenol phyto-constituent and classified as
monoterpene found in certain plants [1]. Thymol improves the digestion by relaxing smooth
muscles, prevents menstrual cramps, attenuates respiratory problems and also have wide used in
food industry [2, 3]. Thymol also shows potential antimicrobial properties, that thymol give the
impression to bind to the ergosterol in the membrane, which increases ion permeability and
ultimately results in cell death [4]. Thymol also shows antiseptic, anti-inflammatory, antioxidant
and healing properties and a broad spectrum of biological activity [5-7]. Thymol is able to inhibit
both Gram-positive 1 and Gram-negative bacteria 2 [8]. Antifungal agents like miconazole,
fluconazole, and ketoconazole may be prescribed instead of thymol [9–11]. However,
unselective use and the less number of available antifungal agents have encouraged the
development of resistant strains, especially in immune compromised individuals like thymol [12,
13].
Thymol used as an individual component shows greater predictability effects, allowing the
minimization of adverse effects of thymol. Rapid absorption limits the luminal availability for
antimicrobial activity. Hence, it is assumed that controlled release product could be improved
their effects on the microflora [14].Controlled release behavior of any drug displays control in
rate time and site of drug release in the body. The desired rate of drug release is obtained and can
1Gram-positive bacteria are bacteria that give a positive result in the Gram stain test. 2Gram-negative bacteria are bacteria that do not retain the crystal violet stain used in the gram-staining method.
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enhance the therapeutic effect of drug [15]. Numbers of biodegradable/biocompatible polymers
is used as controlled release carriers and play an important role in such system. Natural, semi
synthetic and synthetic polymers are nowadays widely used as controlled or sustained release
agent. Ethyl cellulose (EC) a hydrophobic polymer is commonly used as a drug carrier in
controlled drug delivery system [16] due to its low toxicity, film forming ability [17, 18].
Controlled release formulation can be obtained by various methods such as spray drying,
lyophilization, solvent diffusion, nanoprecipitation etc. [19]. Solvent diffusion is one of the
techniques widely used in the preparation of microparticles; shows better results than other
microencapsulation techniques such as interfacial polymerization and coacervation widely used
in pharmaceutical industries for various purposes such as protecting drugs from degradation,
protecting body from the toxic effects of the drugs, controlled drug delivery, masking the taste
and odour of drugs and its formulation process is easy and convenient. There are different single
and double solvent diffusion techniques like O/W, W/O, O/W/O, W/O/W. In O/W single
emulsion solvent diffusion method, the polymer is dissolved in a suitable water miscible organic
solvent, and drug solution is emulsified in this polymeric solution. In O/W solvent emulsion
method the oil (Organic solvent) is media while water is serves as continuous aqueous medium
[20, 21].
Controlled release of thymol:
Martin et al. have studied the release behaviour of thymol and p-cymene used as core materials
through Poly Lactic Alcohol (PLA) microcapsules. The microcapsules were obtained by a
coacervation process [22]. Milovanovic et al. have investigated that Cellulose Acetate (CA) is a
shell material for the controlled release of thymol [23].
Antilisterial activity of thymol:
Xiao et al. in their work produced spray-dried capsules from zein solutions (Zein is a class of
alcohol-soluble storage protein). They demonstrate that its non-ionic surfactant can effectively
improve antimicrobial functions in food systems. Spray drying is a practical technology for the
production of an antimicrobial capsule, which is possessed by a manipulated incorporating
surfactant such as Tween 20 [24]. Xue et al. have studied the ability of whey protein isolate
(WPI) and maltodextrin (R) to conjugate the thymol nano emulsifier with propylene glycol (PG)
to improve the antifungal properties of milk [25].
Antioxidant activity of thymol:
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Liolios et al. isolated carvacrol, thymol, p-cymene, and c-terpinene by hydro-distillation
technique and successfully encapsulated in phosphatidyl choline-based liposomes and the
possible improvement of their antioxidant and antimicrobial activities was tested against selected
microbial [26]. Davoodi et al. reveals the antioxidant capacity and physical properties of potato
starch dispersions enriched with polysorbate-thymol micelles. The results showed that potato
starch has essential antibacterial action only in the presence of polysorbatetimol but below
polysorbate thymol alone [27].
