New Spectroscopic Method for Estimation of Atorvastatin Tablets Using Hydrotropic Solubilization Technique

Dr. N. Usha Rani*, Keerthana. K, Mitha. M, Santhosh Kumar. S, Kotamma. K, Kiran. T.M.N.U.

Maharajah’s College of Pharmacy, Phool Baugh, Vizianagaram -535002. AP, India.

*Corresponding Author E-mail: nusharani.au@gmail.com

ABSTRACT:

A new, simple, accurate, cost effective and sensitive spectrophotometric method has been developed for the estimation of Atorvastatin (ATV) in tablet dosage form using 0.5 M sodium benzoate aqueous solution as a hydrotropic agent. Aqueous solubility of this model drug was enhanced 22 fold with 0.5M sodium benzoate solution without any interference with the method of analysis. Atorvastatin (ATV) showed maximum absorbance at a wavelength of 268 nm and Beer’s law was obeyed in the concentration range of 0.1-0.6 μg/ml. The method was validated for various parameters according to ICH guidelines. Hence it can be adopted for the routine analysis of Atorvastatin (ATV) in tablet dosage forms.

KEYWORDS: Atorvastatin, Hydrotropic solubilization, Method development and Validation.

INTRODUCTION:

Atorvastatin (ATV) chemically, (3R, 5R)-7-[2-(4-Fluorophenyl)-3-phenyl-4-(phenylcarbamoyl)-5-(propan-2-yl)-1H-pyrrol-1-yl]-3, 5-dihydroxyheptanoic acid (Fig. 1) is an inhibitor of 3-hydroxy-3-methylglutaryl-coenzyme A (HMG-CoA) reductase. This enzyme catalyzes the conversion of HMG-CoA to mevalonate, an early and rate-limiting step in cholesterol biosynthesis ^1-6.

The term hydrotropic agent was first introduced by Neuberg (1916) to designate anionic organic salts which, at high concentrations, considerably increase the aqueous solubility of poorly soluble solutes ⁷. Hydrotropy is a molecular phenomenon whereby adding a second solute (the hydrotrope) results in an increase in the aqueous solubility of poorly soluble solutes. Solubility enhancement is one of the advantages of hydrotropes. In the present study, an attempt is made to extend the definition of the term to include cationic and non-ionic organic compounds bearing the essential structural features of Neuberg's hydrotropes.

Hydrotropy is one of the solubility enhancement techniques which enhances solubility to many folds with use of hydrotropes like sodium benzoate, sodium citrate, urea, niacinamide etc

Literature survey reveals that very few methods were reported for the estimation of ATV by using sodium benzoate as hydrotropic agent. Hence the authors proposed to develop simple, accurate, precise, sensitive and economic method for the determination of ATV in bulk and in dosage forms ^8-16.

Fig. 1. Chemical structure of Atorvastatin

EXPERIMENTAL:

Instrument:

The Spectrophotometric analysis was carried out by using a double beam Agilent Cary-60 UV-visible Spectrophotometer with 1cm matched quartz cells.

Reagents and Chemicals:

Standard ATV was supplied as a gift sample by Dr. Reddy’s Laboratories, Pydibhimavaram. Analytical grade sodium benzoate and methanol were used for the proposed work. Millipore water was used were for the present study. Atorvastatin tablets were procured from local market.

Solubility studies:

Solubility of ATV was determined at (28±2)°C. An excess amount of the drug was taken into 25 ml volumetric flasks each containing 15 ml of different aqueous systems like double distilled water, sodium benzoate (0.5, 1, 2, 4, 6 M) using methanol as co-solvent and various combinations of Methanol and Water mixture (30:70, 40:60, 50:50). Solubility of ATV was enhanced by 22 folds in 0.5 M sodium benzoate. This enhancement of solubility was due to the hydrotropic solubilization phenomenon. The solubility enhancement ratio was determined by the following formula:

Solubility of drug in hydrotropic agent

Enhancement ratio=-----------------------------------------

Solubility of drug in Double distilled water

UV spectral studies:

In order to study the interaction between drug and hydrotropic agent, UV spectral studies of ATV were performed using different concentrations of hydrotropic solutions. Possible spectroscopic changes of ATV in the presence of hydrotropes were subsequently investigated.

