Derivative UV Spectrophotometric Method for Validation of Emtricitabine in Bulk and Pharmaceutical Dosage Form

 

Rajan V. Rele *, Prathamesh P. Tiwatane

Central Research Laboratory, D.G. Ruparel College, Matunga, Mumbai 400016

*Corresponding Author E-mail: drvinraj@gmail.com

 

ABSTRACT:

Simple and precise derivative UV spectrophotometric method i.e. first order derivative method have been developed and validated for the estimation of emtricitabine in bulk and its tablet formulation. The standard and sample solutions of emtricitabine were prepared in distilled water. Emtricitabine was estimated by first order at wavelength 269 nm respectively. Beer’s law was obeyed in the concentration range of 1 to 20 μg / ml with coefficient of correlation value 0.9999. These methods were tested and validated for various parameters according to ICH guidelines. The precision expressed as relative standard deviation was 0.7852 %for the method respectively. The proposed method was successfully applied for the determination of emtricitabine in pharmaceutical formulation. Results of the analysis were validated statistically and were found to be satisfactory. The proposed method is simple, easy to apply, low-cost and require relatively inexpensive instruments.

 

KEYWORDS: Emtricitabine, First order derivative UV spectroscopy.

 

 

INTRODUCTION:

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 methods1-2. HPLC3-19, and HPTLC20. This HPLC method can be used for the routine analysis. In the proposed methods optimization and validation of this method are reported.

 

 

 

 

 

 

Structure of emtricitabine:

 

 

MATERIAL AND METHODS:

Shimadzu UV-1800 was used with 10 mm matched quartz cell to measure absorbance of solution.

A Shimadzu analytical balance with 0.01mg was used.

 

Chemical and reagents:

Reference standard of emtricitabine was obtained with certificate analysis. All spectral absorbance measurements were made on Shimadzu UV-1800 with 10 mm matched cell.

 

Preparation of standard solution:

About 10 mg of standard Emtricitabine was weighed accurately and transferred in 100 ml of volumetric flask. About 30 ml of distilled water was added and sonicated for 15 minutes. The volume was adjusted up to the mark with distilled water to give concentration as 100μg/ml.

 

Fig. 1: Overlay spectra of first order derivative of emtricitabine in the concentration range of 2 – 20 µg/ ml.

 

Estimation from tablets:

Twenty tablets were weighed accurately and average weight of each tablet was determined. Powder equivalent to 10mg of emtricitabine was weighed and transferred in 100ml of volumetric flask. A 30ml of distilled water was added and sonicated for 15 minutes and filtered. The filtrate and washing were diluted up to the mark with distilled water to give concentration as 100μg/ml. Such solution was used for analysis.

 

Experimental:

Method: First order derivative method:

For the selection of analytical wavelength, 10μg/ml solution of Emtricitabine was scanned in the spectrum mode from 400nm to 200 nm by using distilled water as blank. The first order derivative spectrum was obtained by using derivative mode by UV probe 2.42 software. From the spectrum, the amplitude of the derivative spectrum was measured at 269 nm.

 

 

Fig. 2(a). Calibration curve for Emtricitabine by First order derivative method in the concentration range of 2-20 µg/ml.

 

Preparation of calibration curves:

Series of solutions containing 2 – 20µg/ml of emtricitabine were used to determine linearity of the proposed method respectively. Solutions were scanned in the spectrum mode and absorbance spectra were converted to first order derivative spectra [Fig. 1].

 

After observing the overlain first order derivative spectra of Emtricitabine, wave length selected was 269 nm, where emtricitabine showed considerable absorbance. The calibration curves were plotted of dA/dλ against concentrations [Fig. 2 (a)].

 

Results of analysis are given in table 1.

Table 1: Values of results of optical and regression of drug     

Parameter

First order derivative

Detection Wavelength (nm)

269

Beer Law Limits (µg/ml)

2-20

Correlation coefficient(r2)

0.9999

Regression equation (y=b+ac)

 

Slope (a)

0.001

Intercept (b)

0.00004

 

Estimation from tablets:

Twenty tablets were weighed accurately and average weight of each tablet was determined. Powder equivalent to 10mg of Emtricitabine was weighed and transferred in 100ml of volumetric flask. A 30ml of 0.1N distilled water was added and sonicated for 15 minutes and filtered. The filtrate and washing were diluted up to the mark with distilled water to give concentration as 100μg/ml of each drug. Such solution was scanned in the range of 200-400 nm against distilled water as blank. The absorbance spectra were converted to first order derivative spectra. Calculations were done as per the equations. The concentrations of Emtricitabine present in tablets were calculated by substituting the values of absorbance in linearity equations.

