INTRODUCTION:

Aceclofenac and Paracetamol are available in tablet dosage form in the ratio of 1:5. Aceclofenac is a non steroidal anti-inflammatory drug with good analgesic and anti-rheumatic properties1. Chemically it is [[[2-[(2, 6-Dichlorophenyl) amino] phenyl] acetyl] oxy] acetic acid. It is used in various pain conditions like rheumatoid arthritis, osteoarthritis and ankylosing spondylatis.^1-4 It is official in British Pharmacopoeia.⁴Paracetamol is N-(4 – hydroxyphenyl) acetamide has analgesic and antipyretic activity. Paracetamol is official in Indian Pharmacopoeia and British Pharmacopoeia.⁴ B.P. suggests a potentiometric assay method for aceclofenac in bulk drugs. The I.P. and B.P. both suggest titrimetric and UV spectrophotometric assay method for paracetamol in bulk and tablet formulations.

Several analytical techniques like titrimetric^4,5, colourimetric⁷, spectroflurimetric⁷, densitometric^8,9, HPLC^9-11, RP–HPLC^12,13, spectrophotometric^14-17 and stripping voltametric¹⁸ have been reported for assay of aceclofenac and paracetamol. This paper describes two simple, rapid, accurate, reproducible and economical methods for the simultaneous estimation of aceclofenac and paracetamol in tablet formulations using simultaneous equation and absorbance ratio methods.

MATERIALS AND METHODS:

Materials:

Spectral runs were made on a Shimadzu UV-Visible spectrophotometer, model- 1700 (Japan) was employed with spectral bandwidth of 1 nm and wavelength accuracy of ± 0.3 nm with automatic wavelength corrections with a pair of 10 mm quartz cells. Glasswares used in each procedure were soaked overnight in a mixture of chromic acid and sulphuric acid rinsed thoroughly with double distilled water and dried in hot air oven. Aceclofenac and Paracetamol reference standards were kindly provided by Aristo Pharma. Ltd. Mumbai (M. S.) while the pharmaceutical preparations of combination of Aceclofenac and Paracetamol used for analysis. Methanol of analytical reagent grade was purchased by Loba Chemie Pvt. Ltd. (India). All the solutions were protected for light and were analyzed on the day of preparations.

Selection of common solvent:

Methanol and distilled water (20:80) was selected as common solvent for developing spectral characteristics of drug. The selection was made after assessing the solubility of both the drugs in different solvents.

Preparation of Standard Drug Solution:

Standard stock solutions containing Aceclofenac (AC) and Paracetamol (PC) were prepared individually by dissolving 10 mg of Aceclofenac and 10mg Paracetamol separately in 20 ml of methanol. It was then sonicated for 10 minutes and the final volume of both the solutions were made up to 100 ml with distilled water to get stock solutions containing 100mcg/ mL each of AC and PC in two different 100 ml volumetric flasks.

Table 1: Linear regression analysis of calibration curves with their respective absorptivity values.

Parameter	Method I		Method II
Parameter	AC	PC	AC	PC
Beer’s law limit (mcg/ml)	5-40	2-20	5-40	2-20
Coefficient of Correlation	0.999894	0.999721	0.999934	0.999912
Molar absorptivity (lit/mole/cm)	8861.8330	9135.7055	8387.2171	3371.0914
Sandal's sensitivity (mcg/Sq.cm/0.001)	0.039968	0.016547	0.04223	0.044843
Slope	0.02502	0.060433	0.02368	0.02230
Intercept	0.0053	0.0050	0.001528	0.006506

Table 2: Results of analysis of laboratory samples.

Analyte	Method I		Method II
Analyte	AC	PC	AC	PC
% Conc. Estimated (Mean ± S.D.)	101.68 ±0.006245	98.87 ±0.005	100.25 ±0.001528	100.49 ±0.006506
Coefficient of variance	0.9996	0.9997	0.9999	0.9998

Table 3: Results of analysis of tablet formulations.

