<p>Fmoc-protected phenylalanine based hydrogels have been widely studied in the literature because of their diverse applications, particularly in the biomedical field, owing to their biocompatibility, self-assembly capabilities, and tunable properties. Despite extensive research on their structural and mechanical characteristics, the fluorescence properties of Fmoc-phenylalanine (Fmoc-F) hydrogels remain less discussed. Fluorescence spectroscopy offers a unique approach for monitoring real-time changes in molecular interactions and structural changes within hydrogels. Fluorescence offers several advantages, including high sensitivity and noninvasive detection, enabling a better understanding of gelation. In this study, we examined the fluorescence properties of Fmoc-phenylalanine hydrogels, with a focus on concentration dependent changes and aggregation-induced quenching (AIQ) effects. AIQ is the decrease in fluorescence emission caused by the close aggregation of fluorophores, which facilitates nonradiative decay pathways. Our findings highlight key aspects of the structural changes, stability, thermoreversibility, critical gelation concentration, and lifetime of hydrogels, providing valuable insights into their performance and potential applications. The study also revealed pH-dependent swelling and morphological characteristics. The moderate swelling rate of Fmoc-F is beneficial for controlled and sustained drug release. This makes it suitable for applications where minimal swelling is desired without the need to absorb excessive body fluids, thus enabling prolonged and stable drug delivery. Additionally, the difference in swelling with respect to small changes in pH can be used for applications such as pH-sensitive drug delivery systems.</p> Graphical Abstract <p></p>