Formulation and in-vitro Evaluation of Disclofenac Microspheres for Sustained Drug Delivery

1. Formulation and in-vitro evaluation of
Diclofenac Microspheres for sustained
drug delivery
PRESENTED BY:
K. Milind kumar (13AD1R0022)UNDER THE GUIDENCE OF:
Dr. UMA MAHESWAR RAO
M Pharm Ph.D

2. contents
• Introduction to microspheres
• Literature review
• Aim and objectives
• Plan of work
• Drug profile
• Excipient profile
• Pre-evaluation parameters
• General methods of preparation
• Formulation method
• Methodology
• Evaluation parameters
• Results
• conclusion

3. Introduction to microspheres
• Microspheres are characteristically free flowing powders
consisting of proteins or synthetic polymers which are
biodegradable in nature and ideally having a particle size less
than 1000 micrometer.

4. Advantages
• Taste and odour masking of drug
• Improves bio- availability and reduces side effects.
• Prolonged and controlled GI transit of the dosage forms.
• Protection of the drug against environment.
• Production of sustained release and controlled release
medication.
• To have better therapeutic efficiency

5. Mechanism of action of
microspheres
• The drug could be released from microspheres by any one of
the three methods;-
1. Osmotically driven burst mechanism
2. Pore diffusion mechanism.
3. Degradation of polymer.
1. Osmotically driven burst mechanism :-
In osmotically driven burst mechanism, water diffuse in to the
core through bio-degradable or non-biodegradable coating
creating sufficient pressure that ruptures the membrane.

6. Mechanism of microspheres
2. Pore diffusion method :-
The pore diffusion method is named so because as penetrating
water front continue to diffuse towards the core.
3. Polymer erosion method :-
In polymer erosion method, loss of polymer is accompanied
by accumulation of the monomer in the release medium. The
erosion of the polymer begins with the changes in the
microstructure of the carrier as water penetrates it leading to
the plasticization of the matrix.

7. • S. Haznedar et al., (2014) were prepared by solvent
evaporation method using acetone/liquid paraffin system. The
influence of formulation factors (stirring speed, polymer:drug
ratio, type of polymer, ratio of the combination of polymers)
on particle size, encapsulation efficiency and in vitro release
characteristics of the microspheres were investigated.
• M Nappinnai et. al., (2010) Microspheres of diltiazem
hydrochloride were formulated using combination of
polyethylene glycol 6000 and Eudragit RS 100 alone by
solvent evaporation. Six formulations prepared by using
different drug to polymer ratios, were evaluated for relevant
parameters and compared with marketed SR capsules
Literature review

8. Aim and objectives
Aim:
• Microspheres appear to be an exploitable delivery system for
controlled or sustained release of drugs.
• The purpose of this study to prepare polymeric microsphere
containing Diclofenac by using Ionotropic gelation technique.
• The resultant microspheres were evaluated for various
parameters
• The aim of this study is to formulate and evaluate of
diclofenac microspheres by Ionotropic gelation technique.
Objective:
• The Objective of the present study was to microencapsulate
the diclofenac drug for sustained release.

9. Plan of work
STEP-1: Literature survey
STEP-2 : API Characterization
Drug –excipient compatability studies
STEP-3: Formulation development by using Ionotropic
gelation
method
STEP-4: Evaluation of microspheres
STEP-5: To Study Invitro Dissolution Profile
STEP-6: To Perform Stability Study of Selected Formulation
STEP-7 : Results
STEP- 8: Conclusion

10. Drug profile
• Name: Diclofenac
• Description: A non-steroidal anti-inflammatory agent
(NSAID) with antipyretic and analgesic actions. It is primarily
available as the sodium salt.
Physical appearance: white to slightly yellow crystalline
powder.
• Solubility: freely soluble in methanol, soluble in ethanol,
sparingly soluble in water, slightly soluble in acetone, and
partially insoluble in ether.
• Category: Anti-Inflammatory Agents, Non-Steroidal
Cyclooxygenase Inhibitors

11. Mechanism of action of diclofenac
Mechanism of action: The anti inflammatory effects of
diclofenac are due to inhibition of both leukocyte migration
and the enzyme cyclooxygenase (COX-1 and COX-2), leading
to the peripheral inhibition of prostaglandin synthesis. As
prostaglandins sensitize pain receptors, inhibition of their
synthesis is responsible for the analgesic effects of diclofenac.
Antipyretic effects may be due to action on the hypothalamus,
resulting in peripheral dilation, increased cutaneous blood
flow, and subsequent heat dissipation.

