• P-ISSN 0974-6846 E-ISSN 0974-5645

Indian Journal of Science and Technology


Indian Journal of Science and Technology

Year: 2023, Volume: 16, Issue: 46, Pages: 4436-4444

Original Article

RP HPLC-Based Method Development, Validation and Stability Indicating Assay of Levosulpiride in Bulk Drug Using Analytical Quality by Design Approach

Received Date:23 November 2023, Accepted Date:08 December 2023, Published Date:20 December 2023


Objectives: This study aimed at the methodical design and validation of a Reversed-Phase High-Performance Liquid Chromatographic method (RP-HPLC) for the estimation of Levosulpiride in bulk form using analytical quality by design approach. Methods: Using custom design, the Critical Method Parameters were methodically optimized. Peak area, retention time, and peak tailing were assessed for flow rate, buffer, and column length as the influencing elements for Critical Analytical Attributes and using an Agilent C8 column in isocratic elution mode with mobile phase NaH2PO4 buffer and methanol (50:50% v/v) and flow rate at 1ml/min. Findings: Chromatographic separation was accomplished on the Agilent C8 (150×4.6 mm, 5m) column. The optimized and predicted data from JMP PRO 14 software consisted of mobile phase NaH2PO4(50%): Methanol (50%), pumped at a flow rate of 1 ml/min which gave the higher desirability function of 77%. LOD and LOQ values were 0.06 g/ml and 0.20 g/ml respectively and models were found to be significant (p < 0.05). The validation parameter findings were within the permitted range. Forcefully testing the drug's stability under various stress situations revealed considerable degradation in the presence of heat. Novelty: The decrease in the retention and run time shows that the method is simple, accurate, precise and economical for the estimation of Levosulpiride that can be adopted in regular quality control tests in industries.

Keywords: Analytical quality by design, Custom design, Desirability function, Levosulpiride, Reversed Phase Liquid Chromatography


