INTRODUCTION:

Carvedilol is a non-selective and β-adrenergic antagonist with no intrinsic sympatomimetic activity and is widely used to treat essential hypertension and angina pectoris. (1-3) Carvedilol is also indicated for the treatment of mild to severe chronic heart failure, Left ventricular dysfunction following myocardial infarction in clinically stable patients and hypertension.Beta‐blockers affect the heart and blood circulation(4-7). Carvedilol is a racemic lipophillic aryoxypropanolamine that blocks α1‐ and ß‐adrenergic receptors(8). Carvedilol significantly decreases systemic blood pressure, pulmonary artery pressure, and pulmonary capillary wedge pressure because of the vasodilatation that occurs with blocking of 1‐receptors(9-13). Blocking of ß‐receptors reduces the heart rate and increases diastolic filling time. Carvedilol blocks the binding to those receptors, which both slows the heart rhythm and reduces the force of the heart's pumping (14).

Chemically, carvedilol (2 RS) - 1- (9H-Carbazol- 4yloxy)- 3- [[2- (2- methoxy phenoxy) ethyl] amino] propan- 2- ol is a non selective β and α1 adrenergic receptor blocking agent (15-16). It also has multiple spectrums of activities such as antioxidant property, inhibition of smooth muscle proliferation and calcium antagonistic blocking activity (17).

Fig. 1 Structure of carvedilol

Analytical methods play a vital role in drug development process including preformulation and formulation studies, stability studies, quality control testing and in quality assurance programmes. Analytical testing of a pharmaceutical product is necessary to ensure its stability, safety and efficacy. Such testing needs an analytical method with reliable result adequate for intended purpose.

Literature survey revealed that several methods such as RP-HPLC (18-19), spectrophotometry (20), gas chromatography-mass spectrometry (21), extractive spectrophotometric (22), liquid chromatography (23) and HPLC (24-26) in biological samples have been reported. A new simple, sensitive UV spectrophotometric method was developed for estimation of carvedilol in pharmaceutical dosage forms without the necessity of sample pretreatment. The proposed method was developed and validated according to the validation parameters.

MATERIALS AND METHODS:

Materials

Carvedilol was obtained as a gift sample from Cipla Ltd., Kurkumbh, India. All analytical grade chemicals and solvents were supplied by S.D. FineChemicals, Mumbai, India. Distilled water was used to prepare all solutions. Freshlyprepared solutions were always employed. Different brands of tablets carvedilol were purchased from local market.

Equipment

The UV-Visible Spectrophotometer (Jasco-V630) with data processing system was used. The sample solution was recorded in 1cm quartz cells against solvent blank over the range 200-400 nm. A Citizen electronic analytical balance was used for weighing the sample. An ultrasonicator bath (PCI Analytics Pvt. Ltd.) was used for sonicating the tablet powder.

Development of method

Accurately weighed 10 mg of carvedilol was solubilized by 10 ml of methanol in a 100 ml volumetric flask, and phosphate buffer pH 6.8 was added to make up the volume so as to give stock solution of concentration 100 μg/ml. The standard solutions were diluted with phosphate buffer pH 6.8 to obtain various dilutions (10, 15, 20, 25, 30, 35μg/ml) in standard volumetric flasks (10 ml). The dilutions were scanned in the wavelength range of 200-400 nm. The λ_max of carvedilol was found at 284 nm. The linear relationship was observed over the range of 10-35 μg/ml. Absorbances were noted at 284 nm against pH 6.8 phosphate buffer as a blank.A calibration graph of the absorbance versus the concentration of the drug was plotted and represented in figure 1.

Figure 1. Calibration curve of carvedilol in pH 6.8 phosphate buffer.

Procedure for dosage forms

For analysis of commercial formulations, twenty tablets were taken and powdered. Tablet powder equivalent to 10 mg of carvedilol was dissolved in small quantity of methanol into a 100 mL volumetric flask and final volume was made up to 100 ml with pH 6.8 phosphate buffer and sonicated for 30 minutes. Then the absorbance of the solution (after suitable dilution) was measured at 284 nm using UV/visible spectrophotometer (Jasco-V630) against pH 6.8 phosphate buffer as a blank. The percentage drug content was calculated with the help of calibration curve (n=3).

Validation of the proposed method

The proposed method was validated according to the International Conference on Harmonization (ICH) guidelines (27, 28).

Linearity (Calibration curve)

The developed method validates as per ICH guidelines. The plot of absorbance verses concentration is shown in figure 1. It can be seen that plot is linear in the concentration range of 5-35 μg/ml with correlation coefficient (r²) of 0.997.

