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International Journal of Advances in Pharmaceutical Sciences 1 (2010) 375-380 http://www.arjournals.org/ijoaps.

html Research article
ISSN: 0976-1055

Development and validation of UV-Spectrophotometric method for determination of Enalapril maleate
Sunil Kumar Dubey1*, Sandeep Kumar1, Rajeev J Mudakavi1, Swapnil Deshpande1, Akash Kumar Jain1

*Corresponding author: Sunil Kumar Dubey 1 Pharmacy Group, Birla Institute of Technology and Science, Pilani-333 031, India. Email: sunbit2000(at)gmail.com Mobile No: 919887146578

Abstract The UV spectroscopic method of analysis is widely applicable for routine analytical procedure for determination of chemical compounds. Enalapril shows maximum absorbance at a wavelenght of 207nm, which is used in this study. The method provides a linear response from a quantitation range of 2μg/ml to 28μg/ml in phosphate buffer pH7.2. The method gave satisfactory results in terms of repeatability and intermediate precision (RSD<1.58%). Also accuracy values were very good for both methods, the recovery being between 101.37 to 100.11%. The method was validated and proved to be robust and rugged. The results showed that this method can be used for rapid determination of Enalapril maleate in bulk as well as formulation. Keywords:- Enalapril; spectroscopic method; peptidyldipeptidase enalapril is rapidly absorbed, with peak serum concentrations of enalapril occurring within one hour. The extent of absorption of enalapril from oral enalapril tablet is approximately 60%. The absorption of oral enalapril is not influenced by the presence of food in the gastro-intestinal tract. Following absorption, oral enalapril is rapidly and extensively hydrolysed to enalaprilat. Peak serum concentrations of enalaprilat occur about 4 hours after an oral dose of enalapril tablet. The effective half-life for accumulation of enalaprilat following multiple doses of oral enalapril is 11 hours. In subjects with normal renal function, steady-state serum concentrations of enalaprilat were reached after 4 days of treatment [1].

Introduction Enalapril is a prodrug which is hydrolysed in the body to Enalaprilat, which is an inhibitor of angiotensinconverting enzyme (ACE). It is indicated for treatment of hypertension, treatment of symptomatic heart failure and prevention of symptomatic heart failure in patients with asymptomatic left ventricular dysfunction (ejection fraction <35%) [1]. Chemically it is ((S)-1-{N-[1-(ethoxycarbonyl)-3-phenylpropyl]L-alanyl}-L-proline, (Z)-2-butenedioate (1:1), a derivative of two amino-acids, L-alanine and Lproline (fig.1). It is a white to off-white crystalline, odourless powder which melts in the range of 143– 144 ◦C [2]. ACE is a peptidyldipeptidase that catalyzes the conversion of angiotensin I to the vasoconstrictor substance, angiotensin II, which stimulates aldosterone secretion by the adrenal cortex. Blocking the conversion of the angiotensin I to the angiotensin II, leads to a reduction in vasopressor activity and a decrease in peripheral vascular resistance [3, 4]. Oral

Fig. 1 : Structure of Enalapril maleate.