Method	Drug	Label Claim	% Label Claim found	%Coefficient of variance (%R. S. D.)	% Recovery*
I	AC	100	100.90	0.9389	101.68
I	PC	500	102.30	0.91743	99.93
II	AC	100	99.30	0.43191	100.41
II	PC	500	99.98	3.1998	100.55

* Average of three determinations; R.S.D.; Relative Standard Deviation.

Table 4: Results of intermediate precisions.

Day	Method I		Method II
	% Label claim estimated (Mean ± % R.S.D.)		% Label claim estimated (Mean ± % R.S.D.)
	AC	PC	AC	PC
Intraday	101.25 ± 1.0267	100.85 ± 0.9854	100.36 ± 0.6826	98.99 ± 2.1974
Interday	99.19 ± 0.8732	103.44 ± 0.5421	102.02 ± 0.7803	100.72 ± 0.9640

Determination of Absorption Maxima:

By appropriate dilution of two standard drug solutions with methanol diluted with distilled water (20:80), solutions containing 10mcg/ml of AC and 10 mcg/ml of PC were scanned separately in the range of 200- 400 nm to determine the wavelength of maximum absorption for both the drugs. AC and PC showed absorbance maxima at 274 nm (λ₁) and 245 nm (λ₂) respectively. The overlain spectra showed λ max of both drugs and also isoabsorptive points at 267.5 nm (Fig. 1).

Method I (Simultaneous equation method):

Two wavelengths selected for the method are 274 nm and 245 nm that are absorption maximas of AC and PC respectively in methanol diluted with distilled water. The stock solutions of both the drugs were further diluted separately with methanol diluted with distilled water to get a series of standard solutions of 5-25mcg/mL concentrations for AC and 3-15mcg/ml for PC respectively. The absorbances were measured at the selected wavelengths and absorptivities (A 1%, 1 cm) for both the drugs at both wavelengths were determined as mean of three independent determinations. Concentrations in the sample were obtained by using following equations-

A₁ ay₂ – A₂ ay₁

Cx = ………….Eq. (i)

ax₁ay₂ - ax₂ay₁

A₁ ax₂ – A₂ ax₁

Cy = …............Eq. (ii)

ay₁ax₂ - ay₂ax₁

Where, A₁ and A₂ are absorbances of mixture at 274 nm and 245 nm respectively, ax₁ and ax₂ are absorptivities of AC at λ₁ and λ₂ respectively and ay₁ and ay₂ are absorptivities of PC at λ₁ and λ₂ respectively. Cx and Cy are concentrations of AC and PC respectively.

Method II (Absorbance ratio or Q-analysis method):

From the overlain spectrum of AC and PC, two wavelengths were selected one at 267.5 nm which is the isoabsorptive point for both the drugs and the other at 274 nm which is λmax of AC. The absorbances of the sample solutions prepared in a similar manner as in the previous method were measured and the absorptivity values for both drugs at the selected wavelengths were also calculated. The method employs Q values and the concentrations of drugs in sample solution were determined by using the following formula,

For AC

Q₀ – Q₂ A

C₁ = X

Q₁–Q₂ a₁

For PC

Q₀ – Q₁ A

C₂ = X

Q₂–Q₁ a₂

Where,

Absorbance of sample at 274 nm

Q₀ =

Absorbance of sample at 267.5 nm

Absorptivity of AC at 274 nm

Q₁ =

Absorptivity of AC at 267.5nm

Absorptivity of PC at 274 nm

Q₂ =

Absorptivity of PC at 267.5nm

A = Absorbance of sample at isoabsorptive point,

a₁ and a₂ = Absorptivities of AC and PC respectively at isoabsorptive point.

