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RESULT AND DISCUSSION
110
5. Mefenamic acid ......................................................................................................111
5.1. Mefenamic acid Spherical crystallization ...........................................................111
5.1.1 Selection of the best liquid proportions ............................................................111
5.1.2 Influence of the various bridging liquids ..........................................................113
5.1.3 Influence of bridging liquid on particle size: ....................................................113
5.1.5 Optimum stirring time or residence time ..........................................................118
5.1.6 Influence of the temperature .............................................................................118
5.1.7 Influence of mode of addition of bridging liquid..............................................118
5.2 Recrystallization of Mefenamic acid: ..................................................................120
5.3 Crystallization by spray drying technique: ..........................................................120
5.4 Crystallization by freeze drying techniques:........................................................122
5.5 Super cooling crystallization (solidification technique): .....................................123
5.6 Characterization of Mefenamic acid spherical agglomerates ..............................123
5.6.1 Percentage yield: ...............................................................................................123
5.6.2 Drug content......................................................................................................124
5.6.3 Melting point:....................................................................................................124
5.6.4 Water content ....................................................................................................125
5.6.5 Residual solvent content: ..................................................................................125
5.6.6 Differential scanning calorimetry (DSC) ..........................................................126
5.6.7 FT-IR Spectroscopy: .........................................................................................129
5.6.8 X-ray analysis: ..................................................................................................132
5.6.9 Surface topography by scanning electron microscopy (SEM): ........................143
5.7 Evaluation of prepared crystals: ...........................................................................146
5.7.1 Micromeritic properties: ...................................................................................146
5.7.2 Mechanical properties: ......................................................................................149
5.7.3 Solubility studies:..............................................................................................154
5.7.4 Dissolution behaviour of crystals: ....................................................................156
5.7.5 Various parameters of model fittings of Mefenamic acid samples...................159
5.7.6 Stability study: ..................................................................................................161
5.8 Preparation of conventional tablet: ......................................................................162
5.8.1 Evaluation of Mefenamic acid tablets: .............................................................162
5.8.2 Dissolution studies of tablets: ...........................................................................162
RESULT AND DISCUSSION
111
5. Mefenamic acid
5.1. Mefenamic acid Spherical crystallization
A typical spherical crystallization solvent system involved a good solvent, a
poor solvent for a drug and a bridging liquid. The selection of these solvents depends
on the miscibility of the solvents and solubility of the drug in individual solvents.
Mefenamic acid is freely soluble in Tetra hydro furan (THF). It is necessary to select
a solvent in which Mefenamic acid exhibits the maximum solubility. In fact, a very
concentrated solution of drug could increase the densification of the material during
the crystallization process. The addition of bridging liquid (isopropyl acetate)
promotes the transfer of the drug to a third emulsified phase in which crystal
agglomerates densify and grows spherically164, 165
. Moreover, Tetrahydrofuran (THF)
is miscible in all proportion with water and isopropyl acetate.
5.1.1 Selection of the best liquid proportions
Mefenamic acid soluble in Tetrahydrofuran, ethanol, methanol, isopropyl
acetate and chloroform. Based upon high volatility, high viscosity and low reactivity
of THF and isopropyl acetate were determined to be a suitable crystallization solvent.
Mefenamic acid showed high solubility in THF (1.75gm/10ml). Hence, THF used as
good solvent.
If the ternary diagram is envisaged, isopropyl acetate and water are like an
emulsion in a large of area of this diagram. To select the best solvent composition, a
ternary diagram was constructed. The points on the vertex correspond to a pure liquid;
those on the sides correspond to a mixture of only two liquids. Since the presence of
three liquids is necessary (good solvent, bridging solvent and poor solvent) for
spherical crystallization, points on the sides of the triangle are excluded. 36 points
RESULT AND DISCUSSION
112
remain for experiments. Each triangle in the ternary diagram was investigated for the
crystallization. The optimal ratio for spherical crystallization is found in zone Figure
20. These proportions of THF/water/ isopropyl acetate were finally chosen for the
study166
.
Mefenamic acid is also soluble in isopropyl acetate, Isopropyl acetate is also
soluble in THF and capable of forming liquid bridges between the particles of
mefenamic acid.
Figure 20: Ternary diagram of Mefenamic acid – THF/water/ isopropyl acetate.
To optimize Mefenamic acid spherical crystallization by THF/water/ isopropyl
acetate system, several parameters were considered; among these are, influence of
bridging liquid, influence of the rotation speed, influence of the temperature,
influence of mode of addition of bridging liquid and stirring time164, 166
.
RESULT AND DISCUSSION
113
5.1.2 Influence of the various bridging liquids
Among Isopropyl acetate, chloroform and toluene, which were used as
bridging liquids, Isopropyl acetate and chloroform produced spherical crystals.
However spherical crystals formed using Isopropyl acetate showed improved
micromeritic properties, solubility and dissolution properties compared to spherical
agglomerates produced by chloroform. Hence Isopropyl acetate was selected as a
bridging liquid for spherical crystallization of Mefenamic acid165, 168
.
5.1.3 Influence of bridging liquid on particle size:
The influence of the composition of the good solvent, poor solvent and
bridging liquid in the system on the particle size of spherically agglomerated crystals
were investigated164, 168
. The solvent composition was varied with respect to isopropyl
acetate and water in the crystallization medium, with the amount of THF being
constant.
The average size of agglomerated crystals increased with increasing amount of
Isopropyl acetate due to enhanced agglomeration of crystals. The excess bridging
liquid on the surface of crystals allowed greater coalescence of crystals. When 9 ml of
isopropyl acetate was used crystals were small, brittle due to lack of enough bridging
liquid. With further increase in isopropyl acetate amount (11 ml), lumps were
obtained. Hence the 10 ml of bridging liquid was selected for the preparation of
spherical agglomerates of Mefenamic acid (Table 9, 10 & Figure 21).
RESULT AND DISCUSSION
114
Table 9 Amount of solvents selected from phase diagram used to prepare
spherical crystals of Mefenamic acid
Amount (ml) of
tetrahydrofuran
(good solvent)
Amount (ml) of
Water used in
ml (poor
solvent)
Amount (ml) of
Isopropyl acetate used
in ml (bridging liquid)
Percent of
bridging liquid
20 71 9 9
20 70 10 10
20 69 11 11
Table 10 Effect of volume of bridging liquid on size distribution of spherical
crystals
Particle diameter (µm)
Percentage Frequency
9 ml 10ml 11 ml
500 11.32 8.14 6.05
710 19.44 16.12 8.72
1000 30.5 25.85 15.44
1200 38.78 29.37 24.89
1450 29.14 24.65 35.09
1700 6.23 9.17 8.65
RESULT AND DISCUSSION
115
Figure 21 Effect of volume of bridging liquid on size distribution of Spherical
crystals
0
5
10
15
20
25
30
35
40
45
0 200 400 600 800 1000 1200 1400 1600 1800
% F
req
uen
cy
Particle diameter (µm)
9 ml 10 ml 11 ml
RESULT AND DISCUSSION
116
5.1.4 Influence of the rotation speed of the stirring
Different stirring rates were studied as they influence the secondary characters
of spherical crystals. Solvent composition (THF: Isopropyl acetate: water) of
crystallization medium was kept constant, different stirring rates 300±25, 400±25,
500±25, 600±25 and 700±25 rpm were used (164, 168). The movement of droplets
within the medium induces circulation inside the droplets. The intensity of this
internal circulation depends on the speed. Higher speed induces crystal agglomerate
destruction (600±25 & 700±25). A lower stirring rate reduces the possibility of
obtaining spherical crystals (300±25 & 400±25). Size distributions of particles at
various degrees of agitation are shown in the Figure 22 & Table 11. The size of
agglomerates was very much dependent on the degree of agitation. For a constant
period of stirring, as the speed of agitation is increased, the size of agglomerates
obtained was decreased. This may be due to the fact that as the speed of agitation
increases the impact energy for collision of particle increases due to increased
turbulence, resulting in agglomerates, which are more compact and dense. Hence
500±25 rpm was selected for the preparation of spherical agglomerates of the
Mefenamic acid.
