ANALYTICAL METHOD DEVELOPMENT AND VALIDATION OF PERINDOPRIL AND INDAPAMIDE USING RP-HPLC

ANALYTICAL METHOD DEVELOPMENT AND VALIDATION OF PERINDOPRIL AND INDAPAMIDE USING RP-HPLC

Banothu Bhadru * and Dasam Bhagya Lakshmi

C. M. R. College of Pharmacy, Kandlakoya, Medchal, Hyderabad, Telangana, India.

ABSTRACT: A rapid and robust RP-HPLC method was developed and validated for the simultaneous quantification of Perindopril and Indapamide. Chromatographic separation was achieved on a Zodiac C18 column (4.6 × 150 mm, 5 µm) at 38°C using a mobile phase of acetonitrile and water (40:60 v/v) with a flow rate of 1.0 mL/min. Detection was performed at 223 nm, and the injection volume was 10 µL, with a total run time of 10 minutes. The method was validated for accuracy, precision, linearity, and reproducibility. Calibration curves demonstrated excellent linearity with correlation coefficients of 0.9991 for Perindopril and 0.9998 for Indapamide. Accuracy and precision studies showed % purity values of 99.86% for both drugs, confirming the reliability of the method. The developed RP-HPLC method is rapid, precise, and suitable for routine simultaneous estimation of Perindopril and Indapamide in pharmaceutical formulations.

Keywords: RP-HPLC, Perindopril, Indapamide, Simultaneous estimation, Method validation

INTRODUCTION: Perindopril is a non-sulfhydryl angiotensin-converting enzyme (ACE) inhibitor and a pro-drug of Perindoprilat Fig. 1 ¹. Chemically, it is (2S, 3aS, 6aS)-1-[(S)-N-(1-ethoxycarbonyl - 3 - phenylpropyl) - L - alanyl] octahydro-1H-quinazoline-2,4-dione ². It is widely used in the management of hypertension and cardiovascular disorders ³. Indapamide Fig. 2 ⁴ is a non-thiazide indole derivative of chloro-sulphonamide used as an oral antihypertensive and diuretic agent. Chemically, it is 4-chloro-N-(2-methyl-2, 3-dihydro-1H-indol - 1-yl)-3 - sulfamoyl benzamide ⁵. Several spectrophotometric and chromatographic methods have been reported for the estimation of these drugs individually and in combination in bulk and pharmaceutical dosage forms ^6–12.

However, there remains a need for a simple, precise, and reliable analytical method for their simultaneous determination. Therefore, a novel RP-HPLC method was developed and validated for the accurate quantification of Perindopril and Indapamide in bulk and formulations in accordance with ICH Q2 (R1) guidelines ^13–20.

Perindopril:

IUPAC Name: (2S, 3aS, 7aS)-1-[(2S)-2-{[(2S)-1-ethoxy-1-oxopentan-2-yl] amino} propanoyl]-octahydro-1H-indole-2-carboxylic acid).

pKa value: 3.79&5.48

Drug Category: Angiotensin-converting enzyme (ACE) inhibitor. The chemical structure of Perindopril is shown in Fig. 1.

FIG. 1: CHEMICAL STRUCTURE OF PERINDOPRIL

Indapamide:

IUPAC Name: 4-chloro-N-(2-methyl-2, 3-dihydroindol-1-yl)-3-sulfamoylbenzamide

pKa value: 8.8 ± 0.2.

Drug Category: Thiazide-like diuretic. The chemical structure of Indapamide is shown in Fig. 2.

FIG. 2: CHEMICAL STRUCTURE OF INDAPAMIDE

MATERIALS AND METHODS:

Preparation of Mobile Phase: Accurately measured 400 mL (40%) of acetonitrile and 600 mL (60%) of water were mixed, degassed in an ultrasonicator for 15 minutes, and filtered through a 0.45 µm membrane filter under vacuum.

Diluent Preparation: The mobile phase itself was used as the diluent.

Preparation of Standard Solution: Accurately weigh and transfer 10 mg each of Perindopril and Indapamide into a 10 mL volumetric flask, dissolve in 7 mL diluent, sonicate, and make up the volume to mark. From this stock, pipette 1.0 mL of Perindopril and 0.5 mL of Indapamide into another 10 mL volumetric flask and dilute to volume with diluent.

Preparation of Sample Solution: Crush tablets and weigh an amount equivalent to 10 mg each of Perindopril and Indapamide, dissolve in 7 mL of diluent, sonicate, and dilute to volume. Pipette 1.0 mL of this solution into a 10 mL volumetric flask and dilute to the mark.

Procedure: Inject three replicate injections of standard and sample solutions.

Calculate assay using the formula:

%Assay = (Sample Area) / (Standard Area) × (Weight of Standard) / (Dilution of standard) × (Dilution of Sample) / (Weight of Sample) × (Weight of Tablet) / (Label Claim) × Purity / 100 × 100

The percentage purity of Perindopril and Indapamide in the formulation was 100.14% and 99.5%, respectively Table 1.

