Difference spectrophotometric method development for the estimation of Acyclovir

 

Pranali Gund1, Shrikrishna Baokar1*, Santosh Undare2

1Department of Pharmaceutical Chemistry, Shivnagar Vidya Prasarak Mandal’s College of Pharmacy, Malegaon (Bk), Tal- Baramati, Dist - Pune, Maharashtra, India – 413115.

2P.G. Department of Chemistry, Marathwada Shikshan Prasarak mandals Balbhim Arts, Science and Commerce College, Beed, Dist- Beed, Maharashtra, India-431122.

 

 

ABSTRACT:

A simple, precise and accurate difference spectroscopic method has been developed for the estimation of Acyclovir in bulk and in pharmaceutical dosage form. The proposed method is based on the principle that Acyclovir can exhibit two different chemical forms in basic and acidic medium that differ in the absorption spectra in basic and acidic medium. Since the drug was freely soluble in distilled water, a stock solution (1 mg/mL) was prepared with distilled water. Further dilution was made by using 0.1 M sodium hydroxide and 0.1 M hydrochloric acid separately. The maxima and minima in the difference spectra of Acyclovir were at 256 nm. Difference in absorbance between these maxima and minima was calculated to find out the amplitude. This amplitude was plotted against concentration. Beer’s law is valid in the concentration range of 1-5 μg/mL. As per ICH guidelines results of analysis were validated statistically and all the results were found satisfactory.

 

KEYWARDS: Acyclovir, difference spectroscopic method, sodium hydroxide, hydrochloric acid.

 

INTRODUCTION:

Acyclovir, chemically known as 9-[(2- hydroxyethoxy) methyl] guanine is a purine nucleoside analogue, active against herpes simplex virus type 1 and 2 and against viricella zoster virus. It inhibits enzyme thymidine kinase and interferes with DNA synthesis1,2. It is official in USP and BP. Survey of literature shows different UV-Vis Spectrophotometric method has been developed to determine different antiviral drug i.e. valacyclovir, gancyclovir, famcyclovir3,4,5. Several methods have been developed for spectrophotometric evaluation of acyclovir. The reported techniques for its estimation include solid phase extraction and HPLC6 and electroimmunoassay7 in serum and cerebrospinal fluid8. Reviewing the literature revealed that nothing has been published concerning the proposed method. These methods are based on difference spectrophotometric analysis by using acidic and basic conditions

 

EXPERIMENTAL:

Chemicals and reagents

Acyclovir pure drug was obtained from Agio Biopharma, Pune (Maharashtra) India. All chemicals were of analytical reagent grade purchased from S. D. Fine chemicals, Mumbai, India. 0.1 N Sodium hydroxide, 0.1 N Hydrochloric acid and double distilled water were used throughout the analysis.

Instrumentation:

A Schimadzu 1700 UV/VIS double beam spectrophotometer with 1cm matched quartz cells was used for all spectral measurements.

 

Preparation of stock solution

The standard Acyclovir (100 mg) was weighed accurately and transferred to volumetric flask (100 ml). It was dissolved properly and diluted with 0.1 N NaOH and 0.1 N HCl separately to obtain the final concentration of 1000 μg/mL and the resulting solution was used as working standard solution.

 

Difference spectroscopy9

The selectivity and accuracy of spectrophotometric analysis of sample containing absorbing interference may be markedly improved by the technique of difference spectrophotometric analysis. The essential feature of difference spectrophotometric assay is that the measured value is the difference in absorbance between two equimolar solutions of the analyte in different chemical forms which exhibit different spectral characteristics. For the Preparation and Analysis of sample solution, each tablet containing 300 mg of Acyclovir, 20 tablets were accurately weighed and. Drug equivalent to 100 mg was taken and treated in similar manner as that of standard. Aliquots of 0.2 - 1.2 ml (1 ml = 10 μg) were transferred into a series of 10 ml volumetric flasks. The volumes of each flask were adjusted to 10 ml with 0.1 N NaOH and 0.1 N HCl separately. The absorbance was measured at 256 nm in acidic and basic solution against reagent blank. For both the solutions, calibration curve was prepared by plotting concentration versus difference in absorbance and found to be linear in the concentration range of 1 – 5 μg/mL.

 

Preparation and Analysis of sample solution

Similarly the difference in absorbance of sample solution was measured and the amount of Acyclovir was determined from standard calibration curve. The differences in absorbance were calculated for the different concentration of the acidic and basic solutions and the concentrations of each component were obtained by analyzing the spectral data of the solutions. The recovery experiments were performed by adding known amounts of the drug to the pre-analyzed formulation and reanalyzing the mixture by proposed methods for both the solutions.

