We further explored if this reaction cycles can drive the generation of transient molecular assemblies. Here we use octanoyl phenylalanine (OA-Phe-OH) as the precursor molecules. When 6.0 equivalents of T3P was added to 5 mM OA-Phe-OH and 5 mM AcCA in Na2CO3 buffer, the reaction system exhibited noticeable turbidity changes, which gradually became clear over time. Next, we monitored the reaction kinetics of the reaction cycles, OA-Phe-AcCA, being hydrophobic, exhibited a peak at 15.5 minutes. As it gradually hydrolyzed over time, it became undetectable by HPLC until 18 hours later (Figure ). The concentration variation curve shown in Figure aligns with the HPLC data, indicating that OA-Phe-AcCA is unstable under alkaline aqueous conditions. Moreover, the concentrations of OA-Phe and AcCA increased steadily over time, corresponding to the hydrolysis of OA-Phe-AcCA via hydrolysis. However, we observed that the amount of AcCA did not fully return to its initial state, resulting in a lower concentration compared to the control experiment. This may be attributed to some side reactions. We further monitored real-time turbidity changes at 600 nm with time as shown in Figure , clearly illustrating the turbidity changes during the reaction process. To further examine the optical characteristics of the OA-Phe-AcCA self-assembly structure, fluorescence microscopy were used to oberve the morphology of the assembly strucures by using 25 µM Nile Red as a dye to monitor the evolution of aggregates formed by OA-Phe-AcCA (Figure ). In the control experiment (without T3P), no higherorder structures are observed under the microscopy. In contrast, when T3P was added, particle-like aggregates were observed first (Figure , and then the structures are converted to long fiber-like structures. At last, with the hydrolysis of thioester, the higherorder assemblies totally disappeared. This indicated the formation of transient molecular assemblies.