Green RP-HPLC method for estimation of trigonelline: AQbD based development, validation, and application to...

Area of Science:

• Analytical Chemistry
• Green Chemistry
• Pharmaceutical Analysis

Background:

• Trigonelline (TRG) is a bioactive alkaloid with significant pharmacological potential.
• Efficient and reproducible analytical methods are crucial for TRG's drug development, especially in nanoformulations.
• Existing methods may lack the efficiency and environmental sustainability required for modern pharmaceutical analysis.

Purpose of the Study:

• To develop and validate a green reversed-phase high-performance liquid chromatography (RP-HPLC) method for trigonelline (TRG) estimation.
• To integrate Analytical Quality by Design (AQbD) principles with Green Analytical Chemistry for method development.
• To ensure the method's suitability for analyzing TRG in both pure and nano-formulated systems.

Main Methods:

• RP-HPLC method development using a rotatable central composite design for optimization.
• Identification of critical method variables (mobile phase composition, flow rate) and responses (tailing factor, retention time, theoretical plates).
• Validation according to ICH Q2(R1) guidelines and comprehensive greenness assessment using multiple tools (AES, GAPI, AGREE, AMGS, AGSA).

Main Results:

• Optimal separation achieved using ethanol/water (40:60, v/v) on a C18 column, with a sharp peak at 5.60 min and flow rate of 1.5 mL/min.
• The method demonstrated excellent linearity (r² = 0.9986) in the 5-15 µg/mL range, with %RSD < 2%.
• Low limits of detection (LOD) and quantification (LOQ) were 0.628 µg/mL and 1.90 µg/mL, respectively; moderate susceptibility to hydrolysis was observed.

Conclusions:

• The developed green RP-HPLC method is efficient, reproducible, and validated for TRG estimation.
• The integration of AQbD and Green Analytical Chemistry principles resulted in a superior, sustainable analytical method.
• The method is suitable for routine analysis of trigonelline in various pharmaceutical applications, offering enhanced environmental performance.