Title : Biohybrid cyclophane-based organic semiconductor frameworks for electrochemical acetylcholine sensing toward point-of-care testing
Abstract:
Acetylcholine (ACh) is a vital neurotransmitter involved in numerous physiological processes, and alterations in cholinergic signaling have been associated with neurodegenerative disorders such as Alzheimer’s disease. Current state-of-the-art approaches for assessing cholinergic dysfunction primarily quantify ACh through chromatographic, mass-spectrometric, or enzyme-coupled approaches that can require specialized instrumentation or multistep sample preparation. Therefore, a direct, rapid, and accessible detection method is needed. Here, we utilize electrochemical impedance spectroscopy (EIS) to establish an electrochemical sensing platform composed of a conductive polymer embedded with a biocompatible cyclophane designed to recognize ACh through non-covalent interactions. UV-Vis, Raman, Fourier-transform infrared (FTIR) spectroscopy, and scanning electron microscopy (SEM) were employed to characterize the structural, chemical, and morphological properties of the sensing platform. Within the medically relevant concentration range of 0-2 uM, the ACh response was well described by a Langmuir binding model (R2=0.992), yielding an apparent KD of 55.8 nM and a maximum modelled fractional response of 27.15%. Selectivity was further evaluated against a panel of potentially interfering molecules, including thioflavin T, ascorbic acid, L-cysteine, acetaminophen, histamine, glucose, urea, bovine serum albumin, ibuprofen, immunoglobulin G, and uric acid.
Selectivity testing demonstrated preferential recognition of ACh, which exhibited a normalized sensitivity approximately 229-fold greater than the average response of the tested interferents (0.095). To further investigate the molecular basis of recognition, a 25-ns molecular dynamics simulation was performed for the cyclophane–ACh system. The simulated complex maintained intermolecular distances below a 0.35-nm contact threshold throughout the trajectory, supporting persistent close association between ACh and the cyclophane framework. Together, the electrochemical, selectivity, spectroscopic, microscopic, and computational results establish an effective electrochemical sensing platform for the direct detection of acetylcholine and provide a foundation for the development of rapid, point-of-care diagnostic tools relevant to the study of cholinergic dysfunction and neurodegenerative disease.