  1. Khan A, Naquvi KJ, Haider MF, Khan MA. Quality by design- newer technique for pharmaceutical product development. Intelligent Pharmacy. 2023;p. 1–8. Available from: https://doi.org/10.1016/j.ipha.2023.10.004
  2. Srujani C, Annapurna P, Nataraj KS, Pawar AKM. Analytical quality by design approach in RP-HPLC method development and validation for the estimation of duvelisib. Asian Journal of Pharmaceutical and Clinical Research. 2021;14(2):99–108. Available from: https://doi.org/10.22159/ajpcr.2021.v14i2.40181
  3. Maheshwari M, Soni P. RP-HPLC Method development and Validation for Rapid estimation of Diazepam in Bulk and Pharmaceutical Dosage Form. Research Journal of Pharmacy and Technology. 2022;15(5):1938–1942. Available from: https://doi.org/10.52711/0974-360X.2022.00322
  4. Santhanam MK, Nagarajan NC, Ponraj PB, Hilurudeen MSM. A Complete Roadmap of Analytical Quality by Design in Various Analytical Techniques. Current Pharmaceutical Analysis. 2023;19(3):184–215. Available from: http://dx.doi.org/10.2174/1573412919666230118105908
  5. Sudhakar B, Akshaya K, Ramya SS. Analytical Method Development and Validation for the Simultaneous Estimation of Bilastine and Montelukast by RP-HPLC. Asian Journal of Research in Pharmaceutical Science. 2023;13(2):79–84. Available from: https://doi.org/10.52711/2231-5659.2023.00015
  6. Samineni R, Chimakurthy J, Konidala SK, Yamarthy V. Development and Validation of Analytical Method for Estimation of Balofloxacin in Bulk and Pharmaceutical Dosage Form by RP-HPLC. Research Journal of Pharmacy and Technology. 2022;15(7):2992–2996. Available from: https://doi.org/10.52711/0974-360X.2022.00499
  7. Hamdache A, Grib L, Grib C, Adour L, Zatout H, Mezrouai A, et al. Forced Degradation Studies of Sofosbuvir with a Developed and Validated RP-HPLC Method as per ICH Guidelines. Chromatographia. 2021;84(12):1131–1140. Available from: https://doi.org/10.1007/s10337-021-04099-8
  8. Marson BM, Concentino V, Junkert AM, Fachi MM, Vilhena RO, Pontarolo R. Validation of analytical methods in a pharmaceutical quality system: An overview focused on HPLC methods. Química Nova. 2020;43(8):1190–1203. Available from: https://doi.org/10.21577/0100-4042.20170589
  9. Patel KK, Patel AM, Patel CN. A new simple RP-HPLC Method development, Validation and Forced degradation studies of Bilastine. Asian Journal of Pharmaceutical Analysis. 2021;11(3):183–187. Available from: http://dx.doi.org/10.52711/2231-5675.2021.00031
  10. Ahmad S, Khairnar M, Bakhshi AR, Tare M, Baheti D, Tare H. QBD approach to develop stability indicating RP-HPLC method development for Levosulpiride and Ilaprazole. International journal of health sciences. 2022;6(S5):7413–7429. Available from: https://doi.org/10.53730/ijhs.v6nS5.11625
  11. Sharma S, Singh N, Ankalgi AD, Rana A, Ashawat MS. Modern Trends in Analytical Techniques for Method Development and Validation of Pharmaceuticals: A Review. Journal of Drug Delivery and Therapeutics. 2021;11(1-s):121–130. Available from: https://doi.org/10.22270/jddt.v11i1-s.4515
  12. Ali AH. High-Performance Liquid Chromatography (HPLC): A review. Annals of Advances in Chemistry. 2022;6:10–20. Available from: https://doi.org/10.29328/journal.aac.1001026
  13. Puranik MP, Mahapatra DK, Soni MA. Analytical quality-by-design (AQBD) approach for the development and validation of RP-HPLC method for the estimation of lamotrigine in bulk and tablet formulation. Journal of Medical pharmaceutical and allied sciences. 2021;10(5):3591–3596. Available from: https://doi.org/10.22270/jmpas.V10I5.1506
  14. Gupta S, Verma P, Mishra AP, Omar N, Mathur R. A Review on Novel Analytical Method Development and Validation by RP-HPLC Method. Indian Journal of Forensic Medicine & Toxicology. 2021;15(4):3479–3486. Available from: https://doi.org/10.37506/ijfmt.v15i4.17798
  15. Bagal D, Nagar A, Joshi A, Chachare A, Shirkhedkar A, Khadse S. Development and validation of stability-indicating RP-HPLC method for estimation of dalfampridine in bulk drug and tablet dosage form. Future Journal of Pharmaceutical Sciences. 2021;7(1):1–7. Available from: https://doi.org/10.1186/s43094-021-00232-4
  16. Pannu S, Thakur M, Gulati P, Kumar M, Dhingra D, Pannu A. A recent review on developed analytical methods for detection of curcumin. International journal of health sciences. 2022;6(S7):173–194. Available from: https://doi.org/10.53730/ijhs.v6n7.10807
  17. Chhalotiya UK. Development of Stability Indicating RP-HPLC Method for Determination of Levosulpiride Hydrochloride In Bulk And Pharmaceutical Dosage Form. International Journal of Advances in Pharmaceutical Analysis. 2012;2(2):41–46. Available from: https://doi.org/10.7439/ijapa.v2i2.18
  18. Tol T, Tawde H, Gorad S, Jagdale A, Kulkarni A, Kasbale A, et al. Optimization of a liquid chromatography method for the analysis of related substances in daclatasvir tablets using design of experiments integrated with the steepest ascent method and Monte Carlo simulation. Journal of Pharmaceutical and Biomedical Analysis. 2020;178:112943. Available from: https://doi.org/10.1016/j.jpba.2019.112943
  19. Pawar SS, Deshmukh DD, Gorde PL, Gosavi SA. Development and validation of RP – HPLC method for quantitation of luliconazole in bulk and formulation. International journal of health sciences. 2022;6(S2):14944–14952. Available from: https://doi.org/10.53730/ijhs.v6nS2.8962


© 2023 Carlota et al. This is an open-access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited. Published By Indian Society for Education and Environment (iSee)


Subscribe now for latest articles and news.