A Non-aqueous Potentiometric Titration Method for Validation of Emtricitabine from Pharmaceutical Dosages
Rajan V. Rele, Prathamesh P. Tiwatane
Department of Chemistry, D.G. Ruparel College Mahim, Mumbai 400 016.
*Corresponding Author E-mail: drvinraj@gmail.com
ABSTRACT:
It is a 4-amino-5-fluoro-1-[(2R,5S)-2 (hydroxyl -methyl)-1,3-oxathiolan-5-yl]-1,2-dihydro- pyrimidin-2-one. Emtricitabine is a nucleoside reverse transcriptase inhibitor (NRTI) for the treatment of HIV infection in adults. EMT is an analogue of cytidine. The drug works by inhibiting reverse transcriptase, the enzyme that copies HIV RNA into new viral DNA.
According to the literature review several methods has been developed for drug, like spectroscopy methods1-6. HPLC7-22, and HPTLC23. These nonaqueous titration methods can be used for the routine analysis. In the proposed methods optimization and validation of this method are reported.
Structure of Emtricitabine:
EXPERIMENTAL:
Instrumentation:
An potentiometric titrator was used (Lab- India auto titrator) for assay method development and validation.
A Simadzu analytical balance with 0.01 mg was used.
Reagents and chemical:
Reference standard of emtricitabine was obtained from reputed firm with certificate of analysis.
Potassium hydrogen phthalate, perchloric acid and glacial acetic acid of A. R. grade were used.
General procedure:
Standardization of 0.1 N perchloric acid:
About 0.350 mg of potassium hydrogen phthalate (previously powdered lightly, dried at 120oC for 2 hours) was weighed accurately into clean and dry titration jar. It was dissolved in 50 ml of glacial acetic acid. About 0.1 ml of crystal violet solution (0.5 % w/v in anhydrous glacial acetic acid) was added. It was titrated with 0.1 N perchloric acid until violet colour changes to emerald green. Blank determination was performed out for necessary correction. The titration was performed in duplicate.
One ml of 0.1 N HClO4 is equivalent to 0.2042 gm of potassium hydrogen phthalate (C8H5KO4)
W
Normality of perchloric acid = ---------------------
B.R. x 0.2042
Where W is weight of potassium hydrogen phthalate in g. B.R. is burette reading in ml.
Quantitative determination of emtricitabine:
About 0.100 g. of emtricitabine test sample was weighted accurately into a clean and dried titration jar. It was dissolved in 35 ml. of anhydrous glacial acetic acid and 15 ml of 5% (w/v) mercuric acetate.
It was titrated with 0.1 N perchloric acid potentiometrically.
Blank determination was also carried out for necessary correction.
One ml of 1 N perchloric acid is equivalent to 0.082416 g. of emtricitabine
% (Percentage) emtricitabine on the dried basis was calculated as below
B.R. x N x 0.082416 x 100
% assay = _____________________________
W
Where B.R. is burette reading in ml at the potentiometric end point.
N is actual normality of 0.1 N perchloric acid.
W is weight of the sample taken in g.
RESULT AND DISCUSSION:
Determination of emtricitabine:
The objective of this work was to determine accurately the content of emtricitabine. The assay of emtricitabine (on the dried basis) of various batches of emtricitabine test sample was analyzed using the above method. It was in the range of 99.85 % to 99.95 %.
Analytical method validation:
The method precision was checked after analyzing six different preparations of homogeneous test sample of emtricitabine. The % RSD of results obtained was found to be 0.06509. It confirms good precision of the method. The results are presented in table 1.
Table No. 1: Method of Precision
|
Weight of drotaverine hydrochloride in g. |
Burrete Reading in ml. |
Normality of perchloric acid |
% Assay |
|
0.1 |
12.12 |
0.0999 |
99.7883 |
|
0.1 |
12.14 |
0.0999 |
99.870 |
|
0.1 |
12.11 |
0.0999 |
99.95297 |
|
0.1 |
12.13 |
0.0999 |
99.80578 |
|
0.1 |
12.12 |
0.0999 |
99.7883 |
|
0.1 |
12.12 |
0.0999 |
99.80827 |
|
|
|
Mean |
99.83571 |
|
|
|
Standard deviation |
0.064983 |
|
|
|
%RSD |
0.06509 |
Linearity:
For the establishment of method linearity, five different weights of emtricitabine test samples corresponding to 20%, 40 %, 60 %, 80 % and 100 % of the about weight (0.100 g.) were taken and analyzed for % (percentage) of emtricitabine content. The results are in table 2.
