Estimation of Nadolol in Bulk and Tablets by Area under Curve and First Order Derivative Spectrophotometry

 

Hemant K. Jain^*, Niraj K. Muley, Neha N. Deshpande

Department of Quality Assurance Techniques, STES’s Sinhgad College of Pharmacy, Vadgaon (Bk.), Pune-411041, Maharashtra, India.

 

ABSTRACT:

Two simple, precise, rapid, accurate and economical spectrophotometric methods have been developed for determination of Nadolol in bulk and tablets. First method was based on measurement of area under curve for spectrum in a wavelength range between 267.6 – 274.6 nm and second method involved first order derivative spectrophotometry at a wavelength 281.6 nm. These methods have been validated as per ICH guidelines. The linearity for first method was found in the concentration range of 80-180 μg/ml and the value of correlation coefficient (R²) was found to be 0.9969. Linearity of second method was found in 80-180 μg/ml concentration and the value of correlation coefficient (R²) was found to be 0.9988. The percent assay values obtained by both methods were found within acceptance limits. Percent R.S.D for precision study by both methods were also found to be satisfactory indicated reproducibility of the methods. The values of % R.S.D were found to be satisfactory suggested both methods were precise. The accuracy of both methods was assessed by recovery studies and % recovery values were found within acceptance criteria. Thus both methods can be applied for routine analysis of Nadolol.

 

 

 

INTRODUCTION:

Nadolol (NAD) is a non-selective beta blocker used to treat hypertension and angina pectoris as antihypertensive agent. It has a preference for beta-1 receptors, which are predominantly located in the heart, thereby inhibiting the effects of catecholamine and causing a decrease in heart rate and blood pressure ^[1]. This drug is official in United States Pharmacopeia ^[2]. It is chemically 2,3-Naphthalenediol, 5-[3-[(1,1-dimethylethyl)amino]-2-hydroxypropoxy]-1,2,3,4 tetrahydro, cis-1-(tertbutylamino)-3-[(5,6,7,8-trahydro-cis-6,7-dihydroxy-1-naphthyl)oxy]-2-propanol[1] (figure 1).

 

Fig. 1: Chemical structure of NAD

 

Literature survey reveals that various visible Spectrophotometric methods ^[3-4] and HPLC methods ^[5-7] have been reported for estimation of NAD. Reported visible methods involved the chemical reactions with the reagents. However, no simple UV spectrophotometric method has not been reported yet. Therefore, the objective of this work was to developed simple, precise, accurate and economical UV spectrophotometric methods for estimation of NAD in pharmaceutical formulations. 

 

MATERIAL AND METHODS:

Instrumentation and Apparatus:

Shimadzu UV-1800 UV-Visible spectrophotometer with two matched quartz cells and UV probe software was used for the work. Shimadzu ATY224 a single pan electronic balance, Biosystems ultrasonic cleaning bath sonicator and the calibrated volumetric glassware’s (Borosil) were used in this study.

 

Chemicals and Reagent:

The gift sample of nadolol was supplied by Lupin Research Park, Pune. Commercially available tablets Corgard^® 80 mg was procured from local market. AR grade of methanol was obtained from Merck India Ltd., Mumbai used as co-solvent. Double distilled water (DDW) was prepared in house using Easy Still 2000, Infusil India Pvt. Ltd. and used as main solvent.

 

Preparation of standard stock solution and working standards:

Standard stock solutions of NAD was prepared by dissolving 100 mg of drug in 100 ml of methanol to get standard stock solution of 1000 μg/ml and this stock solution was further diluted with DDW to get various concentrations between 80-180 μg/ml. Two methods have been used for the analysis of NAD.

 

Method I: Area under Curve

This method involves calculation of integrated value of absorbance with respect to wavelength in indicated range. Area calculation processing item calculates the area bounded by the curve and horizontal axis ^[8]. Here horizontal axis represents baseline.

 

Whereas, α is area of portion bounded by curve data and a straight line connecting the start and end point, β is area of portion bounded by a straight line connecting the start and end point on curve data and horizontal axis, λ1 and λ2 are wavelengths representing start and end point of curve region. In this study area under curve was integrated between wavelength ranges from 267.6 to 274.6 nm (Figure 2). The calibration curve was prepared between concentrations and their respective area.

 

Fig. 2:  Representative area under curve of UV Spectrum in wavelength range from (267.6 to 274.6 nm)

 

Method II: First Order Derivative Spectroscopy

Solutions of NAD were scanned in the spectrum mode from 400 to 200 nm. The first order derivative spectrum (figure 3) was obtained by data processing mode from this spectrum ^[9]. Derivative spectra of all working standards (80-180 μg/ml) were obtained ^{[10, 11]}. The calibration curve was performed between concentration and dA/ dλ.