Application of the Proposed Method for the Determination of Aceclofenac and Paracetamol in Tablets:

Marketed tablet formulation containing aceclofenac 100 mg and paracetamol 500mg was analyzed using this method. From the 20 tablets, an amount equivalent to 100mg of aceclofenac and 500 mg of paracetamol was weighed and from that the amount equivalent to 10mg of aceclofenac and 50mg of paracetamol was weighed and dissolved in 20ml of methanol and sonicated for 10 minutes. Then the volume made upto 100ml with distilled water to get a stock solution containing 100mcg/mlof aceclofenac and 500mcg/ml of paracetamol. Then the solution was filtered through Whatman filter paper no. 41. Appropriate aliquots of aceclofenac and paracetamol within the Beer’s law limit were taken. The absorbances of resulting solutions were measured at 274 nm and 245 nm. The concentration of aceclofenac and paracetamol present in the sample solution was calculated by using the equation generated from calibration curve of respective drugs.

In Method I, the concentration of both AC and PC were determined by measuring the absorbance of the sample at 274 nm and 245 nm. Values were substituted in the respective formula to obtain concentrations. For Method II, the concentration of both AC and PC were determined by measuring absorbance of the sample at 274 nm and 267.5 nm and values were substituted in the respective formula to obtain concentrations. Results of tablet analysis are shown in Table 3.

VALIDATION:

The method was validated according to ICH Q2B guidelines for validation of analytical procedures in order to determine the linearity, sensitivity, precision and accuracy for the analyte.

Accuracy:

To ascertain the accuracy of the proposed methods, recovery studies were carried out by standard addition method at three different levels (80%, 100% and 120%). Percent recovery for AC and PC, by both the methods, was found in the range of 99.93% to 101.68%.

Linearity:

The linearity of measurement was evaluated by analyzing different concentration of the standard solution of AC and PC. For simultaneous equation method and Q analysis, the Beer- Lambert’s concentration range was found to be 5-40 mcg/ml for AC and 2-20mcg/ml PC.

Fig.1. Overlain spectra of Aceclofenac (AC) and Paracetamol (PC) in methanol diluted with distilled water.

Precision:

Precision was studied to find out intra and inter-day variations in the test method of AC and PC. Calibration curves prepared in medium were run in triplicate in same day and for three days. %RSD (relative standard deviation) were calculated which should be less than 2 %. The results are tabulated in Table 4.

RESULTS AND DISCUSSION:

The overlain spectra of AC and PC exhibit λ max of 274 nm and 245 nm for AC and PC respectively which are quite separated from each other. Additionally one isoabsorptive point was observed at 267.5 nm. This wavelength was selected for simultaneous estimation of AC and PC for Q value analysis and it is assume to be sensitive wavelength. Standard calibration curves for AC and PC were linear with correlation coefficients (r) values in the range of 0.9996 - 0.9999 at all the selected wavelengths and the values were average of three readings with % relative standard deviation in the range of 0.001528 – 0.006506. The calibration curves were repeated three times in a day and the average % RSD was found to be 0.8546 for AC and 1.5914 for PC, similarly the method was repeated for three different days and average % RSD was found to be 0.8267 for AC and 0.7531 for PC. The accuracy of the method was conformed by recovery studies from tablet at three different levels of standard additions; recovery in the range of 98 – 102% justifies the accuracy of method.

CONCLUSION:

The most striking feature of this method is its simplicity and rapidity, non- requiring- consuming sample preparations such as extraction of solvents, heating, degassing which are needed for HPLC procedure. These methods can be employed for routine quality control analysis. The described methods give accurate and precise results for determination of aceclofenac and paracetamol mixture in marketed formulation.

ACKNOWLEDGEMENTS:

The authors are thankful to the Principal and Head of Pharmaceutical Chemistry Department, Govt. College of Pharmacy, Karad (M.S.) for providing necessary facilities and Aristo Pharma. Ltd. Mumbai for providing the gift sample of aceclofenac and paracetamol.

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Received on 23.09.2009 Modified on 19.11.2009

Research J. Pharm. and Tech. 3(1): Jan.-Mar. 2010; Page 247-250