 

(a) For first order derivative method, Y = 0.001x + 4E-05

 

 

 

Table 2 : Statistical evaluation of the data subjected to accuracy for first order derivative method

Amount of Sample

Added in (µg/ml)

Amount of Standard Added in (µg/ml)

Total amount recovered

Percentage recovery (%)

Standard deviation

Percentage of relative standard deviation (C.O.V.)

2

0

2.0042

100.21

0.01133

0.5657

2

2

4.0085

100.21

0.00690

0.1721

2

4

6.0057

100.09

0.01272

0.2118

2

6

8.0100

100.12

0.01154

0.1441

 

 

 

Mean=100.16

Mean=0.01062

Mean=0.2734

 

Method Validation:

These methods were validated according to ICH guidelines.

 

Accuracy:

To ascertain the accuracy of proposed methods, recovery studies were carried out by standard addition method. Percent recovery for Emtricitabine was found as 100.16 % for first order derivative method respectively. (Table 2).

 

Linearity:

The linearity of measurement was evaluated by analyzing different concentration of the standard solutions of Emtricitabine in the range of 2-20µg/ml for first order respectively.

 

Precision:

The method precision was established by carrying out the analysis of tablets powder blend containing Emtricitabine. The assay was carried out for the drugs by using proposed analytical method in six replicates. The values of relative standard deviation were well 0.2734 % for Emtricitabine respectively indicating the sample repeatability of the methods. The results obtained are tabulated in table 3.

 

Table 3: Statistical evaluation of the data subjected to method of precision

Sr. No.

Sample No.

First order derivative method values

1

1

100.50

2

2

100.497

3

3

98.504

4

4

99.005

5

5

101.005

6

6

99.004

Mean % assay

99.919

%R.S.D.

0.7852

 

Intra-day precision was estimated by assaying tablets powder blend containing emtricitabine. The assay was carried out for the drugs by using proposed analytical method in six replicates. The results were average for statistical evaluation.

 

Inter-day precision was estimated by assaying tablets powder blend containing Emtricitabine for three consecutive days (i.e. 1st, 3rd and 5th days). The statistical validation data for intra and inter day precision is summarized in table 4.

 

 

Table 4: Summary of validation parameter for intra-day and inter-day

Sr. No.

Parameters

First order Derivative values

1

Intra-day precision

(N=3) amount found ± % R.S.D.

99.78%

0.7129

2

Inter-day precision

(N=3) amount found ± % R.S.D.

99.12

0.5712

 

Both intra- day and inter-day precision variation found to be less in % RSD values. It indicates high degree of precision of the method.

 

RESULT AND DISCUSSION:

The developed first order derivative spectrophotometric methods for determination of emtricitabine in tablet formulation was found to be simple and convenient for the routine analysis of drug. The proposed method is accurate, precise and reproducible. It is confirmed from validation data as given in tables 1 to 4. The % RSD was found to be less than 1, which indicates validity of method. Linearity was observed by linear regression equation method for emtricitabine in different concentration range. The correlation coefficient of these drugs was found to be close to 1.00, indicating good linearity figure 2.

 

The assay results obtained by proposed method is shown in table 2 are in good agreement. Hence proposed method can be used for routine analysis pharmaceutical dosage form. Methods are simple, accurate, precise, reliable, rapid, sensitive, reproducible and economical. It is validate as per ICH guidelines.

 

The proposed methods are simple, precise, accurate and rapid for the determination of emtricitabine pharmaceutical dosage form. The methods do not require any ratio of first order derivative. The amplitude of first  order derivative can be directly used to assay of formulation. This method can be adopted as an alternative to the existing methods. It can be easily and conveniently adopted for routine quality control analysis.

 

ACKNOWLEDGMENT:

Authors express sincere thanks to the Principal, Dr. Tushar M. Desai of D. G. Ruparel college.

 

 

 

 

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Received on 28.10.2019                    Modified on 31.12.2019

Accepted on 20.01.2020                   ©AJRC All right reserved

Asian J. Research Chem. 2020; 13(1): 48-51.

DOI: 10.5958/0974-4150.2020.00011.5