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Aggregation by DLS

Inserting a sample into a Dynamic Light Scattering Instrument.

Background

With the recent boom of complex API based therapeutics coming to market, the Food and Drug Agency (FDA) has recently issued a guidance for Synthetic Peptides and a draft guidance regarding Sameness Evaluations of Complex APIs. Within these guidances, the agency recommends that “the applicant applies orthogonal analytical methods to characterize the Oligomer/Aggregation States.” Oligomer/Aggregation States can be characterized by a variety of orthogonal methods: Size Exclusion Chromatography (SEC-UV-MS), Dynamic Light Scattering (DLS), and Diffusion Ordered Spectroscopy (DOSY).

Recent advances in the field have revolved around the comparison of DLS and DOSY. One publication, by scientists at the Agency, suggests that DLS is more suitable than DOSY to determine whether aggregates that could cause immunogenicity are present in a sample. This is due to the higher sensitivity DLS has for evaluating higher molecular weight compounds.

Novatia as a DLS Research Partner

Novatia has experience in applying DLS and its orthogonal methods for characterizing aggregation states of peptides. Shown below is an example of a DLS autocorrelation function for a lot of Ozempic. In addition to the translational diffusion coefficient (Dt), DLS evaluates the hydrodynamic radius (Rh), %Mass, and %Polydispersity (%Pd) for each autocorrelation. Furthermore, the %Pd characterizes the oligomeric makeup of a specific signal. Such as, whether the species is present as a single conformer or whether there are other unresolved oligomers present. In the case below, a %Pd between 15-30% corresponds to a species that consists of a combination of mainly monomer with a small population of oligomers.

Figure 1: Autocorrelation function of a lot of Ozempic. Autocorrelation 2 was determined to be both an effect of the formulation buffer and insignificant due to a %Mass < 0.1%.

Novatia has assisted research organizations with aggregation characterization for regulatory filing of complex APIs. We offer combined services with our MS team and NMR Team for further characterization. Contact us to see how our team can assist your aggregation characterization needs.