Method Development:

Determination of Absorption maxima:

A quantity of Atorvastatin equivalent to 10 mg was accurately weighed and transferred to a 100 ml volumetric flask. To this, 5 ml of methanol was added as a co-solvent and 50ml of 0.5M freshly prepared sodium benzoate was added as hydrotropic agent. The volume was made up to the mark with HPLC grade water to give a 100 µg/ml stock solution which was diluted suitably to produce 0.4 µg/ml of ATV. This solution was scanned in the spectrum mode from 200-400 nm. From the spectrum of the drug obtained λmax of ATV was determined as 268 nm (Fig. 2).

Linearity Study:

The stock solution of ATV was prepared by dissolving 10mg of the drug in 5 ml of methanol followed by addition of 50 ml of 0.5 M sodium benzoate solution. The final volume was made up to 100 ml with double distilled water to get 100 μg/ml solution. From this, working standard solutions were prepared in the concentration range of 0.1 to 0.6 μg/ml. The absorbances of three replicates of each dilution were measured at 268 nm. The calibration curve was constructed by plotting concentrations against their respective absorbances. The corresponding regression equation was obtained and the values of slope-m, intercept-c and correlation coefficient (r²) were determined as shown in Table No. 1.

Table No. 1 Optimised parameters for the proposed method:

Parameter	Proposed Method
Wavelength	268 nm
Beer’s Law limit	01-0.6µg/ml
Regression equation	y = 1.582x + 0.014
Slope (m)	1.582
Intercept (c)	0.014
Correlation coefficient (r²)	0.999

Fig. 2. UV spectrum of Atorvastatin

Table No. 2 Determination of Atorvastatin in Tablets

Brand name of the Drug

Labeled amount of drug (mg)

Mean (±SD) amount found by the proposed method (n=3)

Mean (±SD) % labeled amount

LIPITOR

AZTOR

9.90

9.96

99.0± 0.0810

99.6 ± 0.1102

Assay of Tablet Formulation:

Twenty tablets of marketed formulation were accurately weighed and powdered. A quantity of powder equivalent to 10 mg of ATV was transferred to 100 ml volumetric flask and dissolved in 5 ml of methanol followed by addition of 50ml of 0.5 M sodium benzoate solution. The final volume was made up with distilled water and filtered through Whatmann filter paper No. 41. From the above solution 10ml was taken and diluted to 100ml with distilled water to get a 10 μg/ml solution. From this, 0.1 ml was taken and made upto 10ml with distilled water to get a concentration of 0.1 µg/ml ATV solution. The absorbance of this solution was found out at 268 nm. The mean of the absorbances of three such determinations was calculated and the drug content in the tablets was quantified using the regression equation obtained for the standard sample as shown in Table No. 2.

RESULTS AND DISCUSSION:

From the literature survey, it has been observed that very few spectroscopic estimations of Atorvastatin using hydrotropic solubilization phenomenon were reported. Hence the authors made an attempt to develop more accurate, sensitive and validated spectroscopic method for estimation of Atorvastatin using hydrotropic solubilization technique. The applicability of this method in determining the drug in commercial dosage forms was also studied. For this 0.5M sodium benzoate was used as the hydrotrope which enhanced the solubility of ATV by 22 fold. The estimation was carried out at 268 nm. The quantification of the drug in the formulations was carried out by calibration curve method using the regression equation y = mx+c where y is the absorbance and x is the concentration of the drug . The developed method was validated as per ICH Q2B (R1) guidelines.