Antimicrobial activity of thymol:
Chang et al. prepared thyme oil-in-water nanoemulsions (pH 3.5) as potential antimicrobial
delivery systems. The nanoemulsions were highly unstable to droplet growth and phase
separation, which was attributed to Ostwald ripening due to the relatively high water solubility of
thyme oil. Nano stable thyme oil emulsions were tested for antimicrobial performance, as
opposed to acid-resistant spoilage yeast, Zygosaccharomyces bailii (ZB) [28]. Li et al. revealed
new antimicrobial films based on colloidal nanoparticles zein coated with sodium caseinate (SC)
emulsifier/stabilizer [29]. Li et al. prepared various sub-micron-thymol emulsions with a high
HLB (hydrophilic-lipophilic balance) surfactant by spontaneous emulsification. The emulsions
were then screened for various provocative pathogens to evaluate antimicrobial efficacy [30].
Antispasmodic activity of thymol:
Engelbertz et al. fractionated Thyme fluid extract by Fast Centrifugal Partition Chromatography
(FCPC), Low Pressure Liquid Chromatography (LPLC), and High Pressure Liquid
Chromatography (HPLC) and compounds isolated were identified by spectroscopic methods.
Thyme extracts have antispasmodic activity, which is at least due to synergistic effects of
phenolic volatile oil compounds and the flavone luteolin [31].
Anti-inflammatory activity of thymol:
Riella et al. assess the anti-inflammatory and cicatrizing activities of thymol in rodents, the
peritonitis models of inflammation and analysis, followed by the evaluation of myeloperoxidase
activity (MPO), total cell counts and histological analysis were used [32].
3. Definition of the Problem The active core material in present research work is thymol and study of its controlled and
sustained release is the main objective. It is obvious that the diffusive flux of the core API will
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be governed by the shell material membrane. Therefore, it is required to study the influence of
the parameter such as the effect of concentration of thymol (the core material), sodium alginate
and ethyl cellulose (the shell material), stirring speed for synthesis on sustained release and
encapsulation efficiency of thymol loaded beads and microparticles. We have selected two shell
materials viz. ethyl cellulose and sodium alginate. Both these materials are biodegradable, health
compatible and easily available. Thymol is an important PAI which has enormous applications
described earlier but its doses need to be properly designed in order to avoid side effects and
undesired reactions at the targeted sites due to high concentration. Control release is the answer
to this problem and encapsulation of the PAI is the technique used for this. To the best of our
knowledge, these systems have not been studied before with these objectives.
4. Objectives and Scope of the work i. To investigate suitable encapsulation techniques (Emulsion solvent diffusion
(emulsification diffusion), Nano-precipitation etc.) for Thymol.
ii. To optimize the important process variables (Process time, stirring speed for synthesis
(RPM), Polymer concentration) on encapsulation efficiency and control release by
Emulsion solvent diffusion and Nano-precipitation techniques: A Comparative study.
iii. To study the sustained release of thymol from Sodium Alginate Beads synthesized by
Emulsion solvent diffusion Microencapsulation.
5. Original contribution by the thesis Microencapsulation of thymol in liposomes, solid microparticles and microemulsions, and
polymeric micro/nanoparticles represents a promising strategy for overcoming thymol’s
limitations, lowering their dose and increasing long-term safety of thymol. Low dosages of
thymol are also found to be effective to lower its long term effects on various parts of body.
Standardization is necessary in terms of purity of product and stability. Microencapsulation
formulation can provide an effective alternative for thymol administration in relatively high or
low dosage depending upon application.
Thymol has potential for maintaining and promoting health, also preventing and potentially
treating some diseases such as eczema, cough, cold, dermatitis, antifungal, antibacterial infection
However, the generally low water solubility and stability as well as the high volatility and side
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effects associated with its application have limited its use in medicine. Encapsulation is an
approach that has potential applications in medicinal and health research.
