The term “oligonucleotides” is broad and encompasses a range of chemical entities from low molecular weight siRNA (15-20 nucleotides) to mRNA and DNA expressing whole genes containing many thousands of nucleotides. In addition, chemical modifications and formulations necessary for the functionality of these therapeutics add a further layer of complexity and potential risk and increase the analytical challenge.
BioPharmaSpec provides specialized, high resolution mass spectrometric analysis of oligonucleotide products to confirm identity, and assess for impurities and degradants. Our scientists use their extensive structural characterization experience to apply many of the below techniques to your oligonucleotide as required to confirm the desired product from simple molecular weight determination through to detailed structural sequence and impurity analysis. The selection of appropriate analytical techniques depends on the oligonucleotide size, structure, chemical composition and modifications. Various chromatographic, electrophoretic and high performance liquid chromatography mass spectrometry techniques are used for data acquisition to support identification and purity assessments of DNA, RNA diagnostic oligonucleotides and oligotherapeutics:
- Molecular weight confirmation using ESI-MS, LC-MS and/or MALDI-TOF
- Detailed structural analysis using LC-MS
- Confirmation of sequence using LC- MS/MS and/or exonuclease treatment followed by MALDI-TOF MS analysis
- Impurity profiling and identification using LC-MS
- pI/charge variants using icIEF/IEX
1. M. Sapru, “CMC Regulatory Considerations for Oligonucleotide Drug Products: FDA Perspective,” (2017). https://pqri.org/wp-content/uploads/2017/02/3-SapruPQRI-FDA-Conference-Oligo-2017-Presentation.pdf