TABLE 1: ASSAY RESULTS OF PERINDOPRIL AND INDAPAMIDE TABLETS

Chromatographic Conditions:

Column: Zodiac C18 (150 × 4.6 mm, 5μm)

Mobile Phase: Acetonitrile: Water (40:60 % v/v)

Flow rate: 1.0 mL/min

Detection Wavelength: 233 nm

Injection Volume: 10 μL

Column Temperature: Ambient

Runtime: 10 min

Retention Times: Perindopril 2.343 min, Indapamide 3.281 min.

Representative chromatograms of the standard mixture are shown in Fig. 3, indicating clear separation of both analytes.

FIG. 3: REPRESENTATIVE CHROMATOGRAM OF PERINDOPRIL AND INDAPAMIDE

RESULTS AND DISCUSSION:

Precision: The precision of the developed RP-HPLC method was evaluated by performing intra-day and inter-day studies. The %RSD values for intra-day precision were 0.17% for Perindopril and 0.42% for Indapamide, while inter-day precision showed values within 2% for both drugs Table 2. These low %RSD values indicate excellent repeatability and reliability of the method. This confirms that the method provides consistent and reproducible measurements suitable for routine analysis.

TABLES 2: INTRA-DAY AND INTER-DAY PRECISION RESULTS

Accuracy: Accuracy was assessed through recovery studies at 50%, 100%, and 150% levels. The % recovery ranged between 99.83% and 100.34% for both Perindopril and Indapamide Table 3, indicating that the method is highly accurate and free from interference. These results are consistent with literature reports where recoveries for these drugs in combination formulations ranged from 98% to 101%, confirming the reliability of the method for quantitative determination in pharmaceutical dosage forms.

TABLE 3: RECOVERY STUDY RESULTS FOR PERINDOPRIL AND INDAPAMIDE

Linearity: The calibration curves for Perindopril and Indapamide showed excellent linearity in the concentration ranges of 10–30 µg/mL and 30–70 µg/mL, respectively. The correlation coefficients (r²) were 0.999 for both drugs Table 4 & 5, Fig. 4 & 5, demonstrating a strong linear relationship between concentration and peak area. The slopes and intercepts indicate consistent detector response. Linearity across this range confirms that the method is suitable for detecting both low and high concentrations of these drugs in combined formulations.

TABLE 4: LINEARITY DATA FOR PERINDOPRIL

TABLE 5: LINEARITY DATA FOR INDAPAMIDE

FIG. 4: LINEARITY GRAPH OF PERINDOPRIL         

FIG. 5: LINEARITY GRAPH OF INDAPAMIDE

Limit of Detection (LOD) and Limit of Quantization (LOQ): The LOD and LOQ were determined to be 2.68µg/mL and 8.04µg/mL for Perindopril, and 3.54µg/mL and 10.62µg/mL for Indapamide, respectively. These low values indicate the method’s high sensitivity, enabling detection and quantification of even small amounts of the analytes, which is critical for quality control and stability studies

Robustness: The robustness of the developed method was evaluated by introducing small, deliberate variations in chromatographic conditions, including changes in flow rate (±0.1 mL/min) and mobile phase composition (±10%). As presented in Table 6, these variations did not produce any significant changes in retention time, peak area, or system suitability parameters.

The results confirm that the method remains unaffected by minor operational fluctuations, thereby demonstrating its robustness and reliability. This observation is consistent with previously reported HPLC methods for the analysis of Perindopril and Indapamide, which also exhibited stable system suitability parameters under slight method modifications.

TABLE 6: ROBUSTNESS STUDY FOR PERINDOPRIL AND INDAPAMIDE

Specificity: Specificity was confirmed by analyzing placebo solutions and standard drug solutions. No interference from excipients or degradation products was observed at the retention times of Perindopril (2.343 min) and Indapamide (3.281 min), demonstrating that the method is highly selective for both analytes in combined formulations. This ensures accurate estimation of the drugs even in the presence of common tablet excipients, consistent with literature findings.

DISCUSSION: Overall, the developed RP-HPLC method demonstrates high precision, accuracy, linearity, specificity, and robustness, making it suitable for routine quality control of combined Perindopril and Indapamide formulations. Compared to previously reported methods, this method offers a shorter runtime (10 min) and simple mobile phase composition, making it cost-effective and efficient without compromising analytical performance. The method’s sensitivity, with low LOD and LOQ values, further supports its applicability for stability studies and detection of minor impurities or degradation products.

CONCLUSION: A robust, accurate, and precise RP-HPLC method has been successfully developed and validated for the simultaneous estimation of Perindopril and Indapamide in bulk and tablet formulations. The method exhibits excellent linearity, high recovery, and specificity, with assay results consistently near 100% purity. Its simplicity, reproducibility, and robustness make it highly suitable for routine quality control, stability studies, and regulatory compliance in pharmaceutical industries, providing a reliable analytical tool for combined dosage forms.