12. Drug profile
Pharmacokinetics
• Absorption: Completely absorbed from gastrointestinal tract
• Volume of distribution: 1.3 L/ kg
• Protein binding: > 99%
• Metabolism: Hepatic
• Half life: 2 hours
• Elimination: Urine-60%, biliary-40%
• Uses:
Diclofenac is used commonly to treat mild to moderate
postoperative or post-traumatic pain, in particular organ when
inflammation is also present, and is effective against menstrual
pain and endometriosis.
• To reduce inflammation and as analgesic reducing pain in
certain conditions.

13. Excipient profile
• Ethyl cellulose
• Synonym: Cellulose, ethyl ether
Solubility: Soluble in a wide variety of organic solvents,
including aliphatic alcohols, ethers, ketones, aromatic
hydrocarbons.
• Pharmaceutical Applications:
• Ethyl cellulose polymers are well suited for controlled release
matrix tablet formulations.
• Ethyl cellulose designed specifically for pharmaceutical
formulations when the EC is used in an un solubilized form
such as in direct compression SR tablets, granulation process.
• In buccal tablets ethyl cellulose is used as a backing layer.

14. Excipient profile
• Sodium alginate
• Synonyms : Algin, alginic acid, sodium salt, E401, Kelcosol,
Keltone, Protanal, sodium polymannuronate
• Solubility : Soluble in cold water, forming a viscous colloidal
solution, insoluble in alcohol, ether and chloroform, but
soluble in mixture of methyl alcohol and methylene chloride
• Applications: It is a suspending, viscosity increasing and
film-forming agent. It is also used as a tablet binder and as an
adhesive ointment ingredient. The E- grades are generally
suitable as film formers and K- grades as thickeners.

15. Excipient profile
• Methanol :- Synonym : Alcohol
• Applications: methanol is highly toxic and unfit for
consumption. At room temperature, it is a polar liquid, and
is used as an antifreeze, solvent, fuel, and as a denaturant for
ethanol. It is also used for producing biodiesel via trans
esterification reaction.
• Eudragit:- Synonym : Dried metha acrylic acid co polymer
• Solubility : eudragit dissolves in methanol, ethanol, isopropyl
alcohol and acetone.
• Applications in Pharmaceutical Technology
• Polymethacrylates are primarily used in oral capsule and tablet
formulations as film coating agent.

16. Pre-Formulations studys
• Determination of absorption maxima (λmax) for Diclofenac
sodium:-
A 10mcg/ml standard solution of Diclofenac was scanned on a
double beam spectrophotometer against respective media
blanks. An absorption maximum (λmax) of 243 nm was
obtained for all solutions and was selected to prepare standard
curve.
• Preparation of standard curve for Diclofenac sodium:-
Standard curves for Diclofenac were obtained in 7.4 pH
buffers and water. Aliquotes of 1, 2, 3, 4 and 5 ml of
Diclofenac sodium standard solution of 100mcg/ml (stock
solution-II) was taken and diluted to obtain concentrations
from 10 mcg/ml with appropriate media.

17. S. no Concentration (µg/ml) Absorbance
1 0 0
2 1 0.236
3 2 0.422
4 3 0.623
5 4 0.834
6 5 0.996
The absorbances of solutions were determined at 243 nm
against respective media as blank. The experiment was
repeated six times for each buffer and a calibration curve
was determined from the mean value.

18. Calibration curve
y = 0.0199x + 0.0203
R² = 0.998
0
0.2
0.4
0.6
0.8
1
1.2
0 10 20 30 40 50 60
Absorbance
Concentration (µg/ml)
Absorbance
Linear (Absorbance)

19. General methods of preparation
• Emulsion solvent evaporation methods
Single emulsion technique
Double emulsion technique
• Polymerization techniques
• Phase separation coacervation techniques
• Spray drying and spray congealing
• Solvent extraction
• Hot melt micro-encapsulation
• Fluidized bed coating
• Centrifugal extrusión technique

20. Formulation table
Ingredients F1 F2 F3 F4 F5
Drug 500 mg 500 mg 500 mg 500 mg 500 mg
Sodium
alginate
1gm - 500 mg
HPMC 1gm - - - -
Ethylcellul
ose
_ - 1gm - -
Eudradit - 1 gm - - -
Methanol 5 ml 5 ml 5 ml 5 ml 5 ml
Tragacanth
- - - - 1 gm
Cacl 2 5gms 5gms 5gms 5gms 5gms

21. Methodology
Drug
dispersed into
Polymer solution and placed on magnetic stirrer
While stirring 400 rpm
Passed through syringe in aqueous solution of calcium chloride
Microspheres are formed
Filtered and Washed with distilled water
Dried and stored in dessicator