       

Fig. 3:  First order derivative spectrum of NAD

 

Assay of tablets:

Twenty tablets of NAD were accurately weighed and average weight of a tablet was calculated. These tablets ware crushed and powdered. Amount of powder equivalent to 100 mg of NAD was dissolved in 20 ml of methanol and the final volume was made up to 100 ml with water. This solution was filtered through whatmann filter paper no. 41 and first few ml of filtrate was discarded. This solution was further diluted in concentration range between 80-180 μg/ml. Results of analysis of tablets by both methods are shown in Table 1.

 

Table 1: Results of Estimation of Nadolol in Tablets.

Method	Label claim (mg/tab)	% Label claim estimated (Mean ±S.D.)*	%R.S.D.
Method I	80	99.56± 0.002	1.438
Method II	80	98.94±0.003	1.361

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

 

Method Validation:

The proposed methods was validated according to ICHQ2 (R1) guideline ^[12]. Linearity, precision, accuracy, robustness, LOD and LOQ parameters were performed.

 

Linearity:

Working standards were suitably prepared between 80-180 μg/ml. Linearity was determined by plotting the curve between concentrations and corresponding values of area in method I. Similarly, Linearity was determined by plotting the curve between concentrations and corresponding values and dA/dλ in method II. Calibration curves of NAD by method I and method II are shown in Fig. 4 and 5, respectively. The optical characteristics of NAD are shown in table 2.

 

Table 2: Optical characteristics of NAD

Sr. No.	Parameters	Method I	Method II
1.	Wavelength/Wavelength range (nm)	267.6-274.6	281.6
2.	Concentration range for linearity (µg/ml)	80-180	80-180
3.	Correlation coefficient (r)	0.996	0.998
4.	Slope (m)	0.0015	-0.0002
5.	Intercept	0.0062	-0.0004

 

 

Fig. 4: Calibration curve of NAD by method I

 

Fig. 5: Calibration curve of NAD by Method II

 

 

Table 3: Results of Recovery Studies

Level	Standard drug added in  sample solution (µg/mL)		%Recovery ± S.D.*		Percent R. S.D.
	Method I	Method II	Method I	Method II	Method I	Method II
80%	96	96	98.00±0.905	99.50±0.404	0.962	0.427
100%	120	120	99.04±0.718	98.87±0.215	0.604	0.181
120%	144	144	98.08±0.209	99.06±0.841	0.148	0.589

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

 

 

Accuracy:

The accuracy of the methods were determined by calculating percent recovery of the drug by standard addition method. Percent recovery of NAD was determined at three different level 80%, 100% and 120% of the target concentration in triplicate for both methods. The results of accuracy study are shown in table 3.

 

Precision:                                                                     

The repeatability study was carried out with six concentrations of NAD. The intraday and interday precision studies were carried out with three concentrations of NAD with three replicates. The values of percent relative standard deviation was calculated. The methods were precise and the % R.S.D. values were within acceptable limit shown in Table 4.

 

 

Table 4: Results of Repeatability, Intraday and Interday Precision

Precision	Method I		Method II
	Amount (µg/ml)	% R.S.D.	Amount (µg/ml)	%R.S.D.
Repeatability (n=6)	120	0.119	120	0.891
Intra-day  Precision (n=3)	100	1.242	100	1.246
	120	0.871	120	0.618
	140	0.611	140	0.652
Inter-day  Precision (n=3)	100	0.626	100	0.666
	120	0.540	120	0.416
	140	0.510	140	1.139

 

Robustness:

Robustness study was carried out by change in wavelength for determination of robustness of methods and the respective absorbance was recorded. The result of robustness study is presented in Table 5.

 

Table 5: Results of Robustness

Change in wavelength	Amount (µg/ml)*	% R.S.D.
		Method I	Method II
270.6 nm	120	0.686	0.392
272.6 nm	120	1.004	0.587

* Average of three determinations =3; R.S.D. = Relative Standard Deviation

 

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

Six sets of concentrations (80-180 μg/ml) were prepared and measured the values of area under curve and dA/dλ were determined its absorbance. Calibration curves were plotted for each set using both methods. The standard deviation of the Y-intercept and average slope of the calibration curve was used to calculate LOD and LOQ using following formulae.

 

Where, SD is standard deviation of y-intercepts of the calibration curves; S is mean slope of six calibration curves.

 

The LOD and LOQ determination is given in the table 6.