Table No. 2: Linearity
|
Level |
Weight of emtricitabine in mg. |
Burette reading in ml. |
Normality of perchloric acid |
% Assay |
|
1 |
20 |
2.42 |
0.0999 |
99.62364 |
|
2 |
40 |
4.84 |
0.0999 |
99.62364 |
|
3 |
60 |
7.27 |
0.0999 |
99.76086 |
|
4 |
80 |
9.69 |
0.0999 |
99.72655 |
|
5 |
100 |
12.12 |
0.0999 |
99.7883 |
|
|
|
|
mean |
99.6911 |
|
|
|
|
Standard Deviation |
0.069792 |
|
|
|
|
%R SD |
0.070009 |
The potentiometric titration was conducted once at each level. Calibration curve was drawn by plotting test sample weight in gram on x axis and titre values on y axis.
The values of correlation coefficient, slope and intercept are given in table 3. And graph is given in fig no. 1
Table No. 3 : Regression values
|
Correlation Coefficient |
0.9999 |
|
Slope (m) |
121.09 |
|
Intercept ( c ) |
0.0007 |
|
Regression equation |
y = 121.09x + 0.0007 |
Fig no. 1: Linearity graph
Table No 4 Accuracy and Recovery
|
Level |
Volume in ml |
Weight of emtricitabine added in mg. |
Weight of emtricitabine found in mg. |
% Assay |
Mean % assay |
|
1 |
1.21 |
0.01 |
0.010042 |
100.4214 |
|
|
|
1.22 |
0.0101 |
0.010125 |
100.2489 |
100.087 |
|
|
1.2 |
0.010 |
0.009959 |
99.59149 |
|
|
2 |
2.42 |
0.020 |
0.020084 |
100.4214 |
|
|
|
2.43 |
0.0202 |
0.020167 |
99.83801 |
99.922 |
|
|
2.41 |
0.0201 |
0.020001 |
99.50891 |
|
|
3 |
3.63 |
0.030 |
0.030126 |
100.4214 |
|
|
|
3.64 |
0.0301 |
0.030209 |
100.3635 |
100.088 |
|
|
3.62 |
0.0302 |
0.030043 |
99.48157 |
|
|
4 |
4.84 |
0.040 |
0.040169 |
100.4214 |
|
|
|
4.85 |
0.0402 |
0.040252 |
100.1283 |
100.171 |
|
|
4.83 |
0.0401 |
0.040086 |
99.96403 |
|
|
5 |
6.025 |
0.050 |
0.050003 |
100.0065 |
|
|
|
6.024 |
0.0501 |
0.049995 |
99.79028 |
99.807 |
|
|
6.026 |
0.0502 |
0.050012 |
99.62456 |
Accuracy and recovery:
Accuracy was determined at five different levels i.e., 20%, 40%, 60%, 80% and 100% of the nominal concentration. (0.100 g.) The titration was conducted in triplicate at each level and the titre value was recorded. The tire value obtained in linearity study was considered as true value during the calculation of percentage (%) recovery. The percentage recovery is calculated using following equation.
Titre value x 100
Percentage recovery = -----------------------------
True titre value
The percentage range recovery of emtricitabine was in 99.80 to 100.17 %. It confirms the accuracy of the proposed method. (Table 4).
Ruggedness:
The ruggedness of the method is defined as degree of reproducibility of results obtained by analysis of emtricitabine sample under variety of normal test conditions such as different laboratories, different analysts and different lots of reagents. Quantitative determination of emtricitabine was conducted potentiometrically on one laboratory. It was again tested in another laboratory using different instrument by different analyst. The assays obtained in two different laboratories were well in agreement. It proved ruggedness of the proposed method.
The proposed method of non-aqueous potentiometric titration was found to be precise, accurate and rugged. The values of percentage recovery and standard deviation showed sensitivity. The method was completely validated. It showed satisfactory data for all the parameters of validation. Hence it can be applied for routine quality control application.
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Received on 27.07.2024 Modified on 12.08.2024
Accepted on 30.08.2024 ©AJRC All right reserved
Asian J. Research Chem. 2024; 17(4):199-202.