6. Methodology of Research, Results / Comparisons 6.1. Methodology 6.1.1. Experimental System: Thymol loaded Ethyl Cellulose (ETHOCEL), Sodium Alginate micro particles were prepared by
1. Nanoprecipitation Techniques (precipitation in principle)
2. Emulsification Solvent Diffusion Method
3. Emulsified beads
6.2. Experimental Procedure: 6.2.1. Techniques of microencapsulation: Emulsion–diffusion method
Preparation of micro-capsules by the emulsion–diffusion method allows both lipophilic
and hydrophilic active substance encapsulation.
The experimental procedure performed to achieve this requires three phases: organic,
aqueous and dilution.
When the objective is the micro-encapsulation of a lipophilic active substance, the organic
phase contains the polymer, the active substance, oil and an organic solvent partially
miscible with water, which should be water-statured.
This organic medium acts as solvent for the different components of the organic phase. If it
is required, the organic phase can also include an active substance solvent or oil solvent.
The aqueous phase comprises the aqueous dispersion of a stabilizing agent that is prepared
using solvent-saturated water while the dilution phase is usually water.
a. Drug (PAI): thymol (pure thymol crystals , purchased from Loba Chemie, Mumbai )
b. Polymer: Ethyl cellulose “ETHOCEL” (as a Gift sample from ICT, Mumbai)
c. Stabilizer/surfactant: Tween 80 (purchased from Loba Chemie, Mumbai )
d. Organic solvent: Ethanol
6.2.2. Techniques of microencapsulation: nanoprecipitation
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Thymol loaded ethyl cellulose nanoparticles were prepared by means of the
nanoprecipitation method using the same concentrations of drug and other excipients as
per above method.
In this technique, the drug and polymer was dissolved in dichloromethane to form
diffusion phase (Solution A). This phase was then added into aqueous dispersing medium
(Tween 80 + distilled water) by using syringe under stirring speed 700 rpm. The
dispersing phase was constituted of a liquid in which the polymer is insoluble – the non-
solvent containing a surfactant (Tween 80).
In this technique the ethyl cellulose deposit on the interface between the aqueous and the
organic solvent, caused by fast diffusion of the organic solvent, leads to the formation of
a colloidal suspension. The freshly formed nanoparticles by this technique in the form of
colloidal particles are separated by Whatman filter paper and dried for further
characterization
6.3. Encapsulation efficiency % (EE) and % drug release (DR): The amount of thymol in thymol loaded ethyl cellulose nanoparticles was determined using a
UV-Vis spectrophotometer (Hitachi U2900). Percent encapsulation efficiency was calculated
using the equation (1):
% EE = (Wt – Ws) x 100/Wt (1)
Where, Wt is the total weight of the drug used in formulation and Ws is weight of the drug in
supernatant.
Drug release study was performed using Dissolution Testing Type II Apparatus at 37 ± 0.5°C
and at 100 rpm using 900 ml phosphate buffer having pH 6.8. Nanoparticles equivalent to 40 mg
of thymol were used for the dissolution study. At predetermined interval 5 ml of aliquot was
withdrawn and analyse spectrophotometrically and an equal amount of fresh buffer was added to
maintain the sink condition.
6.4. Results and Discussions The main objective of this study is to prepare thymol loaded ethyl cellulose micro particles using
solvent diffusion and nanoprecipitation techniques and study the effect of concentration of
thymol, stirring speed on synthesis, and sustained release properties of thymol loaded sodium
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alginate beads using emulsion microencapsulation technique. For optimization method, the
statistical design approach was employed and 3 level 2 factors design was applied using design
expert (version 9.0.3.1) software and design approach of Plackett–Burman factorial Design
(PBD) of experiment was performed using Design-Expert® (DoE) software (Version- 9.0.3.1,
Stat-Ease Inc., Minneapolis, MN).
