Assessment of Phase Diagrams by Cut and
Weigh Method: A Technical Note
K. Pramod1, Shahid H. Ansari2, Javed
Ali1*
1Department of
Pharmaceutics, Faculty of Pharmacy, Jamia Hamdard, Hamdard Nagar, New
Delhi–110062, India.
2Department of
Pharmacognosy & Phytochemistry, Faculty of Pharmacy, Jamia
Hamdard, Hamdard Nagar,
New
Delhi – 110062, India.
*Corresponding Author E-mail:- javedaali@yahoo.com
ABSTRACT:
A good number of
manuscripts are being published where the authors carry out qualitative
assessment of the region of interest in the phase diagrams. These qualitative
assessments can be sensed as a lack of sound technique for quantitative
assessment. In the present technical note we report a simple cut and weigh
method to offer a quantitative method for assessing phase diagrams. The method
was validated and explained using sample analysis. The method was found to be
robust, quantitative and enabled statistical comparison of the results. The
method was found to be excellent even when applied for samples below a weight
of 10 mg and that too on different paper sheets. The suggested technique of
expressing the result as a percentage of the total area could offer a universal
applicability of the method for assessment of phase diagrams. The developed and
validated method would be a great tool for the researchers requiring
quantitative assessment of phase diagrams.
KEYWORDS: Nanoemulsion, Microemulsion,
Validation, p value
INTRODUCTION:
A large number of manuscripts are being published on emulsion
systems and in particular microemulsion/ nanoemulsion systems. Many of them include the preparation
of phase diagrams and selection of some components based on the area of the
region of interest (emulsion/ nanoemulsion/ microemulsion region). But usually the authors make a
qualitative assessment for the conclusion.1-9 This
will work in case where the difference in the areas is large. In this scenario,
our effort to have a reported validated method for cut and weigh method for
assessing areas in the phase diagrams is important. Though the cut and weigh
method is useful in biopharmaceutics in the
assessment of some pharmacokinetic parameters, no reports have been till date
for the use and validation of this method in phase diagrams. We developed a
very simple method for the assessment of phase diagrams and the developed
method is absolutely quantitative and enables statistical assessment of the
results for comparison.
Materials and Methods:
Materials
Paper sheets (A4 size, Century Pulp and Paper, Nainital,
India) were used for the study.
Methods
Method validation
The method was validated by weighing triangles (which represents
100% area in case of ternary and pseudo-ternary phase diagrams) of three
different areas (low, middle and high) simulating the range of possible areas to
be formed in the phase diagrams. The validation was carried out for intra-sheet
(areas within a single A4 size sheet) and inter-sheet (areas within different
A4 size sheets). The outlines of the triangles were printed on the A4 size
sheets. The triangles were carefully separated from the sheets by cutting using
scissors exactly through the outlines of the triangle. The cut triangles were
weighed using an electronic balance (Mettler Toledo
Inc., OH, USA). The data were compared for statistical
significance by one way analysis of variance (ANOVA) followed by Tukey-Kramer multiple comparison tests using GraphPad Instat software (GraphPad Software Inc., CA, USA).
Table 1: Validation data for
cut and weigh method
|
Sl. No. |
Intra-sheet validation |
||||||||
|
Low (mg) |
Middle (mg) |
High (mg) |
|||||||
|
A |
B |
C |
D |
E |
F |
G |
H |
I |
|
|
1 |
29.4 |
29.5 |
28.4 |
80.5 |
78.3 |
80.0 |
152.5 |
154.4 |
159.5 |
|
2 |
28.6 |
29.8 |
29.7 |
80.1 |
77.1 |
79.1 |
155.6 |
159.3 |
154.1 |
|
3 |
27.6 |
29.1 |
28.0 |
80.5 |
76.2 |
78.2 |
161.8 |
157.2 |
157.8 |
|
4 |
28.4 |
26.5 |
28.4 |
77.6 |
79.7 |
79.8 |
153.2 |
158.7 |
152.3 |
|
5 |
26.9 |
27.8 |
27.6 |
81.4 |
79.9 |
79.8 |
150.0 |
152.4 |
153.8 |
|
6 |
27.5 |
26.9 |
27.3 |
83.2 |
79.4 |
80.2 |
150.5 |
159.2 |
155.1 |
|
Mean |
28.07 |
28.27 |
28.23 |
80.55 |
78.43 |
79.52 |
153.93 |
156.87 |
155.43 |
|
±SD |
0.90 |
1.40 |
0.84 |
1.83 |
1.51 |
0.74 |
4.35 |
2.86 |
2.70 |
|
% RSD |
3.21 |
4.95 |
2.98 |
2.27 |
1.93 |
0.93 |
2.83 |
1.82 |
1.74 |
|
Inter-sheet validation |
p>0.05
between all samples |
p>0.05
between all samples |
p>0.05
between all samples |
||||||
Figure 1: Illustration for
sample analysis using cut and weigh method
Sample analysis
As an illustration we have prepared three phase diagrams. The
region marked in the phase diagrams were quantitatively evaluated by
determining the percentage weight (with respect to the total weight for the
triangle in the phase diagram) by the above validated method. A column chart of
percentage weight of shaded area obtained for the samples was prepared.