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  Materials and Methods Pure sample of Enalapril maleate was generous gift from Ranbaxy Pvt. The analytical method was also evaluated for the ruggedness and robustness along with its prospective utilization for the assay of analyte in the marketed formulations. Except for conversion to enalaprilat. time-resolved fluorescence (TR-FIA) method [20]. GC.8. and intact enalapril (about 20%).2 solution. . The absorption of test and reference solutions was recorded in 1-cm quartz cells. Precision The precision was evaluated on the basis of repeatability and intermediate precision. The method developed is simple and transferable to the industry for regular quality control. On at least 376 Experimental Instrumentation A double beam Jasco UV-visible spectrophotometer.1 and rest others were of analytical grade. and reproducible method for determination of Enalapril maleate in pharmaceutical preparations by using UV Spectrophotometric method. Method Validation The method’s accuracy.2 solution to obtain concentration of 100μg/ml. there is no evidence for significant metabolism of enalapril. Mortar pestle. Method Development Phosphate buffer pH7. Accuracy The accuracy of the method was checked by recovery determinations and percentage bias. The Phosphate buffer pH7. The determination of enalapril has been carried out by numerous analytical methods such as HPLC [5–12] . International Journal of Advances in Pharmaceutical Sciences 1(2010) 375-380 Enalaprilat binding to human plasma proteins does not exceed 60%.12. Its scanning range was finalized for study and solutions were scanned on spectrophotometer in the UV range of 200 . The determination was done over three concentration levels in triplicate according to the ICH guidelines.5nm. rapid. capillary zone electrophoretic (CZE) method [13]. Gurgaon.22]. Whatman filter paper no.2 was chosen as solvent because it provided superior recovery and precision for the analyte of interest. polarography [15].400 nm.5nm against phosphate buffer pH7. since it does not utilize costly solvents and exacting requirements for the analysis. 1H nuclear magnetic resonance (NMR) spectroscopy [19].2 (USP). middle. precision. atomic absorption spectrometry [16] and membrane selective electrodes [17]. The absorbance intensity was measured at λ of 207. accounting for about 40% of the dose.Dubey et al.20. and high concentrations across the calibration range. Other reagents like Phosphate buffer pH7. Preparation of Standard solutions and calibration The stock solution was prepared by dissolving 10 mg of Enalapril maleate in 100 ml of phosphate buffer pH7. The concentration levels selected as QC samples were 5µg/ml. Ltd. The aim of the present investigation was to develop accurate. specificity.16.Spectrophotometric methods employing derivatisation and formation of charge transfer complexes have also been reported in literature [15. 15µg/ml and 25 µg/ml of enalapril solution. provided by BITS Pilani. model LAMBDA EZ210 connected to computer loaded with PESSW software (Version 1.2 and Revision E) was utilized.4. LC-MS-MS [18]. model V570. The principal components in urine are enalaprilat. The method developed in UV spectroscopy is less expensive compared to other methods like HPLC. 21.28 µg/ml and absorbance measured at the fixed wavelength of 207. The absorbance was plotted against the concentration.24. HPLC-Mass spectrophotometry [14].The spectral bandwidth was 2nm with scanning speed of 2800nm/min. Mass spectroscopy etc. Excretion of enalaprilat is primarily renal. and recovery were demonstrated by a full validation using QC samples prepared at low. UV spectroscopic method for analysis is an extremely versatile and expeditious method for analysis of drug samples. Distilled water.16.2 was evaluated as possible solvent for the assay of enalapril from Bulk and Formulation. Aliquots of the sample solution were taken having dilutions of 2. Measurement of ruggedness was done on Perkin Elmer UV–Vis spectrophotometer.

Robustness & Ruggedness The robustness was determined by changing pH ±0. International Journal of Advances in Pharmaceutical Sciences 1(2010) 375-380 three occasions.2 while ruggedness evaluation was done on Perkin – Elmer UV spectrophotometer. Good linearity was obtained in the concentration range considered.00403 0. According to the ICH requirements [23]. Parameter Wavelength(nm) Concentration Range(µg/ml) Regression equation Intercept(a) Slope(b) Correlation coefficient(r) r2 Limit of Detection(µg/ml) Limit of Quantification(µg/ml) 0.042 0.5 2-28 377 .2.2. LOD and LOQ was determined and reported. The linearity Table1.019 ± 0. Spectrum of enalapril maleate in Phosphate buffer pH 7. Fig 2. middle. The calibration curve was prepared by plotting concentration (in µg/ml) on the abscissa and absorbance in ordinate axis in the range of 228µg/ml.9995 0. The Limit of detection and Limit of quantitation was calculated according to the ICH guidelines.999 0. three replicates of each QC sample pool at low. and high concentrations were assayed.5nm.2. It was observed that it had only one peak at 207. Analytical Parameters for the assay of enalapril using ultraviolet spectrophotometric method in phosphate buffer pH 1.2 207. Results and Discussion Method Development The spectrum of standard solution of enalapril is given in fig. The results are given in table 1.3149 0.9542 Phosphate buffer pH 7. equation was developed by using least square regression analysis. The results obtained were subjected to student t-test to find any significant difference.Dubey et al. Percentage relative standard deviation (%RSD) was calculated for both the methods.

