Quality by Design Approach in the Development of a Stability-Indicating RP-HPLC Technique for Levodopa and Benserazide

Lakshmana Rao  Atmakuri; Rajani  Vetapalem; Anupama  Baratam; Ramesh  Alluri; Satya Venkata Sakuntala  Mamidi; Apurva  Somala

doi:10.5530/ctbp.2026.2s.3

Authors

Lakshmana Rao Atmakuri V. V. Institute of Pharmaceutical Sciences, Gudlavalleru – 521 356, Andhra Pradesh
Rajani Vetapalem V. V. Institute of Pharmaceutical Sciences, Gudlavalleru – 521 356, Andhra Pradesh
Anupama Baratam KVSR Siddhartha College of Pharmaceutical Sciences, Vijayawada – 520 010, Andhra Pradesh
Ramesh Alluri Vishnu Institute of Pharmaceutical Education & Research, Narsapur – 502 313, Telangana
Satya Venkata Sakuntala Mamidi Department of Pharmacy, Government Polytechnic for Women, Kakinada – 533 003, Andhra Pradesh
Apurva Somala V. V. Institute of Pharmaceutical Sciences, Gudlavalleru – 521 356, Andhra Pradesh

DOI:

https://doi.org/10.5530/ctbp.2026.2s.3

Keywords:

Levodopa, Benserazide, RPHPLC, Quality by Design, Stability-Indicating Method, Method Validation

Abstract

The study demonstrates development & validation of stability-indicating RP-HPLC technique for simultaneous estimation of Levodopa & Benserazide within a Quality by Design (QbD) framework. Critical method parameters (CMPs) - composition of movable phase, rate of flow, & temperature of column were systematically optimized to achieve predefined critical quality attributes (CQAs), including resolution, tailing factor, and theoretical plates. Separation of compounds in chromatography was done on HPLC system of Waters Alliance make equipped with quaternary gradient pump & detection achieved with UV at 228 nm, with C18 column. The optimized mobile phase consisted of acetonitrile & 0.1% triethylamine buffer (pH 2.5, attuned with orthophosphoric acid) in 46.5:53.5 v/v ratio, delivered at rate of flow at 1.0 mL/min. Retention times for Levodopa & Benserazide were 3.422 & 5.330 minutes, correspondingly. Stress studies under various conditions established, method’s stability-indicating capability. Validation performed in accordance with guidelines of ICH Q2(R1) demonstrated excellent linearity (R² > 0.999), accuracy (98-102%), precision (RSD < 2%), robustness, & sensitivity. The integration of QbD principles enhanced method robustness, defined a regulatory - compliant design space, and ensured lifecycle reliability. Developed RP-HPLC technique is therefore reliable & versatile tool for routine quality control of Levodopa & Benserazide in formulations.