Visible Spectrophotometric Determination of Abacavir Sulphate in Bulk Drug and Tablet Dosage Form

 

Prasada Rao C.H. 1*, Seshagiri Rao J.V.L.N.2, Dhachinamoorthi D.1, Lakshmi Aswini G.3 and Ashok K.1

1Dept. of Pharmaceutical Analysis, QIS College of Pharmacy, Ongole -523272, India.           

2Dept. of Pharmaceutical Analysis, University College of Pharmaceutical sciences, Andhra University, Vishakapatnam -530003, India.    

3Dept. of Pharmaceutical Analysis, Vagdevi College of Pharmacy, Gurazala -522415, India.

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

 

ABSTRACT:

A simple visible spectrophotometric method has been developed for the estimation of abacavir sulphate in bulk and tablet dosage form. This method is based on the diazotization of abacavir sulphate with nitrous acid to form diazotized abacavir sulphate, followed by its coupling with β-naphthol to form a red coloured chromogen which shows maximum absorption at 574.0nm and obeys Beer’s law in the concentration range of 5-20mcg/ml. This method was validated for precision, accuracy, ruggedness and robustness. Statistical analysis proves that the method is reproducible and selective for the estimation of said drug.

 

KEYWORDS: Visible spectrophotometry; Abacavir sulphate; β-naphthol; Validation.

 


 

INTRODUCTION:

Abacavir sulphate1 is chemically {(1S, 4R)-4-[2-Amino-6- (cyclopropylamino)-9H-purin-9-yl]-2-cyclopentene-1-methanol}. It is a nucleoside reverse transcriptase inhibitor with antiretroviral activity against HIV. It is administered alone or in combination therapy with other antiretrovirals. Survey of literature2-25 reveals that the drug is determined by using High Performance Liquid Chromatography and some other spectrophotometric methods which includes its estimation in biological fluids also.

 

The present study describes simple, sensitive, accurate, rapid and economical visible spectrophotometric method for the estimation of abacavir sulphate in bulk and its tablet dosage forms.

 

MATERIALS AND METHODS:

An analytical UV/Vis double beam spectrophotometer (model T 60) with 1 cm matched quartz cells was used for all spectral measurements. All the chemicals used in the investigation were of analytical grade. Authentic drug sample of abacavir sulphate was given as a gift sample by Hetero drugs Ltd., Hyderabad. Tablets of abacavir are procured from local market.

 

Working Standard Solution of abacavir sulphate:

Pure abacavir sulphate powder equivalent to 100 mg was accurately weighed and dissolved in 50 ml of methanol in a 100 ml volumetric flask and the volume was made up to 100 ml with distilled water (1 mg/ml). From this, a working standard solution containing 100 mcg/ml was prepared with distilled water.

 

Sample Preparation of abacavir sulphate:

Twenty tablets of abacavir sulphate each containing 300 mg were accurately weighed, average weight was determined and crushed into fine powder. An accurately weighed quantity of powder equivalent to 50 mg of abacavir sulphate was transferred into 50 ml volumetric flask and dissolved in 25 ml of methanol and sonicated for 5 mins. The solution was filtered through Whatmann filter paper no.41. The residue was washed with 5 ml portions of distilled water two times and the total volume of the filtrate was made up to 50 ml with distilled water (1 mg/ml). The final concentration was brought 100 mcg/ml with distilled water.

 

Method development:

Aliquots of standard solution of abacavir sulphate ranging from 0.5 to 2.0 ml (1 ml = 100 mcg) were transferred into a series of 10 ml volumetric flasks. To each flask, 1.0 ml of hydrochloric acid (2 N) and 1.0 ml of sodium nitrite (0.1% w/v) was added and a reaction time of 10 min at 0-5°C was given for the completion of reaction. Then 1.0 ml of alkaline β-naphthol solution (0.1% w/v in 2% aqueous NaOH) was added to each flask with gently shaking and after 10 min, the volume in each flask was made up to 10 ml with distilled water. The absorbances of red colored chromogen were measured at 574.0nm against the reagent blank. The colored chromogen was stable for 3 h. The amount of abacavir sulphate present in the sample solution was computed from the respective calibration curve (Fig.-1 and 2).