Linearity and Range:

The linearity of the method was checked by measuring the absorbances of working standard solutions of Atorvastatin with concentrations ranging from 0.1-0.6 μg/ml. The absorbances of three replicates of each dilution were measured at 268 nm. The absorbances were found to be linear over the analytical range of ATV with regression coefficient value of 0.999 as shown in Fig. 3.

Fig. 3: Calibration curve of Atorvastatin

Table No. 3 Recovery Study of Atorvastatin

Sl.No	Label claim(mg)	Amount added (mg)	Amount recovered ± SD	% Recovery	%RSD
		8	17.84±0.105	99.11	0.588
1.	10	10	19.93±0.0556	99.65	0.278
		12	21.95±0.102	99.786	0.464

Table No. 4 Precision of the proposed method

Amount of ATV taken (µg/ml)	Amount of ATV (µg/ml) obtained
	Inter-day		Intra-day
	Mean	%RSD	Mean	%RSD
0.1	0.090±0.001	1.1	0.091±0.001	1.09
0.2 0.3	0.200±0.001 0.299±0.001	0.5 0.33	0.199±0.002 0.298±0.001	1.0 1.0

Table No. 5 Validation Parameters

S. No.	Validation Parameters	Results
1.	Linearity (r²)	0.999
2.	Range	0.1 – 0.6 µg/ml
3.	Precision (%RSD) Inter Day Intra Day	0.64 0.80
4.	Recovery Study (Average Mean Recovery)	99.51
5.	LOD	0.0019 µg/ml
6.	LOQ	0.0063 µg/ml

Table No. 6. Result of Robustness and Ruggedness Study

Parameter	Modification	% Recovery	Parameter	Modification	% Recovery
		Mean ± SD	Parameter		Mean ± SD
Robustness Study			Ruggedness Study
Solvent System Ratio (0.5M sodium Benzoate solution : Methanol :Water)	40:10:50 30:20:50 50: 5:45 35:15:50 50: 5: 45	98.91±1.258 98.28±1.046 99.00±0.015 98.50±1.205 99.59±0.081	Instrument Analyst	UV-60 UV-630 I II	99.91±0.374 98.45±0.541 99.67±1.190 98.14±0.872

Limit of Detection (LOD) and Limit of Quantitation (LOQ):

Three replicates of each working standard solution were measured and analyzed. The Limit of Detection (LOD) and Limit of Quantitations (LOQ) were then calculated by evaluating the signal to noise ratio of 3:1 and 10:1 respectively using the following equations

LOD = 3.3 σ/S

LOQ = 10 σ/S

The results are shown in Table No. 5.

Robustness:

The robustness of method was studied by changing the composition of solvent system. The results of robustness studies are reported in Table No. 6.

Ruggedness:

Ruggedness of the method was determined by using different models of UV spectrophotometers by different analysts. The results were found to be within the acceptance criteria as shown in Table No.6.

CONCLUSION:

The present paper describes application of hydrotropic solubilization phenomenon for the estimation of ATV in tablet dosage form by calibration curve method. The intra-day and inter-day drug variation studies by the proposed method showed low coefficient of variation indicating that the method is quite precise. The accuracy of proposed method was assessed by recovery studies. The recovery of ATV from the tablet dosage forms were in good agreement with the label claims, which suggested non interference of formulation additives and hydrotropic solubilizing agent in the estimation. Low values of LOD and LOQ indicate good sensitivity of the proposed method. Hence it can be concluded that the proposed spectroscopic method is accurate and reproducible for the analysis of ATV in tablet dosage forms.

ACKNOWLEDGEMENT:

We are grateful to Dr. Reddy’s Laboratories, Pydibhimavaram for providing a gift sample of Atorvastatin and also we acknowledge the laboratory facilities provided by Maharajah’s College of Pharmacy, Vizianagaram for this study.

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Received on 07.05.2015 Accepted on 01.06.2015

Asian J. Pharm. Ana. 5(2): April-June 2015; Page 100-104

DOI: 10.5958/2231-5675.2015.00016.2