Particle size distribution was narrow at 600 rmp, while 400 rpm and 500 rpm
produced agglomerates of broad distribution.
RESULT AND DISCUSSION
117
Table 11 Effect of intensity agitation on size distribution of spherical crystals
Particle diameter (µm)
Percentage frequency
600rpm 500rpm 400rpm
500 7.87 7.52 5.88
710 23.86 13.47 8.75
1000 44.73 22.75 14.34
1200 13.83 28.24 24.21
1450 2.13 22.06 40.53
1700 - 2.57 5.41
Figure 22 Effect of intensity of agitation on size distribution of Spherical
agglomerates.
0
5
10
15
20
25
30
35
40
45
50
0 200 400 600 800 1000 1200 1400 1600 1800
% F
req
uen
cy
Particle diameter (µm
600 rmp 500 rmp 400 rmp
RESULT AND DISCUSSION
118
5.1.5 Optimum stirring time or residence time
Optimum stirring time or residence time for which the spherical crystals
remain suspended in the crystallization medium was found to be 25±5 min. Spherical
crystals did not form when the stirring time was below 25 min. due to incomplete
crystallization/agglomeration. Longer residence time resulted in breakdown of
spherical crystals or may be surface could have rough due to secondary crystallization
164,168.
5.1.6 Influence of the temperature
The temperature of agglomerating solvents was found to have pronounced
effect on the process of spherical crystallization. Agglomeration and hence formation
of crystal agglomerates could not occur when the process was carried out at 2±5oC. It
could be due to reduced solubility of drug in crystallization solvents at such a lower
temperature, which did not affect efficient wetting of drug particles and hence
reduced agglomeration. When the temperature of the process was increased to 45±5oC
very large agglomerates were produced and the amount of the recovery of the drug
was reduced. It could be due to increased solubility of the drug at this higher
temperature. Optimum agglomeration was obtained when the process was carried out
at 25±5oC
164, 165, 168.
5.1.7 Influence of mode of addition of bridging liquid
Uniform distribution of bridging liquid was obtained when it was added drop
wise with continuous stirring of agitator. This process resulted in the formation of
spherical crystals due to efficient crystallization. Pouring of whole amount of bridging
liquid in crystallization vessel at a time produced spherical crystals of irregular
RESULT AND DISCUSSION
119
geometry. It is due to localization of bridging liquid and hence its unavailability for
efficient crystallization164, 165, 168
.
Table 12 Effect of variables on formulation of spherical crystals of Mefenamic
acid
Parameter
Variables
Observation
Bridging liquid
IPA
Chloroform
Toluene
Agglomeration
Agglomeration
No agglomeration
Percentage (%) of
bridging liquid Isopropyl
acetate
9%
10%
11%
No agglomeration
Agglomeration
No agglomeration
Agitation speed
300±25
400±25
500±25
600±25
700±25
Clumps
Spherical & large
Spherical
Spherical & small
Irregular shape & small
Agitation time
˂45 min
45 min
Incomplete agglomerates
Spherical agglomerates
Temperature
5±20 C
20±20C
45±20 C
Large Spherical Agglomerates
Spherical agglomeration
no agglomerates
Mode of addition of
bridging liquid
Whole at a time
Drop wise
Crystals of irregular geometry
Spherical agglomerates
RESULT AND DISCUSSION
120
5.2 Recrystallization of Mefenamic acid:
Recrystallization of Mefenamic acid was done to find out the changes in crystal
lattice, being induced by solvents that can influence the some physicochemical
properties and secondary characteristics of the drug substance. Hence the mechanical,
micromeritic and dissolution properties of prepared crystals were compared with
commercial sample of mefenamic acid168
.
5.3 Crystallization by spray drying technique:
The spray technology being applied to the manufacture of mefenamic acid crystals
using the same solvent system as used in spherical crystallization. In this technique,
energy is applied to the droplet, forcing evaporation of the medium resulting in both
energy and mass transfer through the droplet. Spray drying is the most widely used
industrial process involving crystals formation and drying. It is highly suited for the
continuous production of dry crystals of mefenamic acid169, 173
.
In spray drying method, three important parameters were considered, 1. Inlet
temperature 2. Feed rate/ flow rate of drug solids in the solution and 3. Atomization
pressure rate/ atomization speed rotation. All three parameter have an impact on the
yield of spray dried crystals. Process variables are summarized in Table 13.
RESULT AND DISCUSSION
121
Table 13 Influence of process variables on crystals of Spray dried crystals of
Mefenamic acid
Parameter Variables Observation
Inlet Temperature
100±20 C
110±20 C
120±20 C
Less % yield
Complete dry crystals with high yield
Less % yield
Feed rate/ flow rate of
drug solids in the solution
10%
12%
15%
Less % yield, Irregular shape crystals
Good % yield, Spherical shape crystals
Sticking to wall, Less % yield, Irregular
shape crystals
Atomization pressure
rate/ Atomization speed
rotation
0.5 kg/cm2
1 kg/cm2
2 kg/cm2
Incomplete crystals, Less % yield
Complete spherical crystals, good %
yield
Sticking to wall, Irregular shape
crystals, Less % yield
RESULT AND DISCUSSION
122
5.4 Crystallization by freeze drying techniques:
While optimizing the Freeze-drying process, three parameter were considered;
Time of freezing, Time of primary drying, Time of secondary drying. Process
variables are summarized in Table 14.
Table 14 Influence of process variables on crystals of freeze dried crystals of
Mefenamic acid
Sl.
No. Parameter Variables Observation
1. Time of freezing
-30 0C for 12 hr
-40 0C for 24 hr
-50 0C for 36 h
Incomplete solidification
Complete solidification
Complete f solidification but over
frozen product
2. Time of primary
drying
48 hr
72 hr
96 hr
Incomplete dried sample
Complete dried sample with good
properties
Complete dried sample with higher
water content
3. Time of
secondary drying
18 hr
24 hr
36 hr
Complete dried sample with higher
water content
Complete dried sample with good
properties
Complete dried sample with poor
properties
Optimized parameters of the Freeze-drying process have been selected as
follow: Time of freezing: -40 0C for 24 hr, Time of primary drying: 72 hr, Time of
secondary drying: 24 hr170, 197, 198
.
RESULT AND DISCUSSION
123
5.5 Super cooling crystallization (solidification technique):
In this method Mefenamic acid was heated until it is melted. The melt is solidified
rapidly in an ice bath under rigorous stirring, pulverized, and then sieved. Rapid
congealing is desirable because it results in super saturation of drug. Crystals from
this process can be obtained in pellet form without the necessity of a grinding step
that may alter crystalline modification171, 172, 173
.
The super-cooling process could be considered as a self-seed process in which
the nucleus occurs. The super-cooling process significantly reduce the nucleation time
and increase the overall crystallization rate which result in a higher crystallinity than
that in other crystallization techniques.
5.6 Characterization of Mefenamic acid spherical agglomerates
5.6.1 Percentage yield:
The yield was in the range of 78-95%. The yield of crystals prepared by different
crystallization techniques is presented Table 15:
Table 15 Yield of different crystals of mefenamic acid by different crystallization
techniques
Sl. No. Different crystals % yields*
1 Spherical crystals 78± 2
2 Spray dried crystals 79± 2
3 Freeze dried crystals 86± 2
4 Super cooling crystals 78± 2
5 Recrystallized crystals 95± 2
*Mean SD± n=5
Yield was less for the drug crystallized by spherical crystallization, spray drying,
freeze drying and super cooling. some loss, resulting from transferring solids from
one container to another and leaving a little material behind. Also, because of the
finite solubility of the solid in the crystallization solvent, even at low temperatures,
RESULT AND DISCUSSION
124
any unnecessary prolonged contact with crystallization solvent, especially if the
solvent is not ice-cold will result in loss of product. Some of the product would have
redissolved and lost. During spherical crystallization, particles were sticking to the
wall of container and stirrer, while in case of spray drying, inability of the cyclone
separator to trap particles smaller than 2 μm and adherence of these particles to the
inner wall of the spray drier174, 175
. For super cooling crystals, drugs particles were
sticking to the wall of the container which led to the loss in yield.
