Development of a Stability‐Indicating RP‐HPLC Method for Pioglitazone in Cubosomal and Biological Matrices: A Quality by Design‐Driven, Lean Six Sigma, and Green Chemistry Approach
Vaibhavi D. Torgal, Vinayak Mastiholimath, Rahul Koli
Abstract
ABSTRACT A robust, precise, and environmentally sustainable RP‐HPLC method was developed and optimized via a quality by design (QbD) framework for the quantification of pioglitazone HCl in cubosomal formulations and biological matrices. Method development using Box–Behnken design evaluated three critical parameters—ACN concentration (20%–40% v/v), flow rate (0.9–1.1 mL/min), and column temperature (25°C–35°C)—to optimize analytical performance. The optimal conditions included a mobile phase of ACN and 0.1% formic acid (30:70 v/v), a flow rate of 1.0 mL/min, and a column temperature of 30°C using a Phenomenex Luna C18 (250 × 4.6 mm, 5 µm) column. The method exhibited excellent linearity across the 2 to 10 µg/mL range ( R 2 = 0.9999), with LOD and LOQ of 0.086 and 0.26 µg/mL, respectively. Intra‐ and inter‐day precision (%RSD < 2%) and accuracy with good recovery met ICH Q2 (R1) guidelines. Forced degradation studies confirmed specificity and stability‐indicating capability. Recovery from plasma and cubosomal matrices demonstrated high efficiency (recoveries: 95.3%–101.05%). The method's environmental sustainability was validated using several tools, while Six Sigma evaluation revealed a Cpk > 1.33. This validated RP‐HPLC method offers a green, high‐performance analytical platform for pioglitazone HCl in complex matrices.