Efficient Method Development for Separation of Capped mRNA Fragments

User Benefits

- LabSolutions MD can improve the efficiency of method development for 5' capped mRNA fragments and related impurities. - The LCMS-2050 single quadrupole mass spectrometer and the LCMS-9050 quadrupole time-of-flight mass spectrometer enable confirmation of molecular weight. - The LabSolutions Insight Biologics analysis software enables analysis of modifications and impurities specified by the user.

Introduction

There has been increased attention on new drug discovery modalities for mRNA because of its efficacy in COVID-19 vaccines. Currently authorized mRNA vaccines are synthesized using in vitro transcription to add a Cap-0 (m7GpppR-) or Cap-1 structure (m7GpppRm-) on the 5' end. When the cap structure is added by the post-transfer capping method, related impurities (pppR-, ppR-, and GpppR-) are generated by that process. Cap structures contribute to mRNA recognition, efficiency of translation, and stability of mRNA in cells, making 5' cap structure analysis an important element of quality control. For LC separation, one commonly used mode is reversed-phase ion-pair chromatography. The separation patterns obtained with this mode can vary depending on the concentration of the ion-pair reagent and the composition of the organic solvent. In addition, the separation behavior can differ depending on the length of products, nucleobases, and modifications. Therefore, it is important to optimize the separation parameters for each oligonucleotide sequence. This article describes how to efficiently achieve the optimal peak separation for mRNA fragments and related impurities by utilizing LabSolutions MD, which is dedicated software for supporting method development, together with LC/MS identification results.

May 27, 2025 GMT