Chiral HPLC analysis of formoterol stereoisomers and thermodynamic study of their interconversion in aqueous pharmaceutical formulations

A chiral HPLC method was validated and successfully applied for the determination of formoterol stereoisomers and their inversion products in an aqueous matrix stored at 5-70 °C up to 3 weeks. Analysis was performed on a Chiral-AGP column (100 × 4-mm, 5-μm) using a variable mixture of mobile phase A (50-mM sodium phosphate buffer, pH 7.0) and B (10% v/v IPA) at a flow rate of 1.3 ml min−1, and UV detection at 242 nm. All four formoterol stereoisomers were adequately resolved with acceptable detection and quantitation limits varying from 0.01-0.04 μg/ml and 0.04-0.1 μg/ml, respectively. The method showed acceptable accuracy (≥88%), precision (RSD ≤ 8.5%) and good linearity (r2 ≥ 0.9999) over the concentration range investigated. While interconversion at 5 ± 3 °C and 25 ± 2 °C/60% RH ±5% RH was too low to be determined accurately within the study period, chiral inversion of formoterol stereoisomers measured at high temperatures followed the first order rate kinetics and occurred at a single chiral center, resulting in the reversible formation of diastereoisomers, (R,R) ↔ (S,R) and (S,S) ↔ (R,S). No enantiomerization or diastereomerization occurred. There was no significant difference in inversion of the active components in racemic (R,R/S,S)-formoterol fumarate and the single isomer (R,R)-arformoterol tartrate drug formulations, and both drugs are expected to maintain their stereochemical integrity throughout the proposed shelf-life at the recommended storage condition (5 ± 3 °C).