22. Evaluation parameters
• The prepared sustained release microspheres were evaluated
for various parameters ;-
• Yield of sustained microspheres
• Particle size analysis
• Surface morphology of sustained release microspheres
• Drug entrapment efficiency
• Invitro drug release studies

23. Yield of sustained microspheres
• The dried microspheres were weighed and percentage yield of
prepared microspheres were calculated by using following
formula.
• The yield of microspheres was calculated from the amount of
microspheres obtained divided by the total amount of all non-
volatile components.
Actual weight of the microspheres
% Yield = ----------------------------------------------------× 100
Total weight of all non-volatile components

24. Percentage yield
There was no significant difference observed by solvent
mixture and concentration of emulsifying agent but
concentration of polymer and stirring rate are affecting more
on the percentage yield of microspheres.
Test F1 F2 F3 F4 F5
% yield 85.85 79.55 88.33 84.23 85.23

25. Surface morphology of
microspheres
• The surface morphology of the sustained microspheres was
studied with the aid of a Scanning Electron Microscope
(SEM). The external and internal morphology of microspheres
were studied.The sample was loaded on a copper sample
holder and coated with carbon followed by gold.
• photomicrograph of the microspheres before and after the
release of drug was taken.The quality of the microspheres and
the nature and size of the pores developed on the surface can
be studied.

26. SEM photographs showed the presence of drug crystal
on the surface of microspheres revealing that the
microspheres were having some rough surface. The drug
crystals on microspheres were may be due to the presence
of un entrapped drug in dispersion medium.

27. Particle size analysis
• The particle size of the microspheres was measured by optical
microscopy. The eyepiece micrometer was calibrated using a
stage micrometer and the calibration factor was used further in
the calculation of the size of microspheres.Microspheres were
finely spread over a slide and visualized under an optical
microscope using an eyepiece micrometer. The particle
diameters of more than 50 microspheres were measured
randomly
Test F1 F2 F3 F4 F5
Particle
size (um)
88.39 93.64 96.72 85.24 87.24

28. Drug entrapment efficiency
• The amount of drug entrapped was estimated by crushing 50
mg of microspheres using mortar and pestle, and extracting
drug with aliquots of 7.4 pH buffer repeatedly. The extract was
transferred to a 100 ml volumetric flask and the volume was
made up using7.4 pH buffer. The solution was taken in a
beaker and sonicated in a bath sonicator for 2 hours. The
solution was filtered and absorbance was measured after
suitable dilutions spectrophotometrically at 243 nm against an
appropriate blank.
• The amount of drug entrapped in the microspheres was
calculated using the formula

29. DEE Studies
Amount of drug actually present
DEE = ---------------------------------------------- × 100
Theoretical drug load expected
Test F1 F2 F3 F4 F5
DEE 49.97 49.01 49.93 48.26 49.27
From the table it is observed that the increase in drug
polymer ratio results in increase in entrapment efficiency.
This is due to viscosity of the polymers used which results in
more entrapment of the drug.

30. Invitro drug release studies
• In vitro drug release studies were carried out for all
formulations in Franz diffusion cell. Microspheres equivalent
to 10 mg of Diclofenac sodium were poured. 5ml is
withdrawn at a predetermined intervals and equal volume of
dissolution medium was replaced to maintain sink conditions.
The necessary dilutions were made with 7.4 pH buffer and the
solution was analysed for the drug content
spectrophotometrically using UV-Visible spectrophotometer
at 243 nm against an appropriate blank. Three trials were
carried out for all formulations. From this cumulative
percentage drug release was calculated and plotted against
function of time to study the pattern of drug release.

31. Invitro studies
.
Time (hrs) F1 F2 F3 F4 F5
1 63.519 35.185 37.449 34.225 57.421
2 69.471 52.680 53.109 53.109 53.109
3 75.628 76.452 57.949 67.229 58.910
4 86.990 84.521 63.232 73.541 63.124
5 91.907 85.845 65.664 75.143 65.129
6 94.432 87.997 68.725 78.613 69.907
7 97.520 90.159 70.979 80.812 70.153
8 99.826 91.508 73.656 83.612 76.612
9 ....... 95.743 80.256 88.312 81.908

32. Results
parameters F1 F2 F3 F4 F5
% Yield 85.85 79.55 88.33 84.23 85.23
Particle size 88.39 93.64 96.72 85.24 87.24
DEE 49.97 49.01 49.93 48.26 49.27
Invitro
release
99.826 95.743 80.256 88.312 81.908

33. Conclusion
• From the above results it is clear that out of 5 formulations, F1
is considered as the best formulation because of the maximum
amount of drug release and more DEE with average particle
size and percentage yield when compared to the other
formulations. It is due to the more drug to polymer ratio. So F1
is the best formulation.