Gelatin microparticles cross-linked with second generation anhydride-containing amphiphilic oligomers – cross-linking characteristics by confocal Raman microscopy

Abstract text (incl. figure legends and references)

Introduction: Gelatin-based microparticles (GM) are versatile biomaterials that can serve as enzymatically degradable templates for microtissue assembly and drug delivery [1]. This study explores using second-generation anhydride-containing oligomers as cross-linkers for fabrication of cross-linked GM (cGM). We aim to assess cross-linking density and intraparticle cross-linking characteristics.

Materials and Methods: We adopted a two-step strategy to fabricate oligomer-cross-linked cGM [2]. Initially, GMs (gelatin type A, 300 Bloom) were fabricated using a surfactant-free water-in-oil emulsion method. Subsequently, post-fabrication chemical cross-linking of GM was performed with self-synthesized anhydride-containing oligomers with different anhydride contents. The comonomers constituting these oligomers are stearyl acrylate, acryloyl morpholine, and maleic anhydride. Laser diffractometry was used to analyze cGM size distribution. A trinitrobenzene sulfonic acid assay was used for analyzing degree of cross-linking (DoC), while confocal Raman microscopy (CRM) of individual GM served to extract spectral data and assess cross-linking characteristics.

Results: Particle size and cross-linking density was controlled by the fabrication protocol. We obtained cGM batches with a median size (Dx50) in a size and DoC range of 45-60 μm and 30-60%, respectively. A correlation between higher DoC and increased oligomer concentration or anhydride content was noted. CRM revealed cross-linking by distinct spectral changes, and distinct spectral changes correlated with higher DoC. CRM data from individual cGM suggested cross-linking through the cross-section of the particle.

Conclusion: This study demonstrates effective cGM fabrication using our second- generation oligomers with varying anhydride contents. Intraparticle cross-linking was determined by CRM.

References:

1. Hinkelmann S, et al. MaterialsTodayBio. 2022;13:100190

2. Loth T, et al. Biomacromolecules. 2014;15:2104-2118