5.6.2 Drug content
Drug content in various crystals prepared is presented Table 16.
Table 16 Drug content of different crystals of mefenamic acid
Sl. No. Different crystals Drug content*
1 Spherical crystals 98.43± 0.3
2 Spray dried crystals 99.12± 0.2
3 Freeze dried crystals 99.89± 0.1
4 Super cooling crystals 13.56 ± 2.5
5 Recrystallized crystals 98.67± 0.2
*Mean SD± n=5
5.6.3 Melting point:
Melting point of Mefenamic acid was determined by capillary filled method. The
Melting point of crystals prepared by different crystallization techniques are shown in
Table 17.
Table 17: Melting point of different crystals of Mefenamic acid
Sl. No. Mefenamic acid
crystals
Literature melting point
(20)
Observed melting
point
1 Commercial sample
229-2310C
231±0.5
2 Spherical crystals 230±0.5
3 Spray dried crystals 229±0.5
4 Freeze dried crystals 229±0.5
5 Super cooling crystals 233±0.5
6 Recrystallized crystals 231±0.5
RESULT AND DISCUSSION
125
5.6.4 Water content
This test is important in cases where the drug, like mefenamic acid, is known to be
hygroscopic or degraded by moisture or when the drug substance is known to be a
stoichiometric hydrate. A Loss on drying due to water evaporation may be considered
adequate in some cases, ICH guidelines specify that the water content shall essentially
be determined by Karl Fischer titration176
. The water content in crystals prepared by
different crystallization techniques are shown in Table 18.
Table 18: Water contain of different crystals of mefenamic acid
Sl. No. Mefenamic acid crystals Water content (% w/w)
1 Commercial sample 4.56
2 Spherical crystals 4.34
3 Spray dried crystals 4.47
4 Freeze dried crystals 5.32
5 Super cooling crystals 4.71
6 Recrystallized crystals 4.92
Not More than: 5 % w/w (39).
5.6.5 Residual solvent content:
THF and isopropyl acetate are considered as toxic organic solvent based on its
concentration, however these little detriment to human body. According to the
International Conference on Harmonization (ICH) guidelines for residual solvents, the
solvents are classified into three different categories: Class I, II and III solvents, Class
I solvents are extremely toxic, class II are moderately toxic and class III are low toxic.
Although not listed in the ICH guidelines, THF and isopropyl acetate are fall in the
category of a class II and class III respectively. Therefore, the low level of THF and
isopropyl acetate in the crystals should not be harmful to both animal and human177,
178.
Residual solvents in prepared crystals were determined (THF and isopropyl acetate)
by Gas chromatography presented in Table 19.
RESULT AND DISCUSSION
126
Table 19 Residual solvent in different crystals of Mefenamic acid
Sl. No. Solvents ICH Class and
limit (ppm)
Mefenamic acid
crystals
Residual solvent
(ppm)
1 Isopropyl
acetate
Class 3
5000 ppm
Spherical crystals
Spray dried crystals
Freeze dried crystals
Recrystallized crystals
07
10
11
09
2 Tetrahydrofuran
Class 2
720 ppm
Spherical crystals
Spray dried crystals
Freeze dried crystals
Recrystallized crystals
14
15
18
15
5.6.6 Differential scanning calorimetry (DSC)
DSC studies were performed for different crystals of Mefenamic acid. The
DSC thermograms of Mefenamic acid commercial sample, spherical agglomerates,
spray dried crystals, freeze dried crystals, super cooling crystals and recrystallized
sample are presented in Figure 23. Onset of melt (T0), melting point (Tm) and
completion of melt (Tc) is present in the Table 20. The DSC thermograms show a
sharp endothermic peak for all the Mefenamic acid crystals except super cooling
crystals. This one step melt might be due to only one crystal form (Triclinic) of the
Mefenamic acid formed during the crystallization process. Thus indicating that
Mefenamic acid did not undergo any crystal modification. The temperatures of the
endothermic peaks of all the Mefenamic acid crystals were in the range of 2290C to
2330C. Melting point of the mefenamic acid was 231.32 with enthalpy 184.3 J/g
183.
Melting points show slight variation as the solvent might have affected the nature of
the crystals, the melting points determined by open capillaries are agreement with
DSC studies. The thermogram of super cooling crystals showed at three peaks at
2230C, 241
0C and 272
0C respectively with different enthalpy indicating change in the
polymorphic form of the mefenamic acid.
RESULT AND DISCUSSION
127
The prepared crystals have decreased in crystallinity & Heat of fusion than
commercial sample of mefenamic acid. The crystallinity was in the following order
FD>SD>SA>RS>CS. The decreased crystallinity and Heat of fusion of the different
crystals of Mefenamic acid may be one of the reasons for enhanced solubility and
dissolution of the prepared crystals196
.
The melting point and enthalpy for prepared crystals of mefenamic acid are presented
in Table 20.
Table 20 DSC data of different Mefenamic acid crystals
Crystals T0 Tm Tc Melting range Heat of fusion*
J/gm
Decreased %
crystallinity
Commercia
l sample 225.14 231.32 234.73 9.59 184.32 J/g 100
Spherical
crystals 226.23 230.17 234.63 8.40 167.64 J/g 90.95
Spray dried
crystals 225.02 229.71 233.42 8.40 159.19 J/g 86.36
Freeze
dried
crystals
226.52 229.56 233.58 7.06 145.53 J/g 78.95
Recrystalliz
ed sample 227.43 233.01 234.65 7.22 177.41 J/g 96.25
Super
cooling ---
223
241
272
--- --- --- ---
T0-Onset of melt, Tm -Melting point, Tc - Completion of the melt
RESULT AND DISCUSSION
128
Figure 23 DSC thermograms of different Mefenamic acid crystals
SC- Super cooling crystals, CS-Commercial sample, RS- Recrystallized crystals, SA-
Spherical crystals, SD- Spray dried crystals, FD- Freeze dried crystals.
RESULT AND DISCUSSION
129
5.6.7 FT-IR Spectroscopy:
FTIR studies were performed in order to aid the evaluation of any possible
chemical interaction between the drug mefenamic acid and solvents during
crystallization184, 185
. Moreover, molecular level characterization of crystals can also
be obtained by performing FTIR studies. The FTIR spectrum for commercial sample,
spherical crystals, spray dried crystals, freeze dried crystals, recrystallized crystals
and super cooling crystals are shown in Figure 24 and Table 21.
The FTIR spectrum for mefenamic acid shows a weak peak at 3400 cm ⁻1 due
to the presence of a secondary amine. The broad band in the range of 3200-2900 cm
⁻1 is due to the presence of –OH [1, 2]. The same also represents the intra- and
intermolecular hydrogen bonding due to the –OH groups and also overlaps with the (–
CH3) group. The peak at 1650- 1750 cm⁻1 is due to the presence of a C=O group.
The presence of a peak at 1000 cm ⁻1 indicates the presence of a phenyl group186, 187
.
All the prepared Mefenamic acid crystals have exhibited general characteristic
peaks (Table 21). Specific changes in IR spectra are not very clear, could be due to
variations in the resonance structure, rotation of a part of a molecule or certain bonds.
RESULT AND DISCUSSION
130
Table 21 FT-IR data of Mefenamic acid crystals
Assignment
(Group)
Frequency
CS(in cm-1
)
Frequency
RS(in cm-1
)
Frequency
SA(in cm –1
)
Frequency
SD (in cm–1
)
Frequency
FD(in cm–1
)
Frequency
SC (in cm–1
)
secondary amine 3400 3400 3400 3400 3400 3350-3400
–OH
group
3200-2900 3200-2900 3200-2900 3200-2900 3200-2900 3100-2800
C=O
group
1650- 1750 1650- 1750 1650- 1750 1650- 1750 1650- 1750 1600- 1700
phenyl group 1000 1000 1000 1000 1000 1000
SC- Super cooling crystals, CS-Commercial sample, RS- Recrystallized crystals, SA- Spherical crystals, SD- Spray dried crystals, FD- Freeze
dried crystals.
