# Encapsulation of mRNA in Therapeutics Like Lipid Nanoparticles Probed by Deep-UV Resonance Raman Spectroscopy

## TL;DR

This study uses deep-UV resonance Raman spectroscopy to analyze how mRNA is encapsulated in lipid nanoparticles for therapeutic delivery.

## Contribution

The first use of 2T2D-COS for quantitative analysis of lipid–mRNA interactions in mRNA therapeutics.

## Key findings

- DUVRR spectroscopy can distinguish between fully encapsulated, partially exposed, and free mRNA.
- PCA identified spectral variations between 1270 and 1800 cm–1 linked to lipid presence.
- The 1322 cm–1 band was identified as a quantitative indicator of lipid–mRNA interactions.

## Abstract

Messenger RNA (mRNA) therapeutics have emerged as a powerful
platform
for treating a wide range of diseases, but their clinical success
depends on overcoming the issues of instability and delivery. mRNAs
are typically unstable and require encapsulation in lipid nanoparticles
(LNPs) for efficient delivery. This study quantifies free and encapsulated
mRNAs within LNPs in a model vaccine using deep-UV resonance Raman
(DUVRR) spectroscopy. DUVRR spectroscopy with excitation at 266 nm,
matching the maximum UV absorption of mRNA, enables spectral differentiation
of mRNA based on its degree of encapsulationfrom fully encapsulated
to partially exposed to completely free. Raman spectra were acquired
from the samples with progressively decreasing lipid concentrations
while maintaining a fixed mRNA concentration; vibrational modes associated
with mRNA–lipid interactions were identified. Principal component
analysis (PCA) revealed spectral variations linked to the lipid presence,
especially between 1270 and 1800 cm–1. Two-trace
two-dimensional correlation spectroscopy (2T2D-COS) was employed to
extract the 1322 cm–1 band as a quantitative spectroscopic
indicator of the lipid–mRNA interaction. This is the first
use of 2T2D-COS for quantitative analysis of complex (bio)­chemical
systems. The work highlights the unsurpassed potential of DUVRR spectroscopy
to analyze mRNAs within LNPs.