CHEMOMETRIC EVALUATION OF DARIFENACIN HYDROBROMIDE USING A STABILITY-INDICATING REVERSED-PHASE LC METHOD

Abstract

A set of chemometric tools including in silico simulation and design of experiments (DOE) was used to develop and validate an RP-LC/UV method for darifenacin hydrobromide. Retention factor (k′) from darifenacin and resolution (R) between darifenacin and degradation product were assessed by an exploratory study using organic modifier type, organic modifier content, mobile phase pH, flow rate, and column oven temperature as factors in a full factorial 2⁵ design. The significant factors (organic modifier content, mobile phase pH, and column oven temperature) were selected and applied in a central composite design (CCD) with axial points positioned in α = 1.681. The optimum condition was obtained with Derringer's desirability. The validation was performed with a C₈ (150 mm × 4.6 mm, 5 µm) column, wavelength 205 nm, 1.1 mL · min⁻¹ flow rate, mobile phase with acetonitrile (MeCN) and orthophosphoric acid (OPA) 0.01% (v/v) pH 3.0 in a ratio 28:72 and column maintained at room temperature (25 ± 1°C). Forced decomposition was performed in agreement with the guidelines and collected data were confirmed with LC-MS/MS. The obtained k′ and R were 3.22 and 2.62, respectively. Additionally a Plackett-Burman design was done to study robustness, and the results agreed with the guideline requirements for a stability-indicating method.

Keywords:

• darifenacin hydrobromide
• Derringer's desirability
• design of experiments
• high performance liquid chromatography
• stability-indicating

ACKNOWLEDGMENTS

The authors thank CAPES and CNPq of Brazil, LANAGRO (Brazil) and Labex Inc. (USA).

Notes

*(−1) and (+1) are “low” and “high” levels.

*(−1), (+1), (0) are “low”, “high” and nominal levels, respectively.

^a (−1), (+1), (0) are “low”, “high” and nominal levels, respectively.

^b Acetonitrile content (%) in mobile phase.

^c pH of the mobile phase.

^d Column oven temperature in degree celsius.

^a Relative standard deviation.

^b Retention time.

^c Tail.

^d Theorical plates.

^e Resolution.

^f Concentration in ng mL⁻¹.

^g Confidence limit.

^h Standard error.

^a Standard error.

^b N°1: Day 1, Analyst 1, Instrument 1; N°2: Day 2, Analyst 2, Instrument 2; N°3: Day 3, Analyst 1, Instrument 2.