 

Fig. 01- Absorption spectrum of abacavir sulphate with β-naphthol.

 

Fig. 02- Calibration curve of abacavir sulphate with β-naphthol.

 

RESULTS AND DISCUSSION:

In the present investigation the λmax of the abacavir sulphate was found to be 574.0nm. Abacavir sulphate follows linearity in the concentration range of 5-20mcg/ml. Optimum conditions, optical characteristics and statistical data of the regression equation in the proposed method has been shown in table-1. Two brands of tablets were analyzed and amount of drug were determined by proposed method; it was in good agreement with the label claim (table-2). The proposed method was validated as per the ICH guidelines (table-3 to 6). The recovery studies were carried out by adding a known amount of drug to pre analysed sample at two different levels and the % recoveries were ranges from 99.86-101.08%, which shows the accuracy of method. Intra-day and Inter-day precision of the assay was determined by analyzing the drug sample using same concentration. The intra-day and inter-day % RSD values were calculated and lying in the range of 0.46-0.94%. Ruggedness of proposed method was studied with the help of two different analysts and results were evaluated by calculating the % RSD values; lying within the range of 0.32-0.39%.

 

Table 01: Optimum conditions, optical characteristics and statistical data of the regression equation in the proposed method

Parameter

Values

lmax (nm)

Beer’s law limits (mcg/ml)

Molar extinction coefficient (mol-1 cm-1)

Sandell’s sensitivity

(mcg/cm2-0.001 absorbance units)

Regression equation (Y*)

Slope (b)

Intercept (a)

Correlation coefficient(r2)

% RSD**

Limit of detection (mcg/ml)

Limit of quantitation (mcg/ml)

574

5-20

0.068 X104

…………

0.034

Y=0.0103C + 0.0008

0.0103

0.0008

0.9998

0.82

0.124

0.398

*Y= bC + a where C is the concentration of abacavir sulphate in mcg/ml and Y is the absorbance at the respective lmax.

**Average of five determinations.

 

Table 02:  Results of analysis of marketed formulations.

Brand

used

Label claimed (mg)

Amount found by proposed method (mg)

% label claim

% RSD*

Tab-a

Tab-b

300

300

299.67

300.06

99.89

100.02

0.64

0.47

*Average of five determinations.

 

Table 03: Results of recovery studies

Brand used

Label claimed (mg)

Mean assay value

Known amount of abacavir sulphate added

Mean % recovery ±%RSD*

Tab-a

 

Tab-b

300

 

300

99.89

 

100.02

10mg

20mg

10mg

20mg

99.86±0.34

100.06±0.83

99.97±0.28

101.08±0.96

*Average of five determinations.

 

Table 04:Results of intra day precision studies

Brand used

Label claimed (mg)

Amount found by proposed method(mg)

% label claim

% RSD*

Tab-a

Tab-b

300

300

299.88

299.46

99.96

99.82

0.46

0.82

*Average of five determinations.

 

Table 05: Results of inter day precision studies

Brand used

Label claimed (mg)

Amount found by proposed method(mg)

% label claim

% RSD*

Tab-a

Tab-b

300

300

299.58

298.86

99.86

99.62

0.68

0.94

*Average of five determinations.

 

 

Table 06: Results of ruggedness studies

Brand used

Label claimed (mg)

Normal condition

(Mean assay value)

Changed condition

(Mean assay value) ±%RSD*

Tab-a

Tab-b

300

300

99.89

100.02

99.64±0.39

99.43±0.32

*Average of five determinations.

 

CONCLUSION:

The proposed method is economic, sensitive, accurate, reproducible and useful for the routine determination of abacavir sulphate in tablet formulation.

 

ACKNOWLEDGMENT:

The authors are grateful to the management of QIS College of Pharmacy for providing the necessary facilities and support to carry out this work.

 

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Received on 21.08.2010          Modified on 18.09.2010

Accepted on 30.09.2010         © RJPT All right reserved

Research J. Pharm. and Tech. 4(2): February 2011; Page 234-236