Molecular Design towards Two-Photon Initiators with Reduced Phototoxicity

Abstract

Two-photon polymerization (2PP) has been recently employed within the biomedical sector, since photopolymers can be crosslinked under relatively mild conditions.1 Crosslinked polymeric networks have been frequently used within the field of tissue engineering to mimic native extracellular matrix, which requires the recreation of complex hierarchical structures. 2PP enables photopolymerization right in the focal point of the applied pulsed laser beam. Micro-fabrication of complex structures in the presence of living tissue is possible, by scanning the focus of the laser beam through a photosensitive formulation. The high transparency of biological tissues towards the near infrared laser allows encapsulation of living cells within intimate cell matrixes with very high spatial resolution in the sub-micrometer range. Two photon initiators (2PIs) are the key components of photoresists, transforming irradiation- into chemical energy by the simultaneous absorption of two photons. During this non linear absorption process 2PI molecules are excited to electronic states, which correspond to the sum of the energy of these two photons. Relaxation processes then lead to the formation of reactive radical species, inducing photopolymerization. In order to crosslink networks with high water content, water-soluble 2PIs are necessary. Several 2PIs have been developed and tested towards their two-photon activity during 2PP. However, phototoxicity remains one of the major issues during cell encapsulation experiments. Not only the 2PI itself can be possibly toxic, but also molecular oxygen can promote cell death by the formation of reactive oxygen species upon quenching of excited 2PIs.2 Therefore, we have focused on molecular 2PI design with highly efficient cleaving mechanisms. Cleavage can be implemented by the introduction of photolabile moieties. Two different types of functional groups have been studied in order to develop new 2PIs. The first is based on diazosulfonates (Figure 1, a) and the second approach focuses on oxime ester functionalities (Figure 1, c). Diazosulfonate 2PIs (DAS) have already been tested towards their two photon activity and performance as low toxic 2PIs in the presence of living cells (Figure 1, b). Metabolic cell-activity as well as long term cell survival have been studied and promising results have been shown compared to state-of-the-art 2PIs.3 Furthermore, oxime ester functionalities have been formerly investigated towards their photoinitiation properties during one-photon polymerization.4 These types of co-initiators have shown promising reaction kinetics during photopolymerization. Consequently, new 2PIs should be developed for the two-photon case by introducing oxime ester functionalities onto two-photon active chromophores. Organo-soluble 2PIs (OE) have already been synthesized and tested towards their two photon absorption properties. Subsequently, water soluble target compounds will be established and compared to common watersoluble 2PIs.5 Future experiments will include cell-culture studies to investigate long-term viability of living cells in combination with newly developed 2PIs.