A Study on Synthesis and Upscaling of 2′-O-AECM-5-methyl Pyrimidine Phosphoramidites for Oligonucleotide Synthesis

Abstract

2′-O-(N-(Aminoethyl)carbamoyl)methyl-modified 5-methyluridine (AECM-MeU) and 5-methylcytidine (AECM-MeC) phosphoramidites are reported for the first time and prepared in multigram quantities. The syntheses of AECM-MeU and AECM-MeC nucleosides are designed for larger scales (approx. 20 g up until phosphoramidite preparation steps) using low-cost reagents and minimizing chromatographic purifications. Several steps were screened for best conditions, focusing on the most crucial steps such as N3 and/or 2′-OH alkylations, which were improved for larger scale synthesis using phase transfer catalysis (PTC). Moreover, the need of chromatographic purifications was substantially reduced by employing one-pot synthesis and improved work-up strategies.

THF peroxide as a factor in generating desulphurised products from the solid-phase synthesis of phosphorothioate-modified oligonucleotides

“…In contrast, THF is sometimes used as a reaction solvent for the phosphitylation of chemically modified nucleic acids. 37–41 The above experimental results indicate that phosphoramidites may be oxidised by THF peroxide even during their synthesis, leading to yield loss. In addition, common ethers, such as diethyl ether, which are used for the extraction and recrystallisation of organic compounds, are also known to contain peroxides, 42 and care should be taken with the solvents used in the purification of phosphoramidite after the reaction.…”