A Validated Stability-Indicating and Stereoselective HPLC Method for the Determination of Lenalidomide Enantiomers in Bulk Form and Capsules

Abstract

A simple, rapid and stability-indicating chiral HPLC (CHR-HPLC) method was designed for the enantiomeric separation of lenalidomide (LDM) in the presence of its degradation products. LDM was exposed to different accelerated stress factors. The degradation products were well resolved from the pure drug enantiomers. Separation of the LDM enantiomers was achieved on a LUX 5U cellulose-2 chiral column (250 × 4.6 mm i.d.) with a mobile phase consisting of methanol : glacial acetic acid : triethyl amine (100 : 0.01 : 0.01, v/v/v) at a flow rate of 1.2 mL/min. The detection wavelength was 220 nm, and ornidazole was the internal standard. The chiral method was validated in terms of its specificity, linearity, range, precision and accuracy as well as solution stability, robustness, limit of detection and limit of quantification. The calibration curve was linear for concentrations ranging from 2 to 1,000 ng/mL (r= 0.9999) for both LDM enantiomers. The proposed method, which met International Conference on Harmonization/Food and Drug Administration regulatory requirements, was utilized successfully for the determination of LDM in bulk and in capsules with acceptable accuracy and precision; the label demand percentages were 100.09 ± 0.80 and 99.97 ± 0.93 for the S-(-) and R-(+)-LDM enantiomers, respectively. Based on these results, this method should have great value when applied to quality control and stability studies of LDM.

Figure 1.

Chemical structures of LDM and of the internal standard ORN.

Figure 2.

Typical chromatogram of (a) 500 ng/mL standard LDM and (b) capsule of S-(−)-LDM (1) and R-(+)-LDM (2) spiked with 0.5 µg/mL ornidazole using the proposed CHR-HPLC method.

Figure 3.

Typical chromatogram obtained from stress testing studies of the LDM enantiomers. (a) The standard LDM (500 ng/mL), which was not subjected to stress conditions. (b–f) Samples that were subjected to dry heat, acid hydrolysis, alkaline hydrolysis, UV irradiation and oxidative degradation, respectively.