An analytical study of lipid-oligonucleotide aggregation properties

Area of Science:

• Biophysical Chemistry
• Supramolecular Chemistry
• Analytical Chemistry

Background:

• Lipid-oligonucleotides (LONs) are advanced supramolecular scaffolds for enhanced intracellular nucleic acid delivery.
• Robust analytical characterization of LON assemblies is essential for formulation, in-vivo performance assessment, and quality control.

Purpose of the Study:

• To investigate the utility of size exclusion chromatography (SEC) and capillary electrophoresis (CE) for analyzing LON supramolecular self-assemblies.
• To compare SEC and CE with conventional techniques like gel electrophoresis, dynamic light scattering, and circular dichroism.
• To evaluate the impact of oligonucleotide and lipid modifications on LON assembly.

Main Methods:

• SEC was performed using a TSK-gel G3000PW column at 70°C with water as the mobile phase.
• CE conditions were optimized using a fused-silica capillary at 15°C with various background electrolytes, including a sodium tetraborate buffer with 75mM NaCl.
• Results were correlated with gel electrophoresis, dynamic light scattering, and circular dichroism data.

Main Results:

• SEC successfully separated LON monomers and micelles within 5 minutes.
• Optimized CE conditions provided high resolution and rapid analysis of LON micelles.
• The study confirmed the critical role of lipophilic tails and tetramolecular G-quadruplex formation in LON micelle stability.

Conclusions:

• Capillary electrophoresis (CE) is a highly effective, rapid, and high-resolution technique for analyzing lipid-oligonucleotide (LON) assemblies.
• CE is suitable for assessing LON behavior in biologically relevant media.
• Lipophilic tails and G-quadruplex formation significantly influence the stability of LON micelles.