6.4.1. Formulation and evaluation of thymol loaded ethyl cellulose microparticles using solvent diffusion and nanoprecipitation methods
The solvent diffusion technique and nanoprecipitation technique are used for micro particles
formulation to investigate suitable encapsulation technique for thymol. The Obtained
formulation shows maximum 98% drug release in 10 h. Thymol loaded ethyl cellulose
microparticles were successfully prepared by solvent diffusion as well as nanoprecipitation
method without any incompatibility. This study observed that nanoprecipitation method gives
quite better results than the solvent diffusion method and seems to be promising for sustained
delivery of thymol.
Encapsulation efficiency (EE) of thymol loaded ethyl cellulose micro particles shows direct
relationship with the polymer concentration i.e. encapsulation efficiency is higher at higher
polymer concentration and minimum with lower polymer concentration. The EE obtained in the
range of 69.11 % to 81.3 % for the solvent diffusion method and 63.12 % to 75.47 % for the
nanoprecipitation technique. Design expert software was employed; using 3 level 2 factor design
total 13 runs was carried out for each method to study the effect of independent variables on
dependent variables. Analysis of variance (ANOVA) and all statistical analysis were also
performed using the Design Expert (DoE) software (Version- 9.0.3.1, Stat-Ease Inc.,
Minneapolis, MN) shows linear model.
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Figure 1: 2D counter plot and 3D surface plot (A and B) showing the effect of independent
variables on encapsulation efficiency and drug loading prepared by solvent diffusion and
nanoprecipitation method respectively.
6.4.1.1.Encapsulation efficiency and In-vitro drug release studies:
Figure 2: Display of the release behavior of micro/nanoparticles by solvent diffusion
method (A) and nanoprecipitation method (B).
The EE obtained in the range of 69.11 % to 81.3 % for the solvent diffusion method and 63.12 %
to 75.47 % for the nanoprecipitation technique. Encapsulation efficiency is directly proportional
to the polymer concentration. Drug release profile for all the runs was carried out using in-vitro
release study at pH 6.8. Cumulative percent drug release for solvent diffusion runs are in the
range of 83.43% to 94.38% while in nanoprecipitation technique 84.91% to 98.71 % in 10 h
shows control release (Figure 1). The 2D counter plot and 3D response surface plot in Figure 2A
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shows the effect of polymer concentration and surfactant concentration on EE and DR for the
solvent diffusion technique. The EE efficiency increases with the increase in polymer
concentration and drug release also shows the same behavior. Similarly, in nanoprecipitation
technique 2D counter plot and 3D response surface plot (Figure 2B) shows the direct relationship
with the polymer and surfactant concentration. When we compare the EE and DR data in both
the methods the data obtained by solvent diffusion method is quite better than nanoprecipitation
method in especially in terms of release behavior.
In this study thymol was successfully entrapped in the biodegradable polymer ethyl cellulose by
solvent diffusion and nanoprecipitation method. Drug release from prepared polymer matrix was
observed slowly in in-vitro release profile up to 10 h. Both methods are suitable for the
nanoparticle preparation. No chemical interaction was found in FTIR study and particles
obtained are spherical and distinct in nature. Comparison of two methods showed that
nanoprecipitation method gives better encapsulation efficiency results while particles prepared
by solvent diffusion method gives the more controlled action in in-vitro release.
Formulation shows maximum 98 % drug release in 10 h. Thymol loaded ethyl cellulose
microparticles were successfully prepared by solvent diffusion as well as nanoprecipitation
method without any incompatibility. This study observed that nanoprecipitation method gives
quite better results than the solvent diffusion method and seems to be promising for sustained
delivery of thymol.
6.4.2. Sustained release of thymol from Sodium Alginate Beads synthesized by Emulsion solvent diffusion Microencapsulation
The concentration of polymer (X1) and concentration of surfactant (X2) are considered as
independent variables while percent encapsulation efficiency (Y1) and percent drug release (Y2)
are as dependent variables for both methods Preparation of thymol loaded calcium chloride –
sodium alginate beads was carried out using the emulsion microencapsulation technique.
The PBD is an efficient approach to evaluate the results which are shown in Table 1.
