Impurity profiling of PEGylated myristoyl diglyceride, DMG-PEG 2000, a functional excipient used in mRNA lipid nanoparticle formulations

Abstract

Lipid nanoparticles have gained significant attention during the COVID-19 pandemic, particularly due to their role in mRNA vaccine delivery. However, their rapid advancement has outpaced the development of established harmonized protocols for the quality control of the various excipients. In this study, we focused on the “stealth” lipopolymer 1,2-dimyristoyl-rac-glycero-3-methoxypolyethylene glycol-2000 (DMG-PEG 2000), a critical excipient used in Moderna's Spikevax® mRNA vaccine. We investigated different commercial batches of DMG-PEG 2000 for impurities originating from both synthesis and degradation. Synthesis-related impurities include free glycerol and fatty acids of varying chain lengths, while degradation products result from single or double hydrolysis reactions. These synthetic and degradation-related impurities were primarily analyzed using an optimized high-performance liquid chromatography method with a charged aerosol detector (HPLC-CAD). Applying this validated method, a high purity of commercially available DMG-PEG 2000 was revealed, with every batch investigated exceeding a purity of 98.5%. In addition, gas chromatography (GC), HPLC with an evaporative light scattering detector (HPLC-ELSD), and matrix-assisted laser desorption/ionization time-of-flight (MALDI-TOF) mass spectrometry were employed for comparative purposes and to enable further characterization. Based on these analyses, we developed a streamlined and robust impurity profiling protocol that (i) provides essential insight into the impurity profile of DMG-PEG 2000 in marketed products and (ii) may facilitate more decentralized and standardized validation processes in the future. The analytical approach presented here may also serve as a foundation for a future pharmacopeial monograph proposal for DMG-PEG 2000.