RESULTS
AND DISCUSSION:
Method validation
The method was found to be valid as indicated by the data of
intra-sheet and inter-sheet validations (Table 1). The low percentage relative
standard deviations (< 5%) for the samples in the intra-sheet validation
indicated the robustness of the method. The inter-sheet validation revealed
that there was no statistically significant difference (p>0.05) between the
mean weights obtained from three different sheets.
Sample analysis
As an illustration we have tried to explain
the use of the method using Figure 1. It can be observed that the marked areas
are different in shapes. We have used definite triangular shapes to make
readers understand that how same area itself can confuse the author in
qualitatively assessing the area of region of interest. This becomes worse as
the shapes deviate from definite symmetrical shapes, as is always observed in
real practice. The qualitative assessment by visual method could be a real brain
teaser in these situations and the results depend on the individual. The data
for sample analysis by cut and weigh method is displayed in Table 2.
It is very clear from the results (Figure 2) that there are no
statistically significant differences between the % areas marked in the
triangles. In other words all the areas are similar in magnitude. This method
is thus shape independent also. The use of percentage area for assessment has
the advantage of normalizing the values. Thus the percentage area will remain
same for a particular phase diagram whatever the size of the phase diagram the
researcher use for assessing the region of interest.
Figure 2: Column chart for
percentage weight of shaded area
Table 2: Data for the sample
analysis
|
Sl. No. |
Weight of shaded area (mg) |
% Weight of shaded area* |
||||
|
X |
Y |
Z |
X |
Y |
Z |
|
|
1 |
7.9 |
8.3 |
8.4 |
6.22 |
6.54 |
6.62 |
|
2 |
7.7 |
8.2 |
7.8 |
6.07 |
6.46 |
6.15 |
|
3 |
7.6 |
7.9 |
8.4 |
5.99 |
6.22 |
6.62 |
|
4 |
8.1 |
7.9 |
8.1 |
6.38 |
6.22 |
6.38 |
|
5 |
8.2 |
8.1 |
7.9 |
6.46 |
6.38 |
6.22 |
|
6 |
8.0 |
7.7 |
7.6 |
6.30 |
6.07 |
5.99 |
|
Mean |
7.92 |
8.02 |
8.03 |
6.24 |
6.32 |
6.33 |
|
±SD |
0.23 |
0.22 |
0.33 |
0.18 |
0.18 |
0.26 |
*Calculated using mean weight of triangle (100% area) 126.92±1.65
mg (n=6)
In the present example we have carried out the analysis at the
least areas as is evident from the data that all the weights were well below 10
mg. Also the weights for the individual areas and the triangle were taken from
different sheets. Thus it was confirmed that even in the worst of the
experimental conditions the method is very much practically applicable.
Conclusions:
We have successfully validated the cut and weigh method for use in
the assessment of areas in the phase diagrams. The method was found to be
statistically validated. The sample analysis illustrated helps to understand
and establish the use of the validated method in phase diagrams. The method in
its form can be extended to study the effect of individual components on the
area of interest. This can be done by cutting and weighing the area of interest
at particular regions of the phase diagrams where we suppose to have the effect
of the particular component.
Acknowledgements:
Pramod K. gratefully acknowledges Indian Council of Medical
Research (ICMR), New Delhi, India, for providing Senior Research Fellowship
(No. 35/3/10/NAN/BMS).
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Received on 15.05.2013 Accepted on 27.06.2013
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Asian J. Pharm.
Tech. 2013; Vol. 3: Issue 3, Pg 104-106