RESULT AND DISCUSSION
131
Figure 24 FT-IR spectra of Mefenamic acid crystals SC- Super cooling crystals, CS-Commercial sample, RS- Recrystallized crystals, SA-
Spherical crystals, SD- Spray dried crystals, FD- Freeze dried crystals.
RESULT AND DISCUSSION
132
5.6.8 X-ray analysis:
The characteristic peak of the Mefenamic acid appeared in the 2θ range of 0–
600. All the prepared crystals of mefenamic acid showed similar peak positions (2θ) in
X-ray diffraction, formation of different polymorphs of Mefenamic acid was ruled
out, except super cooling crystals. However, the relative intensities of XRD peaks
were modified (Figure 25). This was attributed to the markedly different crystal habits
of the crystals173, 182
. Therefore the relative abundance of the planes exposed to the X-
ray source would have been altered, producing the variations in the relative intensities
of the peak or this may be due to differences in crystal size189
.
The commercial sample of mefenamic acid exhibits its characteristic
diffraction peaks at various diffraction angles indicating the presence of crystallinity.
The X-ray diffraction of the SC of drug showed the more peaks than other prepared
crystals and commercial sample this could be due to the degradation of drug by
heating or it could be variations in the resonance structure, rotation of a part of a
molecule or certain bonds. Alteration could be due to minor distortion of bond angles
and this lead to chances of conversion of mefenamic acid to another form in super
cooling crystals, the intensity of major peaks were lower than commercial sample due
to the differences in crystal sizes. The X-ray diffraction pattern of the all prepared
crystals showed that peak intensity was much lower than the commercial sample.
The crystalline peaks associated with prepared Mefenamic acid crystals were
of low intensity compare to commercial sample of Mefenamic acid. These findings
suggested that Mefenamic acid crystal habit were modified, to show improve
micromeritic properties by different techniques, these finding are in agreement with
previous study on Different NSAIDs drugs199
.
RESULT AND DISCUSSION
133
The Data obtained from X-ray powder diffraction studies are reported in
Tables 22, 23, 24, 25, 26 and 27. The cell parameters a, b, c, α, β, γ and the cell
volume for all the crystals of Mefenamic acid reported in Table 28.
Table 22: XRD data for commercial Mefenamic acid crystals
Peak
number
h k l 2 Ө
(Observed)
2 Ө
(Calculated)
Diffraction
1 0 0 1 .005766 .005762 .000005
2 1 0 2 .010459 .010544 -.000084
3 1 0 1 .012156 .012141 .000015
4 0 1 1 .015708 .015718 -.000009
5 1 -1 1 .019058 .019028 .000029
6 0 -1 2 .021315 .021240 .000075
7 0 2 0 .024176 .024140 .000036
8 1 1 0 .026675 .026523 .000153
9 1 2 1 .034568 .034576 -.000008
10 -1 -2 1 .056575 .056656 -.000081
11 -1 1 1 .064607 .064634 -.000027
12 1 -3 1 .069008 .069013 -.000004
13 1 -2 1 .076903 .065634 -.000017
14 1 2 4 .087887 .087880 .000007
Lattice Parameters
A 7.5631
B 9.0832
C 10.472
98.942
Β 92.521
57.965
Unit cell Volume 602.361
Space group Triclinic
h, k, l – Miller indices.
a, b, c – three sides of cell expressed in A0.
, β, - three angles of the cell expressed in degrees.
RESULT AND DISCUSSION
134
Table 23: XRD data for Mefenamic acid Recrystallized sample
Peak
number h k l
2 Ө
(Observed) 2 Ө (Calculated) Diffraction
1 0 -1 1 .005988
2 0 0 1 .006142 .006108 .000035
3 1 0 0 .008167 .008170 -.000003
4 1 0 1 .011292 .011272 .000020
5 1 1 0 .013231 .013248 -.000018
6 0 -1 2 .016464 .016476 -.000012
7 -1 1 0 .018395 .018524 -.000129
8 0 1 1 .021542 .021659 -.000117
9 1 1 1 .024176 .024185 -.000009
10 -1 -2 1 .027212 .027202 .000011
11 -1 1 1 .035563 .035473 .000090
12 -1 0 2 .038596 .038612 -.000016
13 -1 2 0 .044334 .044310 .000024
14 1 1 2 .047364 .047338 .000026
15 1 2 1 .027212 .052531 -.000009
16 0 2 1 .021542 .052643 -.000117
17 2 2 0 .052923 .052993 -.000070
18 -1 -3 2 .027212 .053132 -.000016
19 1 -3 2 .056939 .056936 .000003
20 -2 1 1 .035563 .065626 .000090
21 0 -2 4 .065857 .065905 -.000048
22 0 3 0 .047364 .069445 .000026
23 2 0 3 .047364 .069614 -.000009
24 1 3 0 .069807 .069702 .000105
RESULT AND DISCUSSION
135
Lattice Parameters
A 10.443
B 11.743
C 13.053
71.492
Β 99.071
112.02
Unit Cell Volume 1406.87
Space group Triclinic
h, k, l – Miller indices.
a, b, c – three sides of cell expressed in A0.
, β, - three angles of the cell expressed in degrees.
RESULT AND DISCUSSION
136
Table 24: XRD data for Mefenamic acid spherical agglomerates
Peak
number
h k l
2 Ө
(Observed)
2 Ө
(Calculated)
Diffraction
1 0 1 1 .005886 .005861 .000025
2 0 0 1 .006771 .006770 .000001
3 1 0 1 .016344 .006854 .000009
4 0 1 0 .007756 .007744 .000012
5 1 0 0 .010922 .010914 .000009
6 1 0 2 .016344 .016335 .000009
7 1 2 2 .018185 .018144 .000041
8 .021064 .010914 .000009
9 1 2 3 .000046 .023858 .000046
10 1 1 3 .023909 .023863 .000046
11 1 2 1 .025921 .025971 -.000049
12 2 2 2 .034663 .034673 -.000009
13 1 -1 2 .038596 .038663 -.000067
14 2 0 0 .043690 .043655 .000035
15 0 -1 2 .052222 .052132 .000090
16 -1 0 2 .059596 .059653 -.000058
17 2 -2 1 .066161 .066158 .000003
18 0 3 0 .069807 .069699 .000108
19 0 2 4 .059596 .070071 .000090
RESULT AND DISCUSSION
137
Lattice Parameters
A 10.437
B 12.995
C 12.346
85.368
β 91.947
115.84
Unit cell Volume 1501.33
Space group Triclinic
h, k, l – Miller indices.
a, b, c – three sides of cell expressed in A0.
, β, - three angles of the cell expressed in degrees.
RESULT AND DISCUSSION
138
Table 25: XRD data for Mefenamic acid spray dried crystals
Peak
number
h k l
2 Ө
(Observed)
2
Ө(Calculated)
iffraction
1 0 1 0 .003136 .003137 -.000001
2 1 2 0 .014726 .014729 -.000004
3 3 0 0 .019683 .019632 .000051
4 0 1 1 .030965 .031004 -.000039
5 .034695
6 3 3 0 .048033 .047865 .000168
7 1 4 0 .052222 .052374 -.000152
8 1 3 1 .058444 .058281 .000163
9 4 2 1 .075192 .075316 -.000124
Lattice Parameters
A 7.8356
B 10.582
C 17.068
107.256
β 39.080
96.540
Unit cell Volume 836.031
Space group Triclinic
h, k, l – Miller indices.
a, b, c – three sides of cell expressed in A0.
, β, - three angles of the cell expressed in degrees.