Table 1: The Plackett-Burman Experimental Design matrix (in coded level) and
experimental results
Runs Variables Response A B C D E F G H J K L EE (%) DR (%) 1 -1 1 1 -1 -1 1 1 -1 1 -1 -1 65.31± 0.56 63.44±0.54
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2 -1 1 1 -1 1 1 -1 1 -1 -1 -1 96.81± 1.59 87.03±0.65 3 -1 1 -1 -1 1 1 1 1 -1 1 1 91.86± 2.74 92.9±0.74 4 -1 -1 -1 -1 -1 -1 1 -1 -1 -1 -1 95.04± 2.12 30.06±0.73 5 1 1 -1 1 -1 -1 -1 1 1 1 -1 96.20± 1.25 95.07±0.72 6 -1 -1 -1 1 -1 -1 -1 1 -1 1 -1 87.79± 2.55 79.96±0.92 7 1 -1 1 -1 1 1 1 -1 1 1 -1 94.66± 0.57 78.98±0.12 8 1 -1 -1 -1 1 -1 1 1 -1 -1 1 80.27± 2.43 95.18±0.43 9 1 1 -1 1 1 -1 1 -1 1 -1 1 93.24± 1.86 85.72±0.70 10 1 -1 -1 1 1 1 -1 1 1 -1 1 31.18± 0.94 94.63±0.54 11 -1 1 1 -1 1 -1 -1 -1 1 1 1 92.34± 0.83 91.26±0.43 12 1 -1 1 1 -1 1 -1 -1 -1 1 1 84.31± 2.24 92.03±0.87
The release of thymol was evaluated using phosphate buffer (pH 6.8) as the release medium.
Sustained drug release was observed in the range of 30.06±0.73 to 95.18±0.43 % in 12 h study of
all the experimental runs. The EE of different experimental runs of the beads is reported in Table
2. The 3D response surface plots are useful in understanding about the main and interaction
effects of the independent variables. To visualize the effect of independent variables on each
response, 3D response surface plots (Figure 3) were constructed. The % EE of the beads was
calculated to be in the range 31.18 % to 96.81 %.
(A) (B)
Figure 3: 3D surface plots showing independent and interaction effect of thymol and
sodium alginate on (A) encapsulation efficiency; (B) drug release, prepared by
microencapsulation.
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To examine the interaction between the drug and polymer FTIR spectra of pure thymol, ethyl
cellulose and formulation were obtained using Fourier transform infrared spectrophotometer.
No chemical interaction between thymol and sodium alginate was found in FTIR study and
beads obtained were spherical and distinct in nature. Formulation showed that
microencapsulation method gives sustained action in in-vitro release. Formulation shows
maximum 95.18±0.43 % drug release in 12 h. Thymol loaded sodium alginate beads can be
successfully prepared by emulsion microencapsulation method without any incompatibility. This
study observed that the emulsion method gives promising results for sustained delivery of
thymol.
XRD spectra of pure thymol produced peaks at different 2Ɵ angles and found to be crystalline in
nature while, in case of ethyl cellulose, sodium alginate and the formulations, less intense peaks
are observed than that of pure thymol. This result tells that majority of the drug was entrapped
within the polymer and dispersed homogeneously at molecular level. The XRD patterns of
optimized formulation prepared by both methods showed 43.7% crystalline and 56.3 %
amorphous regions. Thus, indicating absence of distinct diffraction peaks of thymol. This result
tells that majority of the drug was entrapped within the polymer and is dispersed homogeneously
at molecular level
Decrease in particle size shows increase in dissolution rate and ultimately the absorption.
The PDI of optimized batch for solvent diffusion method is 0.122(nm) while the PDI of batch for
nanoprecipitation method is 0.223(nm). The results indicate that the particles obtained by the
solvent diffusion method shows narrow particle size distribution than the particles obtained by
nanoprecipitation method (Figure 4).
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Figure 4: Showing the particle size distribution of optimized batches by solvent diffusion
and nanoprecipitation method.