Table No. 1 Data for Linearity of Acyclovir in various medium

Sr. No.

Concn (μg/mL)

Acid 0.1 N HCL

Base 0.1 N NaOH

Difference

1

1

0.28

0.165

0.115

2

2

0.54

0.31

0.23

3

3

0.809

0.466

0.343

4

4

1.07

0.62

0.45

5

5

1.348

0.77

0.578

 

Mean

0.809

0.466

0.343

 

SD

0.421

0.240

0.181

 

RSD

0.520

0.515

0.528

 

%RSD

52.03

51.57

52.84

 

Cor.Coef

0.999

0.999

0.999

 

Intercept

0.009

0.009

-0.008

 

Slope

0.266

0.152

-0.0006

 

RESULTS AND DISCUSSION:

The optical characteristics such as Beer’s law limits, percent relative standard deviation and % range of error were found to be within the limits and satisfactory. All of the analytical validation parameters for the proposed method were determined according to ICH guidelines. After selecting Beers limit linearity was checked and all the results were found within limit (Table No. 1). The method was found to provide high degree of precision and reproducibility. The recovery studies showed that the results were within the limit indicating no interference. Final optical characteristics for acyclovir are mentioned in table No. 2. The % recovery study was carried out for marketed formulation, which is tabulated in Table No. 3. The proposed method is simple, sensitive, accurate and precise and can be successfully employed for the routine analysis of the Acyclovir in Pharmaceutical formulations.

 

Table No. 2 Optical Characteristics of Acyclovir by Differential Spectrophotometry

λ Max (nm)

256

Beers Limit

1 - 5 μg/mL

Correlation coefficient

0.999

Regression Equation

Y = 0.114 x

Slope (m)

-0.0006

Intercept (c)

-0.008

*Y=mx+c, where x is the concentration in (μg/ml) and Y is absorbance unit (ΔA)

 

 

Table No. 3 Analysis of marketed formulation

Method

Label Claim

Amount Estimated

% RSD

SD

% Recovered

Standard Error

Acyclovir

400 mg

391

0.011

0.035

97.75

0.024

 

 

CONCLUSION:

The proposed dissolution method is more precise, accurate and rapid than most of the reported methods and characterized by instrumental simplicity, cost effective in the use of reagents and time. No interference of excipient and coating material was found in this method. All aliquot solutions prepared were stable for 12 hrs. Hence this method can be employed for routine evaluation and validation of Acyclovir tablet during the small scale, pilot scale and large production scale batches.

 

ACKNOWLEDGEMENTS:

The authors are grateful to the Principal and Management of SVPM’S college of Pharmacy, Malegaon for their continuous support, encouragement and for providing the necessary facilities.

 

REFERENCES:

1.       Sweetman, S.C., Eds., In; Martindale, 33rd Edn., The Pharmaceutical Press., Inc., London, 2002, 612.

2.       G.H. Frederick, In: L.L. Brunton, J.S. Lazo, K.L. Parker (Eds.) Goodman & Gilman’s The Pharmacological Basis of Therapeutics, 11th ed. (McGraw-Hill Medical Publishing Division, NewYork, 2006).

3.       .M. Ganesh, C.V. Narasimharao, A. Saravana Kumar, K. Kamalakannan, M. Vinoba, H.S. Mahajan, T. Sivakumar, Spectrophotometric determination and validation of acyclovir, E-J. Chem, 2009, 6(3), 814-818.

4.       P.S. Sarsambi, A. Sonawane, S.M. Malipatil, B. Hiremath, A. Faheem,  Spectrophotometric determination and validation of acyclovir in tablet dosage form , Int. J. PharmTech Res., 2010, 2(2), 1264-1268.

5.       J.A.J. Nesalin, C.J.G. Babu, G.V. Kumar, T.T. Mani, E-J. Chem, 2009, 6(3), 780-784.

6.       Svensson, J.O., Brkhlt, L., Sawe J., J. Chromatogr. B Biomed. Sci. Appl . 1997, 690, 363.

7.       Laurell, C.B. Anal. Biochem., 1996 , 15, 45.

8.       Lycke, J., Malmestrom, C., Stahle, L., Antimicrob. Agents and Chemother., 2003, 47, 2438.

9.       V. Amudhavalli, K. S. Lakshmi, et.al. Difference Spectrophotometric Estimation of Abacavir sulphate in Bulk and Tablet dosage form, J. Chem. Pharm. Res., 2011, 3(3):119-122..

 

Received on 05.01.2013

Modified on 20.03.2013

Accepted on 31.03.2013     

© A&V Publication all right reserved

Research Journal of Pharmaceutical Dosage Forms and Technology. 5(2): March- April, 2013, 88-90