RESULT AND DISCUSSION
139
Table 26: XRD data for Mefenamic acid Freeze dried crystals
Peak
number
h k l 2 Ө
(Observed)
2 Ө
(Calculated)
Diffraction
1 0 1 0 .003242 .003241 .000001
2 0 0 1 .014937 .015029 -.000092
3 3 1 0 .017355 .017420 -.000066
4 2 2 0 .019273 .019267 .000006
5 1 3 0 .030693 .030747 -.000054
6 2 2 1 .034377 .034296 .000080
7 .048033 .019267 .000006
8 0 4 0 .051873 .051861 .000012
9 5 1 1 .057669 .057657 .000012
10 1 2 2 .074778 .074657 .000121
11 6 1 1 -.000066 .074987 .019273
Lattice Parameters
A 10.774
B 11.696
C 10.204
93.462
β 109.49
117.20
Unit cell Volume 1042.90
Space group Triclinic
h, k, l – Miller indices.
a, b, c – three sides of cell expressed in A0.
, β, - three angles of the cell expressed in degrees.
RESULT AND DISCUSSION
140
Table 27: XRD data for Mefenamic acid Super cooling crystals
Peak number h k l 2 Ө Observed) 2 Ө (Calculated) Diffraction
1 1 1 0 .008597 .008551 .000046
2 0 0 1 .011127 .011115 .000012
3 0 2 0 .015514 .015481 .000033
4 -2 0 1 .019951 .019961 -.000010
5 1 1 1 .024661 .024604 .000057
6 0 1 0 .030965 .015481 .000033
7 1 2 1 .036147 .036215 -.000068
8 1 3 0 .039509 .039512 -.000003
9 -2 0 2 .043369 .043430 -.000061
10 -2 1 2 .047364 .047300 .000063
11 3 1 1 .030965 .071925 .000032
12 -3 2 2 .072273 .072438 -.000165
Lattice Parameters
A 8.7353
B 10.695
C 12.132
123.75
β 81.623
86.806
Unit cell Volume 919.452
Space group Triclinic
h, k, l – Miller indices.
a, b, c – three sides of cell expressed in A0.
, β, - three angles of the cell expressed in degrees.
RESULT AND DISCUSSION
141
Table 28: Different cell parameters obtained for Mefenamic acid crystals From
XRD data
Different crystals of
Mefenamic acid
A B C α β γ Unit cell volume
Commercial sample 16.49 13.75 4.61 90 90 90 1046.691
Recrystallized
crystals
19.40 13.53 6.28 90 90 90 1649.86
Super cooling
crystals
11.98 12.38 7.77 90 110 90 1084.10
Spherical crystals 15.53 13.76 6.98 90 90 90 1156.16
Spray dried crystals 15.49 13.70 5.54 90 90 90 1177.77
Freeze dried crystals 15.50 14.01 9.21 90 90 90 2001.88
RESULT AND DISCUSSION
142
Figure 25: X-ray diffraction spectra of Mefenamic acid
SC- Super cooling crystals, CS-Commercial sample, RS- Recrystallized crystals, SA-
Spherical crystals, SD- Spray dried crystals, FD- Freeze dried crystals.
RESULT AND DISCUSSION
143
5.6.9 Surface topography by scanning electron microscopy (SEM):
Crystals of commercial sample are of the smallest size (5-10 µm) and they
have irregular shapes. Recrystallization leads to crystals with intermediate size (7-18
µm) which had rod or needle like shapes. The spherical agglomerates had a rough
surface covered with numerous rod shaped crystals (1221-1624 µm). The shape of
prepared spray dried crystals is uniform and was spherical in shape with small size (4-
12 µm). The freeze dried crystals had a smooth surface with very small in size
(average particle size 257 nm) (Figure 29) and in case of super cooling, crystals were
irregular shape, having rough surface with yellowish colour (21-37 µm).
The photomicrograph of Mefenamic acid commercial sample, recrystallized
sample, spherical crystals, spray dried crystals, freeze dried crystals and super cooling
crystals are shown in the Figures 26, 27 & 28.
RESULT AND DISCUSSION
144
Figure26: SEM of Mefenamic acid commercial and recrystallized crystals
Figure 27: SEM of Mefenamic acid Spherical and spray dried crystals
RESULT AND DISCUSSION
145
Figure 28 SEM of Mefenamic acid Freeze dried and super cooling crystals
Figure 29: Particle size distribution graph of Mefenamic acid Freeze dried
crystals by Malvern analyzer
RESULT AND DISCUSSION
146
5.7 Evaluation of prepared crystals:
5.7.1 Micromeritic properties:
The differences in the bulk densities may be related to their markedly different
crystal habits, leading to different contact points, frictional and cohesive forces
between the crystals. Spherical crystals, spray dried crystals, freeze dried crystals,
recrystallized and super cooling crystals exhibited higher packing ability than
commercial sample. It is due to lower surface area and wide particle size distribution.
The smaller crystals might have settled in voids between larger particles.
Three measures of flowability were utilized to analyze the flow properties (a)
flow rate (b) angle of repose and (c) compression index. Granules or powder must
adequately flow from holding container during pharmaceutical manufacture. Flow
rate measurement allows quick estimation of flow properties. It is the length of time
required for the flow of unit mass of substance through the orifice. Angle of repose is
able to provide gross measurements of the flowability of crystals. Most free flowing
materials have angle of repose less than 300. Powders with angles greater than 50
0
have flow problems. Commercial sample exhibited higher angle of repose than
prepared crystals that could be due to the irregular shape and small size of crystals,
which put hurdles in the uniform flow of crystals from funnel. The higher flowability
of the prepared crystals is due to perfect sphericity and particles size of the crystals.
Carr’s index & Hausner’s ratio were also used to assess the flow and compressibility
properties of the crystals. The compressibility index is a simple and fast method for
estimating flow of powder. Carr’s index showed the relationship between the
compressibility index and flowability. Different micromeritic properties of
Mefenamic acid commercial sample and prepared crystals are given in Table-29.
RESULT AND DISCUSSION
147
Angle of repose properties
less than 20 excellent flow
between 20-30 good flow
between 30-34 Pass flow
greater than 40 poor flow
The carr’s index, Hausner’s ratio of commercial sample is 36.13± 0.11% &
1.56 ± 0.21 respectively (n = 3), indicating extremely poor flow properties. Hausner
ratio was related to interparticle friction. Powders with Hausner’s ratio less than 1.25
indicates good flow (= 20% Carr) due to low inter-particle friction. Value greater than
1.5 indicates poor flow (= 33% Carr), such powder are more cohesive, less free-
flowing powders such as flakes. Flow rates are in agreement with morphology and
bulk density data that spherical crystals with low bulk density exhibits better flow
properties.
Relationship between powder flowability and % compressibility
Flow description % compressibility
Excellent flow 5 – 15
Good 16 – 18
Fair 19 – 21
Poor 22 – 35
Very poor 36 – 40
Extremely poor > 40
RESULT AND DISCUSSION
148
Table 29 Micromeritic and mechanical properties of Mefenamic acid commercial sample and prepared crystals
Properties Commercial
sample
Recrystallized
crystals
Spherical
crystals
Spray dried
crystals
Freeze dried
crystals
Super cooling
crystals
Particle size (m) 5-10 7-18 1221-1624 4-12 257 nm 21-37
Flow rate (gm/Sec) No flow No flow 8.37 4.53 3.93 9.18
Angle of repose (Ө)* 32.17± 1.28 30.96± 2.2 27.63± 1.46 26.23± 1.37 19.26 ± 2.21 25.84± 1.43
Tapped density
(gm/ml) * 0.487± 0.13 0.486± 0.11 0.419± 0.03 0.375± 0.19 0.365± 0.21 0.401± 0.18
Bulk density
(gm/ ml) * 0.311± 0.16 0.343± 0.11 0.395± 0.13 0.358± 0.14 0.367± 0.20 0.338± 0.2
Carr’s index (%)
(compressibility
index)*
36.13± 0.11 29.42± 0.02 5.72± 0.01 4.53± 0.2 1.89± 0.31 15.82± 0.12
Hausner’s Ratio* 1.56± 0.21 1.41± 0.20 1.06± 0.01 1.04± 0.11 1.01± 0.30 1.18± 0.14
Friability (%)* - - 0.654±0.12 - - -
Tensile strength*
(Kg/cm2) 6.56 ±0.11 8.18 ±0.04 5.24 ±0.02 7.39 ±0.03 8.41 ±0.01 10.53 ±0.03
*Standard deviation, n = 3.