 

Table 6: Results of LOD and LOQ determination

*Obtained by Y-intercepts of the calibration curves

 

 

RESULTS AND DISCUSSION:

The values of correlation coefficients obtained by both methods (Table 2) demonstrated the good relationship between response and concentrations. Therefore, the developed methods was linear in concentration range of 80-180 μg/ml of drug. Percent estimation values in assay study (Table 1) of commercial tablets were found within acceptance criteria. Accuracy of proposed methods was ascertained by recovery studies. Percent recovery for NAD and values of R.S.D. obtained by both methods (Table 3) were found satisfactorily indicating the accuracy of both the methods (Table 3). Percent R.S.D. for Intraday and Interday precision was found to be 0.908 and 0.559 for Method I and 0.838 and 0.740 for Method II (Table 4). This study indicates good precision. The values of percent R.S.D. in robustness study (Table 5) was found to be within acceptance criteria which showed the reliability of both methods. The low values of LOD and LOQ indicated that the methods are sensitive.

Table 7: Results of Analytical Method Validation

Parameters		Results
		Method I	Method II
Linearity range (mg/ml)		80-180	80-180
Correlation coefficient (R2)		0.9969	0.9974
Slope		0.0015	0.0002
Y-intercept		0.0062	0.0004
% Assay		98.69	99.14
Accuracy (% Recovery)	80%	98.00	98.50
	100%	99.04	98.87
	120%	98.08	99.06
Precision (Mean % R.S.D.)	Repeatability	0.119	0.891
	Intra-day	0.908	0.838
	Inter-day	0.559	0.740
Robustness	270.6 nm	0.686	0.392
	272.6 nm	1.004	0.587
LOD (mg/ml)		8.756	3.222
LOQ (mg/ml)		26.535	9.766

 

CONCLUSION:

It can be concluded from validation results that the proposed methods were simple, sensitive, accurate, precise, robust and economical for the determination of nadolol in tablets. Thus both methods can be applied for routine estimation of nadolol bulk and tablets.

 

ACKNOWLEDGEMENT:

Authors thanks to Principal, Sinhgad College of Pharmacy, Pune for the required facilities and Lupin Research Park, Pune for providing gift sample of the drug.

 

REFERENCES:

1.     Available on: http:// www.drugbank.ca/drugs/DB01203/

2.     United States Pharmacopoeia-29. National Formulary-24. Pharmacopeial forum; Vol-II 2012, pp. 1466-1467.

3.     Vijayalakshmi R, Nagasariramya Y, Dhanaraju MD. Method Development for Quantification of Oxidation Complexes of Nadolol and Resveratrol by Visible Spectrophotometry. International Journal of Pharmacy and Pharmaceutical Sciences. 7(1); 2015: 304-307.

4.     Adegoke OA, Idowu OS, Olaniyi AA. A New Spectrophotometric Method for the Determination of Nadolol. Journal of the Iranian Chemical Society. 3(3); 2006: 277-284.

5.     Camila T, Pedro LG, Fabio P, Erika RM, Maria SR. Quantitative Determination of Nadolol in Tablets by HPLC and UV Derivative Spectrophotometry. Analysis Letter. 41; 2008: 424-36.

6.     Mark PA, Nuggehally SR, Barbhaiya RH, Blair IA. Simultaneous Determination of Nadolol Enantiomers in Human Plasma by High-Performance Liquid Chromatography using Fluorescence-Detection. Biomed Chromatography. 9; 2005:140–45.

7.     Perlman S, Szyper M, Kirschbaum JJ. High-Performance Liquid Chromatographic Analysis of Nadolol and Bendroflumethiazide Combination Tablet Formulations. Journal of Pharmaceutical Science. 73; 1984: 259-61.

8.     Jain HK, Patel KH. Development and Validation of UV spectrophotometric Area under Curve Method for Estimation of Loratadine in Bulk and Tablet Dosage form. American Journal of Pharmtech Research. 3(4); 2013: 492-499.

9.     Sali MS, Barhate AL, Patil VD, Bhadoriya AS, Choudhari VP, Kuchekar BS. Development and Validation of Area under Curve and First Derivative Spectrophotometric Methods for Ropinirole in Tablet Dosage Form. Der Pharma Chemica. 2(3); 2010: 225-229.

10.  Jain HK, Gujar KN, Randhe VA, First Order Derivative Spectrophotometric Method Development and Validation for Midodrine Hydrochloride in Bulk and Tablets, World Journal of Pharmacy and Pharmaceutical Sciences. 5(7); 2016: 1760-1767.

11.  Jain HK, Gupta R, Agrawal RK. Simultaneous estimation of Isoniazid and Pyridoxine Hydrochloride in Tablets by Spectrophotometric Method. Indian Drugs. 41(3); 2004: 153-155.

12.  ICH Q2 (R1), Validation of analytical procedure: text and methodology, International Conference on Harmonization, Geneva, 2005.

 

 

Accepted on 08.09.2016           © RJPT All right reserved

Research J. Pharm. and Tech 2016; 9(10):1694-1698.