Rapid fluorescence detection of hypoxic microenvironments by nitro-benzyl conjugated chitosan nanoparticles encapsulating hydrophobic fluorophores

In this study, hypoxia-responsive chitosan nanoparticles (HRCNs) were synthesized by the conjugation of nitro-benzyl derivatives into chitosan polymers and the subsequent self-assembly of them with hydrophobic fluorophores. The nitro-benzyl substrate of HRCNs could be selectively reduced by NTR/NADH that was overexpressed under hypoxic conditions, while it was not responsive to reductive agents (GSH or DTT) commonly existing in biological systems. The HRCN encapsulated with Rhodamine 6G (HRCN-R6G) was successfully applied for the rapid determination of the hypoxic status of lung carcinoma cells (A549) within 30 min, while several hours were required in a conventional assay. In the hypoxic microenvironment, the fluorescence intensity of HRCN-R6G was 2.6-fold higher than that in normoxic conditions. Taken together, the HRCN-R6G is expected to be used for the rapid diagnosis and treatment of hypoxia-related acute diseases, after the incorporation of therapeutic drugs into them.