RESULT AND DISCUSSION
149
5.7.2 Mechanical properties:
a. Tensile strength
Prepared crystals exhibited superior compressibility characteristics compared
to commercial sample except super cooling crystals (Figure 30 & Table-29). The FD
crystals showed maximum tensile strength and super cooling crystals shows minimum
tensile strength than the commercial samples of mefenamic acid. This could be due to
the fact that during the process of compression fresh surfaces are formed by
fracturing. Surface freshly formed by fracture enhanced the plastic inter particle
bonding, resulting in a lower compression force required for compressing the
prepared crystals under plastic deformation164, 168
.
Figure 30: Tensile strength of commercial crystals of mefenamic acid and
prepared crystals
b. Crushing strength
The crushing strength of the mefenamic acid spherical crystals was in the
range of 95 - 105 gm and was unaffected by the process variables164, 166, 168
.
0
2
4
6
8
10
12
0 1000 2000 3000 4000 5000 6000
Ten
sile
str
ength
Kg/c
m2
Compaction Pressure Kg/cm2
CS RS SC SA SD FD
RESULT AND DISCUSSION
150
Mefenamic acid: the effect of compression on compact strength is shown in Figure
30. Compact strength were in the order super cooling > freeze dried > recrystallized >
spray dried > commercial sample > spherical crystals. It can be intended that the
increases in compression force increased the compact hardness.
A high slope of a linear relation of compression force verses hardness suggests the
potential problems of capping. Since a small change in pressure could cause
significant change in compact strength.
A linear relationship between tensile strength and compression pressure was observed
for all crystallized sample of mefenamic acid.
c. Heckle analysis
Heckel plot is density-compression force relationship to determine the plastic
behavior of materials.
The slope of the Heckel plot (k) is indicative of the plastic behavior of the
material. A larger value for the slope is related to a greater amount of plasticity in the
material. This indicates that prepared crystals exhibited a higher degree of packing in
the die as a result of die filling than commercial sample. The differences between the
powders are not large, and this is expected because the materials are similar.
Considering slopes and their Standard mean Error, it is difficult to conclude whether
there are real differences between the plastic properties of different types of crystals.
A good rule of thumb is that, if Confidence intervals are not overlapping, then the
differences in plasticity are significant. When two confidence interval are greatly
overlapping, the plasticity of the materials is considered the same (193
). Generally, the
plasticity decreases in the following order: FD >SD >SA >SC >RS >commercial
RESULT AND DISCUSSION
151
sample of mefenamic acid. Constants of Heckel plot are given in Table 30 and Figure
31.
Table 30: Heckel parameters and elastic recovery of piroxicam prepared crystals
Parameters
Commercial
sample
Recrystallized
crystals
Spherical
crystals
Spray dried
crystals
Freeze dried
crystals
Super cooling
crystals
PY 64.4 ±0.21 66.01±0.31 67.21±0.13 67.54±0.37 68.2 ± 0.72 57.01±0.23
D0 0.554 ±0.07 0.484±0.02 0.467±0.01 0.458±0.13 0.442 ±0.03 0.513±0.25
DA 0.639 ±0.03 0.593±0.02 0.519±0.17 0.523±0.03 0.501 ±0.03 0.596±0.01
D’B 0.152 ±0.02 0.182±0.01 0.195±0.02 0.217±0.02 0.237 ±0.01 0.176±0.03
Elastic
recovery (%)
3.83 ± 0.12 4.37±013 4.89±0.11 5.19±003 5.30 ± 0.01 418.±013
Slope 0.00030 0.00035 0.0031 0.0038 0.0042 0.00072
RESULT AND DISCUSSION
152
Figure 31: Heckle profiles of different crystals of Mefenamic acid
The powder column formed by FD & SD was more densely packed than that
formed by CS in the initial stages of rearrangement as indicated by their Do values
(Table 22). The particles of FD & SD powder are larger, highly composed of
spherical- to oval-aggregates with few primary particles which are irregular compared
to those of CS. which are small and irregular primary particle. These features of the
latter could result to formation of bridges and arches, which could in turn prevent
close packing of the particles in the bulk state. Thus it seems that the values for the
relative density or the packing fraction in the bulk state are strongly dependent on the
particle size and shape.
0
0.5
1
1.5
2
2.5
0 1000 2000 3000 4000 5000 6000
In [
1/(
1.-
D)]
Applied pressure (kg)
CS RS SC SA SD FD
RESULT AND DISCUSSION
153
The low Da values for FD & SD are an indication that it opposes the
densification process less strongly than commercial crystal. Da describes the share of
densification due to contact area between particles; therefore, the low Da value of FD
is an indication of low contact area between the particles. However, it has been
reported that the resultant contact area in a powder bed is dependent upon the
interplay of several variables ranging from specific surface area (sequel to particle
size), particle size distribution, particle shape to surface properties of the powders
such as hardness of the surface.
Db, which describes the phase of rearrangement of particles, the extent of
which depends on the theoretical point of densification at which deformation begins,
was also determined for the cellulose powders; FD has a higher value than CS. FD
crystals were more resistant to movement once the initial phase of packing (as a result
of die filling) had been completed. This could be attributed to the high cohesive forces
likely present as a result of its amorphous nature, being an Freeze crystals.
The mean yield pressure, Py, values were calculated from the slope of the
linear line constructed over the compression pressure range (Table 22). Py which is
inversely related to the ability of the material to deform plastically under pressure was
found to be higher for FD. The results therefore indicate that FD underwent plastic
deformation more easily and rapidly than CS. This also confirms that FD is somewhat
resistant to deformation.
RESULT AND DISCUSSION
154
5.7.3 Solubility studies:
All the prepared crystals of mefenamic acid showed increased solubility than
the commercial sample in water and in pH 7.4 Phosphate buffer, The freeze dried
crystals showed highest solubility than commercial sample of mefenamic acid both in
water as well as in pH 7.4 phosphate buffer. The solubility of super cooling particle
showed least solubility than commercial sample both in water as well as in pH 7.4
phosphate buffer (Table 31)164, 166, 168
.
Increased solubility may be due to the increased wettability of the prepared
crystals or reduction in particle size & change surface properties of the prepared
crystals173, 200
.
In the case super cooling crystals, the poor solubility could be due to the
degradation of drug during heating or it could be due to variations in the resonance
structure, rotation of a part of a molecule or certain bonds. Alteration could be due to
minor distortion of bond angles171, 172, 173, 182
.
The solubility of prepared crystals of mefenamic acid in water and in pH 7.4
Phosphate buffer presented in Table 31;
RESULT AND DISCUSSION
155
Table 31: Solubility of different crystals of Mefenamic acid
Mefenamic acid Samples In Water(mg/ml)
(±SD)
In pH 7.4(mg/ml)
( ±SD)
Commercial sample 0.0083±0.01 0.0737±0.01
Recrystallized crystal 0.0094±0.02 0.0778±0.02
Super cooling crystals 0.0037±0.02 0.0239±0.01
Spherical crystals 0.0437±0.02 0.1602±0.01
Spray dried crystals 0.0526±0.01 3.7645±0.02
Freeze dried crystals 0.1237±0.02 4.6520±0.02
*Mean SD± n=3
RESULT AND DISCUSSION
156
5.7.4 Dissolution behaviour of crystals:
Dissolution studies were conducted using USP dissolution apparatus XXIV-
Type II as per the monograph for mefenamic acid tablets. But in this study crystals
were directly added to the dissolution medium. Encapsulating the drug was avoided,
as dissolution of shell will add one more parameter to the result. Amount of
mefenamic acid dissolved – time data are reported in the Table 32 for commercial
sample and for prepared crystals, Dissolution profile of the mefenamic acid
commercial sample, recrystallized sample and spherical crystals is shown in the
Figure 32.