5841 1. while it gave a value of 1.24 25. Evaluation of robustness and ruggedness for the developed methods for the determination of enalapril.68 100.e.5897 0.05. The % bias for LQC was found to be 1.68 0.20 5. The QC samples taken for accuracy determination were similar as for precision study i.2 Accuracy Recovery (%) % bias 101.(Table4). 25µg/ml.4±0. The intraday precision for 5µg/ml was found to be 1. 15.02±0.37 % (Table 2).08±0.38% RSD (Table3).16 25.35 R.D(%) 1.0378 1.05±0.S.08 15.07 15. 15.58%RSD at interday (intermediate precision).11 Phosphate buffer pH 7. QC Concentration (µg/ml) Intraday 5 15 25 Interday 5 15 25 Concentration found(µg/ml) in Phosphate buffer pH 7.2 5 15 25 Accuracy and Precision The precision was carried out at three levels (5.2 Precision Mean ± S. 5.06±0.12±0. QC Concentration (µg/ml) Concentration found(µg/ml) in Phosphate buffer pH 7.37 0. Evaluation of the precision of the developed methods for the determination of enalapril. International Journal of Advances in Pharmaceutical Sciences 1(2010) 375-380 Table2. The accuracy of the method was calculated as percent bias. Table3.37 100. Evaluation of the accuracy of the developed methods for the determination of enalapril. 25 µg/ml).11 1.3861 1.4002 Table4.Dubey et al.D 5.7889 1. Robustness and Ruggedness Robustness and ruggedness of the method was established by applying t-test to compare pH and instrumental variation in which experimental values posed no significant difference at p > 0. 378 .

2 5 99. F = 1. The results obtained are accurate and precise as indicated by the excellent percentage recovery. avisil.6 Applications Assay of dosage forms The applicability of both methods was tested by determination of enalapril in its dosage forms either single or as binary mixture with hydrochlorothiazide. Conclusion The method is suitable for determination of enalapril maleate in bulk form or commercial drug samples.1711 5.0930 15.25 0. International Journal of Advances in Pharmaceutical Sciences 1(2010) 375-380 QC Concentration (µg/ml) Mean ± S.68 0.081±0.S.04) 15 100. Thus the method is applicable for quality control and routine analysis of enalapril in bulk and pharmaceutical preparations. reproducible and sensitive with enough accuracy and precision.91 0. indicating no significance difference between the compared methods regarding accuracy and precision. Pharmaceutical Preparations Concentration found(µg/ml) in Phosphate buffer pH 7.Dubey et al.776 and 19.62 +2. respectively (Table 5).85 101.22 1.44 0.D Robustness: pH 7.83 0.0390 25. non polluting and user friendly. In addition it is rapid and more economical than other methods reported in the literature.62. 23] was performed using the Student’s t-test and the Variance ratio Ftest.84 +2. Theoretical values of t and F at P = 0.9 ± 0.57 (t = 1. The method described is simple.D. Application of the proposed method to the determination of the studied compounds in dosage forms via proposed Method.06 +0.28 +.0928 15. UV spectroscopy offer other advantages of being less expensive.092±0.05 +1. The calculated values did not exceed the theoretical ones.95 101.0568 5.62 102.65±0.06 100. respectively 379 .3741 25.053±0.1010 24.69 1.18 99.15 +1.0963 15.11 Phosphate buffer pH 7.4 5 15 25 pH 7. starch.37 0.09 +2.96±0. Proposed method Reference Method 1.1750 Concentration found(µg/ml) Precision R.00. such as talc. lactose.67 0.83 2.84 102.11 a Mean recovery of three separate determinations.09 102.05 are 2.28 100.18 +0.68 25 100.34±0. gelatin or magnesium stearate did not interfere with the assay.0 5 15 25 Ruggedness 5 15 25 5.03±0.2 Recoverya ± S. Table 5.31±0. Common excipients in tablets.6 % bias +1. Statistical analysis of the results obtained by proposed method and by those given by the reference UV derivative spectrophotometric method [22.D(%) 1.24±0.68 Accuracy Recovery (%) 101.

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