ACKNOWLEDGEMENTS: Nil

CONFLICTS OF INTEREST: Nil

REFERENCES:

1. The Merck Index. 14th ed. Merck & Co., Inc., Whitehouse Station, NJ, USA 2006; 1238.
2. British Pharmacopoeia. The Stationery Office on behalf of the Medicines and Healthcare Products Regulatory Agency, London 2010; 1: 158–159.
3. Bounhoure JP, Bottineau G and Lechat P: Value of perindopril in the treatment of chronic congestive heart failure: A multicentre double-blind placebo-controlled study. Clin Exp Hypertens A 1989; 11(2): 575–586.
4. Indian Pharmacopoeia. Government of India, Ministry of Health and Family Welfare, Indian Pharmacopoeia Commission, Ghaziabad 2010; 1489.
5. Aleti Rajareddy and Divya A: Stability indicating RP-HPLC method development and validation for simultaneous estimation of ertugliflozin and sitagliptin in bulk and pharmaceutical dosage form. World J Pharm Sci 2021; 9(11): 68–75.
6. Jain PS: RP-HPLC assay method for simultaneous determination of perindopril erbumine and indapamide combination in bulk and tablet dosage form. J Pharm Res 2012; 11(3): 76–80.
7. Siva Sai Kiran B: Stability indicating isocratic RP-HPLC method development and validation for indapamide and perindopril erbumine in pure and combined tablet dosage form. Int J Pharm Sci Res 2015; 6(8): 3428–3438.
8. Mujawar T, Ahmad S and Khatik S: Development and validation of stability indicating RP-HPLC method for estimation of perindopril erbumine and indapamide in bulk and pharmaceutical dosage form. Int J Health Sci 2022; 6(6): 5159–5177.
9. Patel KP: A new RP-HPLC method for simultaneous quantification of Perindopril erbumine, indapamide, and amlodipine besylate in bulk and pharmaceutical dosage form. Future J Pharm Sci 2020; 6: 80.
10. Varma PSR Ch NP: Validated stability indicating RP-HPLC method for simultaneous estimation of perindopril and indapamide in pharmaceutical dosage forms. Int J Pharm 2013; 3(1): 277–289.
11. Bhadru B, Srinath P and Kedarnath: Development, estimation and validation of Aripiprazole in bulk and its pharmaceutical formulation by HPLC method. Int J Chem Tech Res 2012; 4(1): 124–128.
12. Pravalika P, Jephthah G, Raja Reddy A and Rama Rao T: A new RP-HPLC method for estimation of vericiguat in bulk drug and pharmaceutical dosage form. J Adv Sci Res 2023; 14(9): 37–43.
13. Katta S, Boggula N, Bhadru B, Pathakala N and Peddapalli H: Method development and validation of RP-HPLC method for estimation of tolvaptan in bulk and tablet dosage form. Eur Chem Bull 2023; 12(10): 5158–5171.
14. Bhaskara Raju V and Lakshmana Rao A: Development and validation of new HPLC method for estimation of perindopril in tablet dosage forms. Rasayan J Chem 2011; 4(1): 113–116.
15. Padol SG and Waghmare SA: Development and validation of RP-HPLC method for estimation of indapamide: A review. Int J Pharm Sci 2024; 2(5): 1588–1597.
16. Pannu HK, Mahajan MP and Sawant SD: Validated RP-HPLC method for determination of indapamide in bulk and tablet dosage form. Scholars Res Lib 2012; 4(3): 996–1002.
17. Bhadru B, Venkata Rao V and Boggula N: Simultaneous estimation of levodopa and benserazide using RP-HPLC in API and marketed formulation. Int J Recent Adv Multidiscip Res 2023; 10(10): 9020–9024.
18. Bhadru B, Bhagya Lakshmi D and Rama Rao T: Analytical method development and validation for simultaneous estimation of perindopril and indapamide in bulk and pharmaceutical dosage form by RP-HPLC. Int J Res Innov Appl Sci 2025; 10(6): 122–126.
19. Venkatesh A, Raja Reddy A and Ramarao T: A review on method development and validation of atenolol by spectrophotometric and HPLC techniques. World J Pharm Pharm Sci 2022; 11(3): 593–607.
20. Raja Reddy A and Kavya Reddy H: Analytical RP-HPLC method development and validation for simultaneous estimation of bilastine and montelukast in bulk and pharmaceutical dosage form. Ymer 2022; 21(11): 1504-16.
    

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    How to cite this article:

    Bhadru B and Lakshmi DB: Analytical method development and validation of perindopril and indapamide by using RP-HPLC. Int J Pharm Sci & Res 2026; 17(5): 1561-65. doi: 10.13040/IJPSR.0975-8232.17(5).1561-65.

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