Percentage drug dissolved in 10 min is rapid in all the samples. The
commercial sample showed a plateau region in its dissolution profile. This could be
attributed to the super saturation of unstable form of drug with its stable form. Drug in
contact with solvent mixtures the transformation of rate depends on the mobility of
the molecule in the solid, the type of structural change that take place and
environment factor. The larger the difference between the packaging arrangements of
two forms the slower the rate of conversion from unstable form to stable form.
The dissolution profiles of mefenamic acid exhibited improved dissolution
behaviour for prepared crystals than commercial sample, except super cooling
crystals. The reason for this faster dissolution could be linked to the better wettability
of the prepared crystals or reduction in particle size of the prepared crystals. The
amount of drug dissolved in 60 min greatly varied for all prepared crystals164-168
.
Difference in the dissolution and solubility profile of the modified Mefenamic
acid may also be due to the difference in crystal structure. Solvents with varying
polarities and desolvation process affect crystal formation, leading to different
RESULT AND DISCUSSION
157
morphologies and properties of crystals obtained from different solvent mixtures and
present research work result are accordance with the previous research finding result
201.
Dissolution of super cooling crystals of mefenamic acid was not improved
than commercial sample; these findings are in agreement with the previous findings
171, 173.
Further, the results of dissolution studies were submitted to two analysis of
variance using graph pad instat programme in order to determine significant
differences. ANOVA results showed that the critical value of F at 5% level of
significance (4.366%) with p value 0.0038, between the treatments. Freeze dried
crystals has more significant influence on the dissolution of than others prepared
crystals. (26.71), and super cooling crystals shown no significant influence on the
dissolution (0.06). Paired t – test was applied so that comparisons of two means are
made on related samples. Pairing was found to be effective for all the crystals
dissolution profiles except super cooling crystals. Significant differences were
observed for dissolution rate of all the prepared crystals except super cooling crystals
(p>0.05), in the following order FD (0.004)>SD (0.0002) >SA (0.00002)>RS
(0.003)>SC (0.12).
RESULT AND DISCUSSION
158
Table 32: Dissolution data of Mefenamic acid crystals in Phosphate buffer pH7.4
Sl. No Time
(in min)
CS
% release
mean ±S.D.*
RS
% release
mean ±S.D.*
SA
% release
mean ± S.D*
SD
% Release
mean ± S.D*
FD
% release
mean ± S.D*
SC
% release
mean ± S.D*
1 0.00 0.00±0.34 0.00 0.00 0.00 0.00 0.00
2 10 1.17±0.04 2.13±0.04 17.13±0.30 15.71±0.01 7.91±0.05 0.090±1.01
3 20 4.06±0.01 5.56±0.01 20.41±0.02 19.52±0.02 18.04±0.01 0.091±2.02
4 30 5.09±0.02 7.78±0.20 22.91±0.02 25.36±0.03 25.32±0.02 --
5 40 9.47±0.05 11.92±0.02 26.76±0.12 29.97±0.03 35.62±0.01 --
6 50 13.9±0.01 14.61±0.04 30.65±0.23 36.55±0.04 43.77±0.34 --
7 60 18.5±0.11 20.40±0.01 35.73±0.25 48.13±0.02 56.31±0.04 --
*Standard deviation, n = 3.
Figure 32: Dissolution profile of different Mefenamic acid Crystals in Phosphate
buffer pH7.4
CS-Commercial sample, RS- Recrystallized sample, SC- Super cooling, SA-
Spherical crystallization, SD- Spray dried, FD- Freeze dried.
0
10
20
30
40
50
60
70
0 10 20 30 40 50 60 70
% R
elea
se
Time (min.)
Pure M.acid RS SC SA SD FD
RESULT AND DISCUSSION
159
5.7.5 Various parameters of model fittings of Mefenamic acid samples
Table 33: Data of various parameters of model fittings of Mefenamic acid
samples
Drug Different
crystals
Zero
Order
(r2)
First
Order
(r2)
Higuchi
(r2)
Korsemeyer
/ Peppas
(r2)
n Value
Peppas
Mefanamic
acid
CS 0.9522 0.3388 0.7744 0.7791 0.5671
SA 0.8804 0.3248 0.9837 0.9380 0.6723
RS 0.9795 0.3407 0.8408 0.9184 0.5972
FD 0.9961 0.2343 0.8904 0.9975 0.7842
SD 0.9612 0.2842 0.9402 0.9704 0.8734
SC - - - - -
The release kinetics was evaluated by using zero order, first order, Higuchi’s
diffusion and Korsemeyer - Peppas equation. Calculated regression co efficient values
for different crystals are tabulated in Table 33. These values are compared with each
other for model equation. Based on the higher regression values (r2), the best fit model
was zero order for all the prepared crystals and the drug release kinetics followed
diffusion controlled mechanism. Release kinetic profile of Mefenamic acid crystals
for zero order, first order, Peppas and Higuchi are presented in Figure 33. The Peppas
model is widely used, when the release mechanism is not well known or when more
than one type of release phenomenon could be involved. ‘n’ value could be used to
characterize different release mechanism.
‘ n’ Mechanism of drug release
0.5 Fickian diffusion
0.5 < n < 1 Non fickian diffusion
1 Case II transport
The ‘n’ values obtained graphically from peppas plot are shown in Table 25. As the
values obtained were more than 0.5, indicates that the release approximates non
fickian diffusion for all the prepared crystals of mefenamic acid.
RESULT AND DISCUSSION
160
Figure 33: Release mechanism of Mefenamic acid
RESULT AND DISCUSSION
161
5.7.6 Stability study:
The dissolution behaviour of Mefenamic acid prepared crystals shall remain unchanged during storage. The results of the
stability study (Table 34) of prepared crystals of Mefenamic acid stored at 400C and 75% relative humidity for 6 month with
accordance with ICH guidelines194
. The influence of physical stability on the prepared crystals was investigated. The results of
stability presented in the Table 34.
Table 34: Stability data of prepared crystals of mefenamic acid by different crystallization techniques
Sample name: Mefenamic acid Spherical crystals
Storage condition: 400C /75% RH
Testing interval Description of drug FT-IR Study XRD Study Drug content ( ±SD) Dissolution Study (±SD) Water content (%w/w)
Initial Light yellow crystals Complies Complies 98.46±0.01 35.73±0.03 4.34
1 month Complies Complies Complies 98.36±0.01 35.71±0.02 --
2 month Complies Complies Complies 98.22±0.02 35.71±0.02 --
3 month Complies Complies Complies 98.14±0.01 35.67±0.01 --
6 month Complies Complies Complies 97.53±0.02 35.54±0.02 4.39
Sample name: Mefenamic acid Spray dried crystals
Storage condition: 400C /75% RH
Initial Light yellow crystals Complies Complies 99.43±0.02 48.13±0.01 4.47
1 month Complies Complies Complies 99.38±0.03 48.11±0.02 --
2 month Complies Complies Complies 99.29±0.01 47.08±0.01 --
3 month Complies Complies Complies 99.16±0.01 46.98±0.02 --
6 month Complies Complies Complies 98.32±0.02 46.86±0.03 4.62
Sample name: Mefenamic acid Freeze dried crystals
Storage condition: 400C /75% RH
Initial Light yellow crystals Complies Complies 99.12±0.02 56.31±0.02 5.32
1 month Complies Complies Complies 99.11±0.03 56.29±0.03 --
2 month Complies Complies Complies 99.10±0.02 56.24±0.02 --
3 month Complies Complies Complies 99.08±0.01 56.24±0.01 --
6 month Complies Complies Complies 99.02±0.02 55.14±0.02 5.71
Prepared crystals of mefenamic acid were stable and compiled with its all the properties when compare to freshly prepared
crystals of Mefenamic acid.
RESULT AND DISCUSSION
162
5.8 Preparation of conventional tablet:
Mefenamic acid conventional tablets were prepared by direct compression using direct
compressible excipients.