The surface morphology of thymol loaded ethyl cellulose micro particles was examined by FE-
SEM. The obtained micro particles are smooth, discrete, and spherical with sharp edges without
any cracks or erosion at the surface. The particles prepared by solvent diffusion method are in
the range of 6 to 19 μm and particles prepared by nanoprecipitation method have size range 27 to
38 μm. The surface morphology of thymol loaded 2% sodium alginate beads was examined by
FE-SEM as depicted in Figure 5 (A & A1). Thymol loaded 4% sodium alginate beads were
examined by FE-SEM (Figure 5 (B & B1)). The obtained beads are smooth, discrete, and
spherical with sharp edges without any cracks or erosion at the surface before drying (Figure 5
(A & B)), but after drying, an irregular and rough surface was observed (Figure 5 (A1 & B1)).
The beads prepared by microencapsulation method are in the range of 6 to 38 μm
Figure 5: Microscopic analysis of beads obtained by microencapsulation : A, A1 before and
after drying at 2% sodium alginate beads respectively; and B, B1 before and after drying
at 4% sodium alginate beads respectively.
A1 A
B1 B
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7. Achievements with respect to objectives i. Microencapsulation of thymol is successfully carried out with three methods/systems. For
optimization method, the statistical design approach of Plackett–Burman and factorial
design of experiment was performed using Design-Expert® (DoE) software.
ii. Optimization of the important process variables on encapsulation efficiency and control
release by Emulsion solvent diffusion is achieved.
iii. Optimization of the important process variables on encapsulation efficiency and control
release by Nano-precipitation techniques has been achieved.
iv. Sustained release of thymol from Sodium Alginate Beads synthesized by Emulsion
solvent diffusion Microencapsulation has been studied.
v. Comparison of Emulsion solvent diffusion Microencapsulation and Nano-precipitation
techniques on microencapsulation of thymol has been done.
8. Conclusion In this study thymol was successfully entrapped in the biodegradable polymer ethyl cellulose by
solvent diffusion and nanoprecipitation methods. Drug release from prepared polymer matrix
was observed slowly in in-vitro release profile up to 10 h. Both methods are suitable for the
microparticle preparation. No chemical interaction was found in FTIR study and particles
obtained are spherical and distinct in nature. When we compare both the methods with respect to
the encapsulation efficiency and drug release, both methods have their own better results i.e.
nanoprecipitation method gives better encapsulation efficiency while particles prepared by
solvent diffusion method gives more controlled action in in-vitro release.
Thymol was successfully entrapped in the calcium-alginate by microencapsulation method. The
% EE of the beads ranged from 31.18 % to 96.81 %. Drug release from prepared polymer matrix
was observed slowly in in-vitro release profile up to 12 h. Formulation showed that
microencapsulation method gives more sustained action in in-vitro release. Formulation shows
maximum 95.18±0.43 % drug release in 12 h. No chemical interaction was found in FTIR study
and beads obtained were spherical and distinct in nature.
Thymol loaded calcium-alginate beads were successfully prepared by microencapsulation
method without any incompatibility. This study observed that microencapsulation method gives
satisfactory results and seems to be promising for sustained delivery of thymol. As the thymol
concentration increases, the EE also increases. On the other hand with increasing concentration
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of sodium alginate the EE slightly decreases. Whereas an increasing pattern of drug release was
observed with increase in the concentration of thymol and sodium alginate.
9. Copies of papers published and a list of all publications arising from the thesis i. Yashawant. P. Bhalerao, Shrikant J. Wagh. Formulation and evaluation of thymol loaded
ethyl cellulose microparticles using solvent diffusion and nanoprecipitation methods: A
comparative factorial design approach. International Journal of Pharmaceutical Research
July- Sept 2018. Vol 10, Issue 3.
ii. Yashawant. P. Bhalerao, Shrikant J. Wagh. In vitro sustained release study of Thymol from Sodium Alginate Beads synthesized by Emulsion Microencapsulation. International
Journal of Pharmaceutical Research, Apr - June 2019. Vol 11, Issue 2.
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