Precision (repeatability)

Intraday and interday precision was determined by measurement of theabsorbance for three times on same day and on three different day. The relative standard deviation for replicates of sample solution was less than 2 % which meet the acceptance criteria for established method.The obtained results are presented in table 1.

Table 1 Precision for proposed method.

Concentration (μg/ml)	Absorbance mean	Standard deviation	% Relative standard deviation
Intraday precision (n=3)
10	0.194	0.002517	1.29
20	0.447	0.006658	1.48
30	0.698	0.003055	0.43
Interday precision (n=3)
10	0.192	0.003914	1.98
20	0.452	0.007000	1.54
30	0.697	0.004726	0.57

Accuracy (recovery study)

Recovery studies were carried out by adding a known quantity of pure drug to the preanalysed formulations and the proposed method was followed. From the amount of drug found, percentage recovery was calculated as per ICH guidelines. The data were presented in table 2.

LOD and LOQ

The limit of detection (LOD) and limit of quantification (LOQ) of the drug were separately determined based on method of the intercept and the average value of slope. (i.e. 3.3 for LOD and 10 for LOQ) ratio using the followingequations designated by International Conference on Harmonization (ICH) guideline.

LOD = 3.3 σ/S

LOQ = 10 σ/S

Where, σ = the standard deviation of the response.

S = slope of the calibration curve.

RESULTS AND DISCUSSION:

Beers law is obeyed over the concentration range of 10 - 35 μg/mL, using regression analysis the linear equation y = 0.024x - 0.045 with a correlation coefficient of 0.997. The limit of detection was found to be0.3796 μg/mL. The limit of quantification was found to be 1.15 μg/mL. The percentage purity of carvedilol in Brand I, Brand II and Brand III was found to be 99.04 %, 96.64% and 100.96% respectively. Precision was calculated with intra and interday variation. Recovery study was performed on formulations and % RSD was found. The optical parameters such as Beer’s law limits, slope, and intercept values were calculated and given in table 3. Method was validated for accuracy and precision. The accuracy of method was proved by performing recovery studies in commercially available formulations. The results were given in table 2 and shows relative standard deviation of less than 2 % was observed for analysis of three replicate samples, indicating precision and reproducibility. The percentage recovery value indicates that there is no interference from the excipients present in the formulation. The applicability of the proposed method for the assay of carvedilol in tablet formulation was examined by analyzing commercial formulations and the results are tabulated in table 4. The result obtained were good agreement with the label claims. The results of analysis of the commercial tablets and the recovery study of the drug suggested that there is no interference from any excipients such as starch, lactose, magnesium stearate etc. which are commonly present in tablets.

Table 3 Optical parameters for determination of carvedilol

Sr. No.	Parameters	Data
1	λ-Max	284 nm
2	Beer’s law limit	10 - 35 μg/mL
3	Regression equation	y = 0.024x - 0.045
4	Correlation coefficient	R² = 0.997
5	Slope	0.024
6	Intercept	0.045
7	Limit of detection	0.3796 μg/mL
8	Limit of quantification	1.15 μg/mL

Table 4 Results of assay

Formulation	Label claim, mg/tablet	*Amount found, mg/tablet**	% Amount found	% C.V.
Brand I	6.25	6.19 ±0.053	99.04	0.856
Brand II	6.25	6.04±0.040	96.64	0.669
Brand III	6.25	6.31 ±0.012	100.96	0.200

*Mean of three determinations.

CONCLUSION:

The simple spectrophotometric method for determination of carvedilol have been developed and validated as per ICH guidelines. The developed method is found to be sensitive, accurate and reproducible and can be used for the routine quality control analysis of carvedilol in bulk and pharmaceutical formulations.

ACKNOWLEDGMENT:

The authors are thankful to M/s Cipla Pharmaceuticals Ltd., Kurkumbh, India for providing gift sample of carvedilol. Authors are delighted to say thank you to Prof. Dr. R. B. Saudagar, Principal and also management of KCT’S R. G. Sapkal College of Pharmacy, Anjaneri, Nashik for support and providing necessary facilities to carry out the research work successfully.

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Received on 19.07.2013 Modified on 01.08.2013

Asian J. Research Chem. 6(10): October 2013; Page 956-959

Sr. No.	Label claim, mg/tablet	Amount of standard added, mg	Total amount recovered, mg	% Recovery	Standard deviation	% relative standard deviation
1	6.25	5	11.41	101.42	0.002645	0.96
2	6.25	10	16.05	98.76	0.002647	0.68
3	6.25	15	21.98	103.43	0.003214	0.60