5.8.1 Evaluation of Mefenamic acid tablets:
Tablets containing pure sample and prepared crystals were evaluated for Average
weight, hardness and percentage friability the limit (Table 35). Percentage weight
variations, content of active ingredient were within IP limits.
Table 35: Evaluation of prepared Mefenamic acid tablets
Different
crystals Formulations
Average
weight
(mg) ( ±SD)
Hardness in
kg/cm2
(±SD)
Friability*
(%) ( ±SD)
Content
uniformity*
(%) ( ±SD)
Commercial
sample
CS1 251.7±2.9 5.12±0.96 0.56±0.71 101.31±0.23
CS2 253.3±2.8 5.02±0.13 0.41±0.92 99.18±0.21
Spherical
crystals
SA1 250.7±1.8 5.23±0.41 0.63±0.51 99.66±0.13
SA2 251.5±3.0 5.51±0.43 0.34±0.11 99.33±0.11
Spray dried
crystals
SD1 251.3±2.4 5.17±0.25 0.72±0.65 99.86±0.03
SD2 249.7±2.9 5.52±0.76 0.44±0.11 99.51±0.83
Freeze
dried
crystals
FD1 251.6±2.8 5.22±0.63 0.21±0.32 99.88±0.41
FD2 248.7±1.8 5.30±0.51 0.34±0.21 99.36±0.57
Super
cooling
crystals
SC1 252.05±1.0 5.16±0.48 0.89±0.47 24.63±0.13
SC2 250.23±1.9 5.51±0.26 0.73±0.66 23.96±0.03
Marketed
product MP 250.93±0.9 5.42±0.79 0.26±0.21 99.81±0.43
1-Sodium starch glycolate,
2-Povidone, MP-=Market Product
*Mean SD± n=3
5.8.2 Dissolution studies of tablets:
Dissolution studies of prepared tablets were performed using USP XXIV
dissolution test apparatus Type II. The dissolution profile is shown in the Figure 34 & 35.
The dissolution profiles of Mefenamic acid tablets contains spherical crystals, spray dried
and freeze dried crystals exhibited better dissolution behavior than tablets contains
RESULT AND DISCUSSION
163
commercial sample. In case of crystals prepared by Recrystallization and super cooling
techniques dissolution is not enhanced as compare to other techniques. Percentage release
of mefenamic acid from super cooling crystals was very low when compare to commercial
sample, this may be due to the change in crystals habit or change in the crystal morphology
due to heat But the dissolution profile of the marketed tablets is much better than the tablets
prepared by different crystallization techniques. Table 36 & 37 contain the release data of
Mefenamic acid from prepared Mefenamic acid tablets (1-
Sodium starch glycolate, 2-
Povidone) and marketed tablets respectively.
Further, the results of dissolution were submitted to two analysis of variance using
graph pad instat program in order to determine significant differences. ANOVA results
showed that the critical value of F at 5% level of significance (4.03), between the
treatments. Tablets containing Freeze dried crystals has more significant influence on the
dissolution of than others Tablets contains crystals. (47.39) and super cooling crystals had
shown not significant influence on the dissolution (1.67).
Paired t – test was applied so that comparisons of two means are made on related
samples. Pairing was found to be effective for all the crystals dissolution profiles except
super cooling crystals. Significant differences were observed for dissolution rate of all the
prepared crystals i.e. (p>0.05), except super cooling crystals, in the following order FD
>SD >SA >RS >SC.
Table 36: Dissolution data of prepared Mefenamic acid and market tablet in
Phosphate buffer pH7.4
Sl.
No.
Time
(in
min)
CS1
Average
% release
mean±SD*
RS1
Average
% release
mean±SD*
SA1
Average %
release
mean±SD*
SD1
Average
% release
mean±SD*
FD1 Average
% release
mean±SD*
SC1 Average
% release
mean±SD*
MP Average
% release
mean±SD*
1 0 0.00 0.00 0.00 0.00 0.00 0.00 0.00
2 10 17.1±0.03 15.72±0.11 18.0±0.33 19.3±0.05 20.2±0.03 0.36±0.03 21.21±0.02
3 20 20.4±0.01 19.63±0.11 25.2±0.22 29.4±0.06 29.4±0.02 0.94±0.02 32.52±0.04
4 30 23.1±0.13 25.35±0.13 35.5±0.01 47.8±0.03 47.8±0.04 1.54±0.06 48.76±0.05
5 40 27.1±0.11 29.91±0.03 43.6±0.02 60.9±0.04 64.9±0.01 2.15±0.07 68.10±0.01
6 50 31.6±0.12 36.52±0.13 56.2±0.04 69.2±0.04 79.5±0.05 2.82±0.02 83.20±0.04
7 60 36.0±0.12 45.01±0.15 67.2±0.17 79.8±0.03 89.9±0.06 3.91±0.01 95.21±0.05
1-Sodium starch glycolate
RESULT AND DISCUSSION
164
Figure 34: Dissolution profile of Mefenamic acid from prepared tablets in Phosphate
buffer pH7.4
CS-Commercial sample, RS- Recrystallized sample, SC- Super cooling, SA- Spherical
crystallization, SD- Spray dried, FD- Freeze dried, MP-Market product.
Table 37: Dissolution data of prepared Mefenamic acid and market tablet in
Phosphate buffer pH7.4
Sl.
No.
Time
(in min)
CS2
Average %
release
mean±SD*
RS2
Average %
release
mean±SD*
SA2
Average %
release
mean±SD*
SD2
Average %
release
mean±SD*
FD2
Average %
release
mean±SD*
SC2
Average %
release
mean±SD*
MP
Average %
release
mean±SD*
1 0 0.00 0.00 0.00 0.00 0.00 0.00 0.00
2 10 14.11 14.22 18.53 18.40 19.53 0.35 21.21
3 20 18.20 18.43 25.52 27.86 27.54 0.98 32.52
4 30 21.43 23.53 33.64 45.85 44.63 1.46 48.76
5 40 25.42 27.43 41.53 61.32 63.53 2.13 68.10
6 50 29.89 33.64 55.63 68.43 78.53 2.91 83.20
7 60 34.42 43.65 65.78 77.63 86.32 3.92 95.21
2-Povidone
-20
0
20
40
60
80
100
120
0 10 20 30 40 50 60
% R
ele
ase
Time (min)
CS RS SC SA SD FD MP
RESULT AND DISCUSSION
165
Figure 35: Dissolution profile of Mefenamic acid from prepared tablets in Phosphate
buffer pH7.4
CS-Commercial sample, RS- Recrystallized sample, SC- Super cooling, SA- Spherical
crystallization, SD- Spray dried, FD- Freeze dried, MP-Market product.
Dissolution data analysis:
Differential and Similarity factor for prepared tablets formulations
Formulations
Differential
Factor
(f 1)1
Differential
Factor
(f 1)2
Similarity
Factor
(f 2)1
Similarity
Factor
(f 2)2
CS 62.22 63.88 66.00 65.72
RS 52.75 54.18 69.40 67.53
SA 29.43 30.92 74.13 73.61
SD 16.19 18.47 80.63 79.18
FD 5.58 9.34 92.21 86.60
SC 95.94 95.93 61.26 61.30
CS-Commercial sample, RS- Recrystallized sample, SC- Super cooling, SA- Spherical
crystallization, SD- Spray dried, FD- Freeze dried.
Differential factor (f1) and Similarity factor (f2) was calculated for prepared tablets
containing crystals (1 & 2 formulation). The Differential factor (f1) for FD was nearer to 0
and for SC it shows >95. The Similarity factor (f2) for SA, SD and FD was more than 100
and for SC it was ~60 for both 1 & 2 formulations respectively.
From the above result it can be conclude that tablets containing FD crystals were
similar to marketed product and tablet containing SC crystals least similar222
.
0
20
40
60
80
100
120
0 10 20 30 40 50 60
% R
elea
se
Time (min)
CS
RS
SC
SA
SD
FD
MP